title: Histone methyltransferase Ezh2 coordinates mammalian axon regeneration via epigenetic regulation of key regenerative pathways

abstract: Current clinical treatment for neurodegenerative diseases and neural injuries falls short of success, and one primary reason is that neurons in the mammalian central nervous system (CNS) lose their regeneration ability as they mature. Previous studies indicated that the regeneration ability of neurons is governed by complex signaling networks involving many genes. Therefore, here we investigated the roles of Ezh2, a histone methyltransferase, in regulation of mammalian axon regeneration at the epigenetic level. We found that Ezh2 level was gradually downregulated in the mouse nervous system during maturation but significantly upregulated in mature sensory neurons during spontaneous axon regeneration in the peripheral nerve system (PNS), suggesting its role in supporting axon regeneration. Indeed, Ezh2 loss-of-function in sensory neurons impaired PNS axon regeneration in vitro and in vivo. In contrast, overexpression of Ezh2 in retinal ganglion cells in the CNS induced optic nerve regeneration after optic nerve injury in both methyltransferase-dependent and -independent manners. Mechanistic exploration with multiomics sequencing, together with functional analyses, revealed that Ezh2 supported axon regeneration by systematically silencing the transcription of genes regulating synaptic function and axon regeneration inhibitory signaling, while broadly activating factors promoting axon regeneration. Our study not only reveals that Ezh2 coordinates axon regeneration via epigenetically regulating multiple key regenerative pathways, but also suggests that modulating chromatin accessibility is a promising strategy to promote CNS axon regeneration.

category: neuroscience


title: Enhancing Polysaccharide Immunity through Intranasal Immunizations

abstract:
ObjectiveA model conjugate vaccine incorporating a pneumococcal polysaccharide with EcoCRM197 was used to determine vaccine efficacy when administered with our clinical-stage mucosal stimulating adjuvant, NE01. This was the first attempt at combining NE01 with a polysaccharide conjugate vaccine to elicit mucosal immunity against respiratory bacterial infection through intranasal immunizations.

ResultsIntranasal immunizations using the NE01 adjuvant incorporating a polysaccharide conjugate resulted in the generation of a robust IgG and IgA responses in both serum and mucosa. Our data also demonstrated the active homing of immunological memory cells to the lower respiratory tract as evidenced by an increase of IgG- and IgA-producing memory B-cells in the lungs. Further, intranasal immunization enhanced the induction of a balanced Th1/Th2/ Th17 immune response with clear homing of memory T-cells to the lungs. Serum antibodies generated by this formulation and route of administration demonstrated excellent efficacy in killing S. pneumonia in an in vitro OPK assay, a biomarker of vaccine efficacy. Our data suggest that further optimization of the dose and schedule as well as evaluation of the efficacy of this new formulation in animal models of colonization and infection are warranted.

category: immunology


title: The transcriptional regulator Sin3A balances IL-17A and Foxp3 expression in primary CD4 T cells

abstract:
The Sin3 transcriptional regulator homolog A (Sin3A) is the core member of a multi-protein chromatin-modifying complex known to control gene transcription via epigenetic mechanisms. Its inactivation in developing thymocytes halts T cell maturation. We and others had previously shown that Sin3A controls STAT3 transcriptional activity. Given the role of STAT3 in the differentiation of T helper 17 cells critical in inflammatory disorders and against opportunistic infections, we asked whether Sin3A could also contribute to their differentiation. To this aim, we exploited CD4-Cre and CD4-CreERT2deleter strains for conditional and inducible Sin3A deletion in CD4 cell subsets. We report that Sin3A inactivation in vivo arrested thymocyte development at the double positive stage, hindering the characterization of mature T cells. At difference, tamoxifen-inducible Sin3A deletion proved permissive for in vitro proliferation of T cells in Th17 skewing conditions and the acquisition of memory markers. Transcriptional profiling indicated that while Sin3A inactivation imprinted T cells with a mTORC1 signaling gene signature, Sin3A deficient cells lacked the expression of IL-17A, the signature Th17 cytokine. This reflected a defective induction of Il17a, and also of the Il23R and Il22 genes, which occurred in spite of proper upregulation of the lineage defining transcription factor ROR{gamma}t. We found that Sin3A inactivation was paralleled by increased STAT3 phosphorylation and nuclear representation, and by higher fractions of IL-2 and FoxP3 expressing cells. Such events proved causally linked as inhibiting Foxp3 partially rescued IL-17A expression, and neutralizing IL-2 simultaneously lowered the representation of FoxP3+cells, while rescuing IL- 17A+ ones. Thus, together our data underline a previously unappreciated role for Sin3A in Th17 differentiation and the shaping of their immunoregulatory potential.

StatementThis study identifies a new role for the transcriptional regulator Sin3A in the shaping of Th17 cell differentiation. Data indicate that by controlling IL-2 expression, and mTORC1 signaling, it balances IL-17A and Foxp3 levels, shaping Th17 inflammatory potentials.


title: Intercellular collectivity is governed by enzyme secretion strategies in marine polysaccharide degrading bacteria

abstract: Polysaccharide breakdown by bacteria requires the activity of enzymes that degrade polymers extracellularly. This generates a localized pool of breakdown products that are accessible to the enzyme producers themselves as well as to other organisms. Marine bacterial taxa often show marked differences in the production and secretion of degradative enzymes that break down polysaccharides. These differences can have profound effects on the pool of diffusible breakdown products and hence on the ecological dynamics. However, the consequences of differences in enzymatic secretions on cellular growth dynamics and interactions are unclear. Here we combine experiments and models to study the growth dynamics of single cells within populations of marine Vibrionaceae strains that grow on the abundant marine polymer alginate, using microfluidics coupled to quantitative single-cell analysis and mathematical modelling. We find that strains that have low extracellular secretions of alginate lyases show stronger aggregative behaviors compared to strains that secrete high levels of enzymes. One plausible reason for this observation is that low secretors require a higher cellular density to achieve maximal growth rates in comparison with high secretors. Our findings indicate that increased aggregation increases intercellular synergy amongst cells of low-secreting strains. By mathematically modelling the impact of the level of degradative enzyme secretion on the rate of oligomer loss to diffusion, we find that enzymatic capability modulates the propensity of cells within clonal populations to cooperate or compete with each other. Our experiments and models demonstrate that marine bacteria display distinct aggregative behaviors and intercellular interactions based on their enzymatic secretion capabilities when growing on polysaccharides.

category: microbiology


title: Functional characterization of bacterial isolates from dye decolorizing consortia and a step-up metabolic engineering based on NADH-regeneration

abstract: Azo dye decolorizing acclimatized decolorizing consortia are enriched microbial sources of potential azoreductase-efficient bioremediation strains. Here, we characterized eight selected consortial members for their azo decolorization and azoreductase profiling. These efficient dye decolorizing bacterial isolates were affiliated to two major phyla viz. Firmicute (genus-Enterococcus) and Proteobacteria ({gamma}-group). Redox-mediators such as AQDS and AQS were found to significantly increase decolorization except for menadione, and IR functional group signatures highlighted the azo bond reduction and degraded metabolites profiles of each strain. Among isolates, Enterococcus sp. L2 was found to be the most effective strain as it could reduce >90mg/L Reactive violet 5R (RV5R) dye in 3h of incubation. Furthermore, strain L2 possesses profound high NADH and NADPH-dependent azoreductase activity which also corroborated with its superior azo decolorization. As per physicochemical parameters, strain L2 showed an optimum decolorization at pH 8, 40 {degrees}C and up to 2% w/v salinity. To channelize reducing equivalence (NADH) to further enhance the dye decolorization in NADH-azoreductase efficient Enterococcus sp. L2, we augmented an NADH co-factor regeneration system. Using pMGS100, a Gram-positive expression vector a constitutive heterologous expression of Mycobacterium vaccae encoded NAD+-dependent formate dehydrogenase enhanced NADH pool which led to a significant 3.2 fold increased dye decolorization in Enterococcus sp. L2 harboring pMGS100 fdh along with a positive effect on growth. Ultimately, an augmentation of formate utilization step could further accelerate azo dye decolorization by fulfilling the co-factor (NADH) requirement of azoreductase along with a growth advantage in the non-model azoreductase-efficient environmentally important strain L2.


title: Nonstructural protein 1 (nsp1) widespread RNA decay phenotype varies among Coronaviruses

abstract:
AO_SCPLOWBSTRACTC_SCPLOWExtensive remodeling of the host gene expression environment by coronaviruses nsp1 proteins is a well-documented and conserved piece of the coronavirus-host takeover battle. However, whether and how the underlying mechanism of regulation or the transcriptional target landscape differ amongst coronaviruses remains mostly uncharacterized. In this study we use comparative transcriptomics to investigate the diversity of transcriptional targets between four different coronavirus nsp1 proteins (from MERS, SARS1, SARS2 and 229E). In parallel, we performed Affinity Purification followed by Mass-Spectrometry to identify common and divergent interactors between these different nsp1. For all four nsp1 tested, we detected widespread RNA destabilization, confirming that both - and {beta}-Coronavirus nsp1 broadly affect the host transcriptome. Surprisingly, we observed that even closely related nsp1 showed little similarities in the clustering of genes targeted. Additionally, we show that the RNA targeted by nsp1 from the -CoV 229E partially overlapped with MERS nsp1 targets. Given MERS nsp1 preferential targeting of nuclear transcripts, these results may indicate that these nsp1 proteins share a similar targeting mechanism. Finally, we show that the interactome of these nsp1 proteins differ widely. Intriguingly, our data indicate that the 229E nsp1, which is the smallest of the nsp1 proteins tested here, interacts with the most host proteins, while MERS nsp1 only engaged with a few host proteins. Collectively, our work highlights that while nsp1 is a rather well-conserved protein with conserved functions across different coronaviruses, its precise effects on the host cell is virus specific.

SO_SCPLOWIGNIFICANCEC_SCPLOWCoronaviruses extensively co-opt their host gene expression machinery in order to quicky benefit from the host resources. The viral protein nsp1 plays a major role in this takeover as nsp1 is known to induce a widespread shutdown of the host gene expression, both at the RNA and the translational level. Previous work characterized the molecular basis for nsp1-mediated host shutdown. However, this was mostly conducted in the context of {beta}-coronaviruses and in particular SARS-CoV1, CoV2 and MERS due to the important public health burden that these viruses represent. Here instead, we explored the impact of nsp1 on the host using a comparative approach, defining the influence of 4 nsp1 protein from - and {beta}-coronaviruses. We delineated the impact of these 4 nsp1 on the host transcriptome and mapped their interactome. We revealed that host target range and interactomes vary widely among different nsp1, suggesting a viral-specific targeting. Understanding how these differences shape infection will be important to better inform antiviral drug development.


title: eIf2α-regulated translation modulates the early adhesion of mesenchymal-like cells

abstract:
Cellular invasion is a complex process that requires several interdependent biological mechanisms, which are initiated by changes in adhesion that establish a morphology favorable for migration. Hence, the regulation of adhesion potential is a rate-limiting step in metastasis. Our previous work revealed that de novo translation is necessary to regulate the adhesion of mesenchymal-like cells; however, the underlying translational regulatory mechanism and the identity of newly synthesized proteins needed for the adhesion process remain unidentified. Here, we describe a translational regulatory mechanism that modulates mesenchymal cell adhesion. We observed a drastic decrease in translation during the initial phase of adhesion, followed by a reprogramming of the translatome, characterized by an orchestrated wave of mRNA translation that increases the expression of specific proteins involved in adhesion. We observed that phosphorylation of eukaryotic translation initiation factor 2 subunit alpha (eIF2), which inhibits general translation initiation, was drastically increased at the beginning of cell adhesion. As adhesion progressed, the selective increase in the translation of adhesion-related mRNAs intensified as eIF2 phosphorylation gradually decreased over time in mensenchymal cells, but not in epithelial cells. Taken together, we have identified a translational regulatory mechanism specifically affecting the adhesion process of mesenchymal cells, as well as metastatic cells that have undergone epithelial-to-mesenchymal transition.

One sentence summaryTranslation regulates mesenchymal cell adhesion

category: cell biology


title: Cystin is required for maintaining fibrocystin (FPC) levels and safeguarding proteome integrity in mouse renal epithelial cells Running title: Cystin deficiency reduces FPC levels through enhanced autophagic degradation.

abstract: Autosomal recessive polycystic kidney disease (ARPKD) is primarily caused by mutations in PKHD1, encoding fibrocystin (FPC). Recently, patients with CYS1 mutations, encoding cystin, have also been identified. While Pkhd1 mutant mice do not exhibit ARPKD-like phenotype, cystin deficient Cys1cpk/cpk (cpk) mice do. We explored potential functional interactions between FPC and cystin in mouse kidneys and cortical collecting duct (CCD) cell lines. FPC levels were markedly reduced in both cpk kidneys and CCD cells through selective autophagy. Re-expression of Cys1 in cpk kidneys restored and siRNA silencing of Cys1 in wild type-CCD cells reduced FPC levels. In contrast, FPC deficiency in Pkhd1 mutant kidneys did not diminish cystin levels. Consistent with the previously identified role of FPC in E3 ubiquitin ligase complexes (NEDD4), we observed reduced polyubiquitination with elevated epithelial sodium channel (NEDD4 target) levels and function in cpk-CCD cells. Our results imply that cystin stabilizes FPC in E3-ligase complexes and helps to maintain proteome integrity. In cpk mice, cystin loss causes increased MYC expression and also leads to severe FPC reduction and altered proteome. Together these perturbations promote cystogenesis. We speculate that in Pkhd1 mutants FPC loss alone is insufficient for kidney cyst formation, because cystin prevents c-Myc overexpression.


title: Inducing Specific Chromosome Mis-Segregation in Human Cells

abstract: Cancer cells display persistent underlying chromosomal instability that results in chromosome mis-segregation, the formation of micronuclei, and abnormal numbers of chromosomes (aneuploidy). These features are common to nearly all human cancers, with individual tumour types intriguingly exhibiting characteristic subsets of whole, and sub-chromosomal aneuploidies. To date, few methods to induce specific aneuploidies at will exist, hampering the investigation of functional consequences of recurrent aneuploidies. Moreover, although some human cell lines with specific aneuploidies exist, the acute cellular responses to specific chromosomal instability events remain unknown. We therefore investigated the possibility of sabotaging the mitotic segregation of specific chromosomes using nuclease-dead CRISPR-Cas9 (dCas9) as a cargo carrier to specific genomic loci. We recruited the kinetochore-nucleating domain of centromere protein CENP-T to assemble ectopic kinetochores either near the centromere of chromosome 9, or the telomere of chromosome 1. Ectopic kinetochore assembly led to increased chromosome instability and aneuploidy of the target chromosomes, providing the potential to create  designer karyotypes and study their immediate downstream cellular responses in a wide range of cell types. Overall, our findings provide new insights into ectopic kinetochore biology, and also represent an important step towards investigating the role of specific aneuploidy and chromosome mis-segregation events in diseases associated with aneuploidy.


title: Why resource dynamics matter in the optimization of microbial communities

abstract: The impact of resource supply on microbial community dynamics is rarely treated explicitly in research into the control and optimization of natural and synthetic microbial communities. Using simulations, we show that compositional overlap between microbial communities decays rapidly with a change in the frequency of resource pulsing, particularly in the presence of a metabolic trade-off in resource affinity and maximum growth rate. We conclude that resource supply dynamics should be considered both a constraint and a tuning mechanism in microbial community optimization.

category: ecology


title: Do Neotropical adult dobsonflies (Megaloptera, Corydalidae) feed on solid material in the nature? Information from a gut content study

abstract: Females of two species of corydalids (Megaloptera), Corydalus magnus Contreras-Ramos, 1998 and Platyneuromus soror (Hagen, 1861), are studied with the aim to determine if solid food is present in their guts, which would indicate the existence of feeding on solid food during the imaginal life in nature. Gut anatomical architecture is also studied and described for females of both species, showing no peculiarities regarding the existing information in other close related species and confirming that digestive morphology is conserved in Megaloptera, as previously reported in literature. No significant solid food was detected in any of the species, so in the nature, they may rely more on liquid sources and/or fat reserves produced in the larval stage for the energetic requirements of adult life.


title: NIPBL and WAPL balance cohesin activity to regulate chromatin folding and gene expression

abstract: The relationship between cohesin-mediated chromatin looping and gene expression remains unclear. We investigated the roles of NIPBL and WAPL, two regulators of cohesin activity, in chromatin folding and transcription in human cells. Consistent with their opposing roles in cohesin regulation, depletion of these factors showed opposite effects on levels of chromatin-bound cohesin and spatial insulation of neighboring domains. We find that NIPBL or WAPL depletion each alter the expression of ~2,000 genes, most of which are uniquely sensitive to either regulator. We find that each set of differentially expressed genes are enriched at chromatin loop anchors and clustered within the genome, suggesting there are genomic regions sensitive to either more or less cohesin. Remarkably, co-depletion of both regulators rescued chromatin misfolding and gene misexpression compared to either single knockdown. Taken together, we present a model in which the relative, rather than absolute, levels of NIPBL and WAPL are required to balance cohesin activity in chromatin folding to regulate transcription.

category: genetics


title: High-resolution transcriptomics of bovine purified protein derivative-stimulated peripheral blood from cattle infected with Mycobacterium bovis across an experimental time course

abstract:
ObjectivesImproved bovine tuberculosis (bTB) diagnostics with higher sensitivity and specificity are urgently required. A better understanding of the peripheral blood transcriptional response of Mycobacterium bovis-infected animals after bovine purified protein derivative (PPD-b) stimulation of whole blood--an important component of current bTB diagnostics--will provide new information for development of better diagnostics.

MethodsRNA sequencing (RNA-seq) was used to study the peripheral blood transcriptome after stimulation with PPD-b across four time points (-1 wk pre-infection, and +1 wk, +2 wk, and +10 wk post-infection) from a 14-week M. bovis infection time course experiment with ten age-matched Holstein-Friesian cattle.

ResultsIn vitro PPD-b stimulation of peripheral blood from M. bovis-infected and non-infected cattle elicited a strong transcriptional response. Comparison of PPD-b stimulated, and unstimulated samples revealed higher expression of genes encoding cytokine receptors, transcription factors, and interferon-inducible proteins. Lower expression was seen for genes encoding proteins involved in antimicrobial activity, C-type lectin receptors, inhibition of signal transduction, and genes encoding metal ion transporters.

ConclusionsA transcriptional signature associated with the peripheral blood response to PPD-b stimulation consisting of 170 genes was identified exclusively in the post-infection time points. Therefore, this represents a panel of potential biomarkers of M. bovis infection.

category: genomics


title: Molecular diversity and phenotypic pleiotropy of genomic regulatory loci derived from human endogenous retrovirus type H (HERVH) promoter LTR7 and HERVK promoter LTR5_Hs and their impacts on pathophysiology of Modern Humans.

abstract:
Targeted DNA sequences conservation analyses of 17 primate species demonstrated that human endogenous retroviruses (HERV) LTR7/HERVH and LTR5_Hs/HERVK appear to have distinct evolutionary histories charted by evidence of the earliest presence and expansion of highly-conserved (HC) LTR sequences. HC-LTR7 loci were mapped to genomes of Old World Monkeys (18% of all HC-LTR7 loci), suggesting that LTR7/HERVH have entered germlines of primates in Africa after the separation of the New World Monkey lineage. HC-LTR5_Hs loci have been identified in the Gibbons genome (24% of all HC-LTR5_Hs loci), suggesting that LTR5_Hs/HERVK successfully colonized primates germlines after the segregation of Gibbons species. Subsequently, both LTR7 and LTR5_Hs undergo a marked [~]4-5-fold expansion in genomes of Great Apes. Timelines of quantitative expansion of both LTR7 and LTR5_Hs loci during evolution of Great Apes appear to replicate the consensus evolutionary sequence of increasing cognitive and behavioral complexities of non-human primates, which seems particularly striking for LTR7 loci and 11 distinct LTR7 subfamilies.

Consistent with previous reports, identified in this study 351 human-specific (HS) insertions of LTR7 (175 loci) and LTR5_Hs (176 loci) regulatory sequences have been linked to genes implicated in establishment and maintenance of naive and primed pluripotent states and preimplantation embryogenesis phenotypes. Unexpectedly, HS regulatory LTRs appear linked with genes encoding markers of 12 distinct cells populations of fetal gonads, as well as genes implicated in physiology and pathology of human spermatogenesis, including Y-linked spermatogenic failure, oligo- and azoospermia.

Granular investigations of genes linked with eleven LTR7 subfamilies revealed that mammalian offspring survival (MOS) genes seem to remain one of consistent regulatory targets throughout [~]30 MYA of the divergent evolution of LTR7 loci. Differential GSEA of MOS versus non-MOS genes identified clearly discernable dominant enrichment patterns of phenotypic traits affected by MOS genes linked with LTR7 (562 MOS genes) and LTR5_Hs (126 MOS genes) regulatory loci across the large panel of genomics and proteomics databases reflecting a broad spectrum of human physiological and pathological traits. GSEA of LTR7-linked MOS genes identified more than 2200 significantly enriched records of human common and rare diseases and gene signatures of 466 significantly enriched records of Human Phenotype Ontology traits, including 92 genes of Autosomal Dominant Inheritance and 93 genes of Autosomal Recessive Inheritance.

One of the most informative categories of genes linked with LTR7 elements were genes implicated in functional and morphological features of central nervous system, including genes regulating synaptic transmission and protein-protein interactions at synapses, as well as gene signatures differentially regulated in cells of distinct neurodevelopmental stages and morphologically diverse cell types residing and functioning in human brain. These include Neural Stem/Precursor cells, Radial Glia cells, Bergman Glia cells, Pyramidal cells, Tanycytes, Immature neurons, Interneurons, Trigeminal neurons, GABAergic neurons, and Glutamatergic neurons. GSEA of LTR7-linked regulatory targets identified significantly enriched sets of genes encoding markers of more than 80 specialized types of neurons and markers of 521 human brain regions, most prominently, subiculum and dentate gyrus amongst top significantly enriched records. These observations were validated and extended by identification and characterization of 1944 genes comprising high-fidelity down-steam regulatory targets of LTR7 and/or LTR5_Hs loci, which are markedly enriched for genes implicated in neoplasm metastasis, intellectual disability, autism, multiple cancer types, Alzheimers, schizophrenia, and other brain disorders. Despite distinct evolutionary histories of retroviral LTRs, genes representing down-stream regulatory targets of LTR7 and LTR5_Hs elements exert the apparently cooperative and exceedingly broad phenotypic impacts on human physiology and pathology. Observations reported in this contribution highlight the need to accelerate the in-depth experimental and translational explorations of these important genomic determinants of Modern Humans health and disease states.


title: Clearing of ribosome collisions by the ribosome quality control trigger complex RQT

abstract:
After translational stalls, colliding eukaryotic ribosomes are cleared through dissociation into subunits by the ribosome quality control trigger complex, RQT, by an unknown mechanism. Here we show that RQT requires accessible mRNA and the presence of a neighboring ribosome. Cryo-EM of several RQT-ribosome complexes revealed the structural basis of splitting: RQT engages the 40S subunit of the lead ribosome and can switch between two conformations. We propose a mechanistic model in which the Slh1 helicase subunit of RQT applies a pulling force on the mRNA, causing destabilizing conformational changes of the 40S subunit. The collided ribosome functions as a ram or giant wedge, ultimately resulting in subunit dissociation. Our findings provide a first conceptual framework for a helicase driven ribosomal splitting mechanism.

One-Sentence SummaryRQT clears collided ribosomes by pulling mRNA to trigger destabilizing conformational transitions for subunit dissociation.

category: biochemistry


title: Tissue Specific Knockout of the Cardiolipin Transacylase Enzyme TAFAZZIN in Both Liver and Pancreatic Beta Cells Protects Mice From Diet-Induced Obesity

abstract: Mutations in the TAFAZZIN gene result in the X-linked genetic disease Barth Syndrome. The protein product tafazzin is a transacylase enzyme that remodels the phospholipid cardiolipin with fatty acids. Some Barth syndrome boys exhibit a lean phenotype. In addition, whole body knockdown of tafazzin in mice protects them from diet-induced obesity through increased hepatic fatty acid oxidation and reduced basal insulin secretion. We thus hypothesized that tafazzin deficiency in both the liver and beta cells of the pancreas contribute to this lean phenotype. Through a Cre-Lox approach we generated control, liver-specific, pancreatic beta cell-specific and liver and pancreatic beta cell-specific double knockout male mice. The animals were fed a high fat diet for 8 weeks and body weight, liver weight and fat pad weights determined. Liver-specific or pancreatic beta cell-specific male tafazzin knockout mice accumulated weight gain ({approx}40% increase in body weight) at the same rate as control animals. In contrast, the liver- and beta cell-specific double tafazzin knockout mice exhibited a reduced rate of weight gain by 8 weeks ({approx}26% increase in body weight) compared to control or the single tafazzin knockout animals. In addition, at 8 weeks the double tafazzin knockout mice exhibited reduced weight gain in tissues known to accumulate fat including the liver, the gonadal, inguinal and perirenal white adipose tissues and the brown adipose tissue. Thus, liver- and pancreatic beta cell-specific double tafazzin knockout male mice are protected from high fat diet induced weight gain and fat accumulation. These results may partially explain why some Barth Syndrome boys exhibit a lean phenotype.


title: Engineering and characterization of a long half-life relaxin receptor RXFP1 agonist

abstract: Relaxin-2 is a peptide hormone with important roles in human cardiovascular and reproductive biology. Its ability to activate cellular responses such as vasodilation, angiogenesis, and anti-inflammatory and anti-fibrotic effects have led to significant interest in using relaxin-2 as a therapeutic for heart failure and several fibrotic conditions. However, recombinant relaxin-2 has a very short serum half-life, limiting its clinical applications. Here we present protein engineering efforts targeting the relaxin-2 hormone in order to increase its serum half-life, while maintaining its ability to activate the G protein-coupled receptor RXFP1. To achieve this, we optimized a fusion between relaxin-2 and an antibody Fc fragment, generating a version of the hormone with a circulating half-life of up to five days in mice while retaining potent agonist activity at the RXFP1 receptor both in vitro and in vivo.

category: pharmacology and toxicology


title: The mitogen-activated protein kinase kinase kinase, ILK5, regulates plant purinergic receptor-mediated, innate immunity

abstract:
Mitogen-activated protein (MAP) kinase signaling cascades play important roles in the regulation of eukaryotic defense against various pathogens. Activation of the extracellular ATP (eATP) receptor P2K1 triggers MAP kinase 3 and 6 (MPK3/6) phosphorylation, which leads to elevated defense responses in Arabidopsis. However, the mechanism by which P2K1 activates the MAPK cascade is unclear. In this study, we identified Raf-like MAPKKK ILK5 as a downstream substrate of the P2K1 kinase. P2K1 phosphorylates ILK5 on serine 192. The interaction between P2K1 and ILK5 was confirmed both in vitro and in planta and their interaction was enhanced by ATP treatment. Similar to P2K1 expression, ILK5 expression levels were highly induced by treatment with ATP, flg22, Pseudomonas syringae pv. tomato DC3000, and various abiotic stresses, such as wounding. ILK5 interacts with both MKK4 and MKK5, but only MKK5 is phosphorylated by ILK5. Moreover, phosphorylation of MPK3/6 was significantly reduced upon ATP treatment in ilk5 mutant plants, relative to wild-type. The ilk5 mutant plants showed higher susceptibility to P. syringae pathogen infection relative to wild-type plants. Plants expressing only the mutant ILK5S192A protein, lacking kinase activity, did not activate the MAPK cascade upon ATP addition. Taken together, the results suggest that eATP activation of P2K1 results in transphosphorylation of the Raf-like MAPKKK ILK5, which subsequently triggers the MAPK cascade, culminating in activation of MAPK3 and 6 associated with an elevated innate immunity response.

Significance statementPathogens invasion and subsequent wound stress activates extracellular ATP-mediated purinergic signaling cascades, a danger associated molecular pattern (DAMP) signal, which induces phosphorylation of MAPKs. Previous studies revealed that the P2K1 purinergic receptor increases MPK3/6 phosphorylation in response to eATP signaling cascades in Arabidopsis. However, the specific mechanism by which this occurs remains unknown. Here, we describe the isolation and characterization of Raf-like MAPKKK ILK5 (Integrin-linked Kinase 5) as a downstream substrate of P2K1 kinase activity. Initiation of an eATP-dependent signaling pathway by phosphorylation of ILK5 with subsequent activation of MKK5, leading to activation of MPK3/6 and downstream events is crucial to the plant innate immunity response.

category: plant biology


title: Vitis vinifera plants edited in DMR6 genes show improved resistance to downy mildew.

abstract: The production and cultivation of vines (Vitis vinifera) tolerant or resistant to diseases such as downy mildew (DM) is a promising strategy to reduce fungicides and help viticulture sustainability. In many crops, generation of knock-out mutants in host genes controlling susceptibility to DM, such as Downy Mildew Resistant 6 (DMR6) is a strategy of proven success to obtain resistant plants, while the effect of mutations in DMR6 genes has yet to be demonstrated in grapevine. In addition, small mutations in genes governing important traits can be obtained by gene-editing while maintaining the genetic background of commercially important clones. Moreover, very recent advances in the technology of gene-editing allowed to produce non-transgenic grapevine mutants, by regeneration of protoplasts previously edited with the CRISPR-Cas9 ribonucleoprotein. This approach may revolutionize the production of new grapevine varieties and clones, but it requires knowledge on the targets, and an extensive evaluation of the impact of their mutation on plant phenotype and fitness. In this work we generated single and double knock-out mutants in DMR6 susceptibility (S) genes in multiple grapevine cultivars with improved resistance to DM.


title: Transcriptome profiling of the dorsomedial prefrontal cortex in suicide victims

abstract: The default mode network (DMN) plays an outstanding role in psychiatric disorders. Still, gene expressional changes in its major component, the dorsomedial prefrontal cortex (DMPFC), have not been characterized. We used RNA-sequencing in postmortem DMPFC samples to investigate suicide victims compared to control subjects. Most of the data variance (79%) was associated with expression changes between suicide and control samples. 1400 genes differed using log2FC > {+/-} 1 and adjusted p-value < 0.05 criteria between groups. Genes associated with depressive disorder, schizophrenia and impaired cognition were strongly overexpressed in top differentially expressed genes. Gene set enrichment and protein-protein interaction network analysis revealed that pathways related to cytokine receptor signaling were enriched in downregulated while glutamatergic synaptic signaling in upregulated genes in suicide individuals. A validated differentially expressed gene, which is known to be associated with mGluR5, was the N-terminal EF-hand calcium-binding proteins 2 (NECAB2). In situ hybridization histochemistry and immunohistochemistry proved that NECAB2 is expressed in 2 different types of inhibitory neurons located in layers II-IV and VI, respectively. Our results imply extensive gene expressional alterations in the DMPFC related to suicidal behavior. Some of these genes may contribute to the altered mental state and behavior of suicide victims.


title: An intranasal stringent response vaccine targeting dendritic cells as a novel adjunctive therapy against tuberculosis

abstract: Lengthy tuberculosis (TB) treatment is required to address the ability of a subpopulation of persistent Mycobacterium tuberculosis (Mtb) to remain in a non-replicating, antibiotic-tolerant state characterized by metabolic remodeling, including induction of the RelMtb-mediated stringent response. We developed a novel therapeutic DNA vaccine construct involving fusion of the relMtb gene with the immature dendritic cell-targeting gene encoding chemokine MIP-3/CCL20. To augment mucosal immune responses, intranasal delivery was also evaluated. We found that the intramuscular MIP-3/relMtb (fusion) vaccine potentiates isoniazid activity more than a similar DNA vaccine expressing relMtb alone in a chronic TB mouse model (absolute reduction of Mtb burden: 0.63 log10 colony-forming units, P=0.0001), inducing pronounced Mtb-protective immune signatures. The intranasal fusion vaccine, an approach combining relMtb fusion to MIP-3 and intranasal delivery, demonstrated the greatest therapeutic effect compared to each approach alone, as evidenced by robust Th1 and Th17 responses systemically and locally and the greatest mycobactericidal activity when combined with isoniazid (absolute reduction of Mtb burden: 1.13 log10, P<0.0001, when compared to the intramuscular vaccine targeting relMtb alone). This DNA vaccination strategy may be a promising adjunctive approach combined with standard therapy to shorten curative TB treatment, and also serve as proof-of-concept for treating other chronic infections.


title: T cell Aryl Hydrocarbon Receptor Activity Tunes the Gut Microenvironment to Sustain Autoimmunity.

abstract:
Multiple sclerosis (MS) is an incurable T cell driven autoimmune disease of the central nervous system. The contribution of the environment and, in particular, the gut flora to MS etiology is well documented but not well understood. The aryl hydrocarbon receptor (AHR) binds numerous molecules present in the environment and is a therapeutic target for MS: however, its precise function in T lymphocytes, the orchestrators of MS, has not been described. Here, we show that CD4 specific Ahr knockout drives recovery in EAE by a decreasing T cell fitness. At the mechanistic level, we show that the absence of AHR changes the gut microenvironment composition to generate metabolites, such as bile salts and short chain fatty acids, that impact T cell viability and influence disease recovery. Our study demonstrates a newly emerging role for AHR in mediating the interdependence between T lymphocytes and the microbiota while identifying new potential molecular targets for the treatment of MS and other autoimmune diseases.

One Sentence SummaryThe aryl hydrocarbon receptor modulates the gut microenvironment to promote autoimmunity in a mouse model of multiple sclerosis.


title: Canal number and configuration are predictors of external root morphology

abstract: Within tooth roots canals can vary in shape and configuration, and it is not uncommon for a single root to contain multiple canals. Externally, root morphology also varies, though the range of variation, and its relation to canals remains little explored. This investigation of modern human post-canine teeth uses data from computerized tomography scans of a global sample of 945 modern humans to identify the most frequent phenotypes of root and canal morphologies, and investigate how canal number, shape, and configuration relate to external root morphology. Results (1) include descriptions and definitions of root and canal morphologies, counts, and configurations; (2) indicate that certain canal counts, morphologies, and configurations can predict external morphologies; and (3) that this pattern varies in individual teeth and roots in the maxilla and mandible.

category: evolutionary biology


title: Development of transformation for genome editing of an emerging model organism

abstract:
With the advances in genomic sequencing, many organisms with novel biological properties are ripe for use as emerging model organisms. However, to make full use of them, transformation methods need to be developed to permit genome editing. Here, we present development of transformation for the fungus fly Sciara (Bradysia) coprophila; this may serve as a paradigm for development of transformation for other emerging systems, especially insects. Sciara has a variety of unique biological features including locus-specific developmentally-regulated DNA amplification; chromosome imprinting; a monopolar spindle in male meiosis I; non-disjunction of the X chromosome in male meiosis II; X chromosome elimination in early embryogenesis; germ line limited (L) chromosomes; high resistance to radiation. Mining the unique biology of Sciara requires a transformation system to test mutations of DNA sequences that may play roles for these features. We describe a Sciara transformation system using a modified piggyBac transformation vector and detailed protocols we have developed to accommodate Sciara-specific requirements. This advance will provide a platform for us and others in the growing Sciara community to take advantage of this unique biological system. In addition, the versatile piggyBac vectors described here and transformation methods will be useful for other emerging model systems.

Author BiographiesSusan A. Gerbi (Ph.D. with Joseph Gall at Yale University 1970) is the George Eggleston, Professor of Biochemistry at Brown University. Her research includes chromosomes, DNA replication and ribosomal RNA. She was President and is a Fellow of ASCB, a Fellow of AAAS and received the RI Governors Award for Scientific Achievement. Other honors include RNA Society/CSHL Press Distinguished Research Mentor award; GSA George Beadle award; ASCB Senior Leadership/ Mentoring Award. She is a national leader in graduate education, including member of the National Academy of Sciences Panel on Bridges to Independence that led to the NIH K99 program, Chair of the AAMC Graduate Research Education Training Group; Chair of the FASEB Consensus Conference on Graduate Education. Yutaka Yamamoto (M.D. Kansai Medical University 1990; Ph.D. with Walter Gehring at Biozentrum - Basel 1995; postdoc with David Glover at Dundee University and University of Cambridge) is a research associate at Brown University.

Graphical Abstract

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title: High resolution shotgun metagenomics: the more data, the better?

abstract: In shotgun metagenomics (SM), the state of the art bioinformatic workflows are referred to as high resolution shotgun metagenomics (HRSM) and require intensive computing and disk storage resources. While the increase in data output of the latest iteration of high throughput DNA sequencing systems can allow for unprecedented sequencing depth at a minimal cost, adjustments in HRSM workflows will be needed to properly process these ever-increasing sequence datasets. One potential adaptation is to generate so-called shallow SM datasets that contain fewer sequencing data per sample as compared to the more classic high coverage sequencing. While shallow sequencing is a promising avenue for SM, detailed benchmarks using real data are lacking. In this case study, we took two public SM datasets, one moderate and the other massive in size and subsampled each dataset at various levels to mimic shallow sequencing datasets of various sequencing depths. Our results suggest that shallow SM sequencing is a viable avenue to obtain sound results regarding microbial community structures and that high depth sequencing does not bring additional elements for ecological interpretation. One area, however, where ultra-deep sequencing and maximizing the usage of all data was undeniably beneficial was in the generation of metagenome-assembled genomes (MAGs).

category: bioinformatics


title: Bats as ecosystem engineers in iron ore caves in the Carajas National Forest, Brazilian Amazonia

abstract: Ecosystem engineers are organisms able to modify their environment by changing the distribution of materials and energy, with effects on biotic and abiotic ecosystem components. Several ecosystem engineers are known, but for most of them the mechanisms behind their influence are poorly known. We detail the role of bats as ecosystem engineers in iron ore caves in the Carajas National Forest, Brazilian Amazonia, an area with > 1,500 caves, some holding [~]150,000 bats. We analyzed the chemical composition of guano deposits in bat caves, radiocarbon-dated those deposits, and elucidated the chemical mechanisms involved and the role the bat guano has on structuring those caves. The insect-composed guano was rich in organic matter, with high concentrations of carbon, nitrogen, phosphorus pentoxide and ferric oxide, plus potassium oxide, calcium and sulfur trioxide. Radiocarbon dating indicated guano deposits between 22,000 and 1,800 years old. The guano pH was mainly acid (from 2.1 to 5.6). Percolating waters in those bat caves were also acid (pH reaching 1.5), with the presence of phosphate, iron, calcium, nitrate and sulfate. Acid solutions due to guano decomposition and possible microbial activity produced various forms of corrosion on the cave[s] floor and walls, resulting in their enlargement. Caves with active or inactive bat colonies had, on average, horizontal projections 4.5 times larger, areas 4.4 times larger, and volumes 5.0 times bigger than the reginal average, plus more abundant, diversified and bigger speleothems. In an example of bioengineering, the long-term presence of bats (> 22,000 years) and the guano deposits they produce, mediated by biological and chemical interactions along millennia, resulted in very unique ecological, evolutionary and geomorphological processes, whose working are just beginning to be better understood by science. However, the current expansion of mineral activities coupled with the loosening of licensing and cave protection rules is a real conservation threat to the bat caves in Carajas. The destruction of those caves would represent and unacceptable loss of both speleological and biological heritage and we urge that, whenever they occur, bat caves and their colonies must be fully protected and left off-limits of mineral extraction.


title: Motor outcomes congruent with intentions may sharpen metacognitive representations

abstract:
We can monitor our intentional movements, in order to describe how we move our bodies. But it is unclear which information this metacognitive monitoring relies on. For example, when throwing a ball to hit a target, we might use the visual information about how the ball flew to metacognitively assess our performance. Alternatively, we might disregard the ball trajectory -- given that it is not directly relevant to our goal -- and metacognitively assess our performance based solely on whether we reached the goal of hitting the target. In two experiments we aimed to distinguish between these two alternatives and asked whether the distal outcome of a goal-directed action (hitting or missing a target) informs the metacognitive representations of our own movements. Participants performed a semi-virtual task where they moved their arm to throw a virtual ball at a target. After each throw, participants discriminated which of two ball trajectories displayed on the screen corresponded to the flight path of their throw and then rated their confidence in this decision. The task included two conditions that differed on whether the distal outcome of the two trajectories shown matched (congruent) or differed (incongruent). Participants were significantly more accurate in discriminating between the two trajectories, and responded faster in the incongruent condition and, accordingly, were significantly more confident on these trials. Crucially, we found significant differences in metacognitive performance (measured as meta-d/d) between the two conditions only on successful trials, where the virtual ball had hit the target. These results indicate that participants successfully incorporated information about the outcome of the movement into both their discrimination and confidence responses. However, information about the outcome selectively sharpened the precision of confidence ratings only when the outcome of their throw matched their intention. We argue that these findings underline the separation between the different levels of information that may contribute to body monitoring, and we provide evidence that intentions might play a central role in metacognitive motor representations.

HighlightsO_LIParticipants threw a virtual ball to hit a target
C_LIO_LIFollowing their throw participants selected between two plausible trajectories
C_LIO_LIOn half the trials, the two alternative trajectories differed in their distal outcome.
C_LIO_LIDiscrimination and confidence were higher in trials with different outcome.
C_LIO_LIMetacognitive performance was best in hit trials when the alternative missed.
C_LI


title: Disruption of anterior temporal lobe reduces distortions in memory from category knowledge

abstract: Memory retrieval does not provide a perfect recapitulation of past events, but instead an imperfect reconstruction of event-specific details and general knowledge. However, it remains unclear whether this reconstruction relies on mixtures of signals from different memory systems, including one supporting general knowledge. Here, we investigate whether the anterior temporal lobe (ATL) distorts new memories due to prior category knowledge. In this experiment (N=36), participants encoded and retrieved image-location associations. Most images locations were clustered according to their category, but some were in random locations. With this protocol, we previously demonstrated that randomly located images were retrieved closer to their category cluster relative to their encoded locations, suggesting an influence of category knowledge. We combined this procedure with transcranial magnetic stimulation (TMS) delivered to the left ATL before retrieval. We separately examined event-specific details (error) and category knowledge (bias) to identify distinct signals attributable to different memory systems. We found that TMS to ATL attenuated bias in location memory, but only for atypical category members. The magnitude of error was not impacted, suggesting that a memorys fidelity can be decoupled from its distortion by category knowledge. This raises the intriguing possibility that retrieval is jointly supported by separable memory systems.


title: Language and the cerebellum: structural connectivity to the eloquent brain

abstract: Much remains to be learned about the role(s) of the cerebellum in the neurobiology of language. Contemporary models have focused extensively on the left-hemisphere perisylvian cortex with the assumption that the cerebellum supports more peri-linguistic cognitive processes (e.g., verbal working memory). The goal of this study was to map the structural connectivity profile of domain-specific language regions of interest (ROIs) implicated in semantics, phonology, and syntax relative to connectivity for working memory. Functional imaging data and diffusion-weighted imaging data from the Human Connectome Project (HCP) were analyzed. We found that (a) working memory, motor activity, and language comprehension activated partially overlapping but mostly unique regions of the cerebellum; (b) the linguistic portion of the cerebello-thalamo-cortical circuit was more extensive than the linguistic portion of the cortico-ponto-cerebellar tract; (c) there was a frontal-lobe bias in the connectivity from the cerebellum to the cerebrum; (d) there was some degree of specificity; and (e) for some of the cerebellar tracts, individual differences in picture identification ability correlated with variation in fractional anisotropy. These findings yield insights into the structural connectivity of the cerebellum as it relates to language comprehension - an ability that is unique to humans.


title: Binding of the peptide deformylase on the ribosome surface modulates the structure and dynamics of the exit tunnel interior

abstract: Proteosynthesis on ribosomes is regulated at many levels. Conformational changes of the ribosome, possibly induced by external factors, may transfer over large distances and contribute to the regulation. The molecular principles of this long-distance allostery within the ribosome remain poorly understood. Here, we use structural analysis and atomistic molecular dynamics simulations to investigate peptide deformylase (PDF), an enzyme that binds to the ribosome surface near the ribosomal protein uL22 during translation and chemically modifies the emerging nascent peptide. Our simulations of the entire ribosome-PDF complex reveal that the PDF undergoes a swaying motion on the ribosome surface at the sub-microsecond time scale. We show that the PDF affects the conformational dynamics of parts of the ribosome over distances of more than 5 nm. Using a supervised-learning algorithm we demonstrate that the exit tunnel is influenced by the presence or absence of PDF. Our findings suggest a possible effect of the PDF on the nascent peptide translocation through the ribosome exit tunnel.

category: biophysics


title: C. elegans Sine oculis/SIX-type homeobox genes act as homeotic switches to define neuronal subtype identities

abstract:
The classification of neurons into distinct types reveals hierarchical taxonomic relationships that reflect the extent of similarity between neuronal cell types. At the base of such taxonomies are neuronal cells that are very similar to one another but differ in a small number of reproducible and select features. How are very similar members of a neuron class that share many features instructed to diversify into distinct subclasses? We show here that the six very similar members of the C. elegans IL2 sensory neuron class, which are all specified by a homeobox terminal selector, unc-86/BRN3A/B, differentiate into two subtly distinct subclasses, a dorsoventral subclass and a lateral subclass, by the toggle switch-like action of the SIX/Sine-oculis homeobox gene unc-39. unc-39 is expressed only in the lateral IL2 neurons and loss of unc-39 leads to a homeotic transformation of the lateral into the dorsoventral class; conversely, ectopic unc-39 expression converts the dorsoventral subclass into the lateral subclass. Hence, a terminal selector homeobox gene controls both class-, as well as subclass-specific features, while a subordinate homeobox gene determines the ability of the class-specific homeobox gene to activate subtype-specific target genes. We find a similar regulatory mechanism to operate in a distinct class of six motor neurons. Our findings underscore the importance of homeobox genes in neuronal identity control and invite speculations about homeotic identity transformations as potential drivers of evolutionary novelty during cell type evolution in the brain.

SIGNIFICANCE STATEMENTAnatomical and molecular studies have revealed that in many animal nervous systems, neuronal cell types can often be subclassified into highly related subtypes with only small phenotypic differences. We decipher here the regulatory logic of such cell type diversification processes. We show that identity features of neurons that are highly similar to one another are controlled by master regulatory transcription factors and that phenotypic differences between related cell types are controlled by downstream acting transcription factors that promote or antagonize the ability of such a master regulatory factor to control unique identity features. Our findings help explain how neuronal cell types diversify and suggest hypothetical scenarios for neuronal cell type evolution.


title: Structural Basis of MicroRNA Biogenesis by Dicer-1 and Its Partner Protein Loqs-PB

abstract: In animals and plants, Dicer enzymes collaborate with double-stranded RNA-binding proteins to convert precursor-microRNAs (pre-miRNAs) into miRNA duplexes. We report six cryo-EM structures of Drosophila Dicer-1 and its partner Loqs-PB. The structures show Dicer-1*Loqs-PB (1) before binding pre-miRNA, (2) after binding and in a catalytically competent state, (3) after nicking one arm of the pre-miRNA, (4) following complete dicing and initial product release. Our reconstructions suggest that pre-miRNA binds a rare, open conformation of the Dicer-1*Loqs-PB heterodimer, enabling conformational proofreading. The Dicer-1 dsRBD and three Loqs-PB dsRBD domains form a tight belt around the pre-miRNA, distorting the RNA helix to place the scissile phosphodiester bonds in the RNase III active sites. Pre-mRNA cleavage shifts the RNA- binding domains and tightens Dicer-1, promoting product release. Our data suggest a model for how the Dicer-1*Loqs-PB complex effects a complete cycle of pre-miRNA recognition, stepwise endonuclease cleavage, and sequential product release.

category: molecular biology


title: A motor-based approach to induce chromosome-specific mis-segregations in human cells

abstract: Various cancer types exhibit highly characteristic and recurrent aneuploidy patterns. The origin of these cancer type-specific karyotypes, and the extent to which they contribute to cancer progression, remains to be elucidated, partly because introducing or eliminating specific chromosomes in human cells still poses a challenge. Here, we describe a novel strategy to mis-segregate specific chromosomes at will in different human cell types. We employed Tet repressor (TetR) or nuclease dead Cas9 (dCas9) to link a plant-derived microtubule minus-end-directed kinesin (Physcomitrella patens Kinesin14VIb) to integrated Tet operon repeats and chromosome-specific endogenous repeats, respectively. By live- and fixed-cell imaging, we observed poleward movement of the targeted loci during (pro)metaphase. Kinesin14VIb-mediated pulling forces on the targeted chromosome were often counteracted by forces from kinetochore-attached microtubules. This tug of war resulted in chromosome-specific segregation errors during anaphase, and revealed that spindle forces can heavily stretch chromosomal arms. Using chromosome-specific FISH and single-cell whole genome sequencing, we established that motor-induced mis-segregations result in specific arm-level, and to a lesser extent, whole chromosome aneuploidies, after a single cell division. Our kinesin-based strategy to manipulate individual mitotic chromosomes opens up the possibility to investigate the immediate cellular responses to specific (arm level) aneuploidies in different cell types; an important step towards understanding how recurrent aneuploidy patterns arise in different cancer types.


title: Sorting of secretory proteins at the trans-Golgi network by TGN46

abstract: Secretory proteins are sorted at the trans-Golgi network (TGN) for export into specific transport carriers. However, the molecular players involved in this fundamental process remain largely elusive. Here, we identified the transmembrane protein TGN46 as a cargo receptor for the export of secretory proteins in CARTS - a class of protein kinase D-dependent TGN-to-plasma membrane carriers. We show that TGN46 is necessary for cargo sorting and loading into nascent carriers at the TGN. By combining quantitative fluorescence microscopy and mutagenesis approaches, we further discovered that the TGN46 lumenal domain encodes for its cargo sorting function. Interestingly, this domain forms liquid droplets in vitro by means of liquid-liquid phase separation, a physical mechanism that may assist in TGN46-mediated cargo sorting. In summary, our results define a cellular function of TGN46 in sorting secretory proteins for export from the TGN. These findings could open new therapeutic avenues for diseases that parallel secretory malfunction.


title: The stoichiometric interaction model for mesoscopic molecular dynamics simulations of liquid-liquid phase separation

abstract:
Liquid-liquid phase separation (LLPS) has received considerable attention in recent years for explaining the formation of cellular biomolecular condensates. The fluidity and the complexity of their components make molecular simulation approaches indispensable for gaining structural insights. Domain-resolution mesoscopic model simulations have been explored for case in which condensates are formed by multivalent proteins with tandem domains. One problem with this approach is that interdomain pairwise interactions cannot regulate the valency of the binding domains. To overcome this problem, we propose a new potential, the stoichiometric interaction (SI) potential. First, we verified that the SI potential maintained the valency of the interacting domains for the test systems. We then examined a well-studied LLPS model system containing tandem repeats of SH3 domains and proline-rich motifs. We found that the SI potential alone cannot reproduce the phase diagram of LLPS quantitatively. We had to combine the SI and a pairwise interaction; the former and the latter represent the specific and non-specific interactions, respectively. Biomolecular condensates with the mixed SI and pairwise interaction exhibited fluidity, whereas those with the pairwise interaction alone showed no detectable diffusion. We also compared the phase diagrams of the systems containing different numbers of tandem domains with those obtained from the experiments, and found quantitative agreement in all but one case.

SIGNIFICANCECells organize their interior structures as not only membrane-bound organelles but also as membrane-less organelles. Membrane-less organelles, such as stress granules, Cajal bodies, and postsynaptic density, are biomolecular condensates in which many biomolecules are gathered owing to their interactions. In some cases, biomolecular condensates are formed by tandemly connected multidomain proteins. In such cases, a mesoscopic simulation model representing each domain as a particle is effective; however, the problem with this approach is that a domain-domain pairwise interaction cannot regulate the well-defined valency. To overcome this problem, in this study, we have developed a new potential, viz. the stoichiometric interaction potential, and confirmed that this potential can reproduce the liquid-liquid phase separation of multidomain proteins, a hallmark of the membrane-less organelles.


title: Weak tension accelerates hybridization and dehybridization of short oligonucleotides

abstract: The hybridization and dehybridization of DNA subject to tension is relevant to fundamental genetic processes and to the design of DNA-based mechanobiology assays. While strong tension accelerates DNA melting and decelerates DNA annealing, the effects of tension weaker than 5 pN are less clear. In this study, we developed a DNA bow assay, which uses the bending rigidity of double-stranded DNA (dsDNA) to exert weak tension on a single-stranded DNA (ssDNA) target in the range of 2pN to 6pN. Combining this assay with single-molecule FRET, we measured the hybridization and dehybridization kinetics between a 15 nt ssDNA under tension and a 8-9 nt oligo, and found that both the hybridization and dehybridization rates monotonically increase with tension for various nucleotide sequences tested. These findings suggest that the nucleated duplex in its transition state is more extended than the pure dsDNA or ssDNA counterpart. Our simulations using the coarse-grained oxDNA2 model indicate that the increased extension of the transition state is due to exclusion interactions between unpaired ssDNA regions in close proximity to one another. This study highlights an example where the ideal worm-like chain models fail to explain the kinetic behavior of DNA in the low force regime.


title: A genotoxic antibody drug conjugate targeting CD276/B7H3 demonstrates efficacy across multiple biomarker defined classes of treatment refractory metastatic prostate cancer

abstract:
B7H3, a cell surface protein often associated with higher grade solid tumors of various origins, is an attractive therapeutic target due to its frequent upregulated expression relative to normal tissue. In a large cohort of treatment-resistant, metastatic castrate resistant prostate cancer (mPC) patient derived xenografts and organoids, we present a comprehensive therapeutic and biomarker analysis. The 26 models tested encompass the heterogeneous genomic and histological categories of clinical mPC. In vitro and in vivo treatment with an anti-B7H3 targeted antibody conjugated to a pyrrolobenzodiazepine (PBD) cytotoxic agent were efficacious in 70% of models. All models expressed some B7H3 protein, the levels of which did not correlate with responses. Underlying susceptibilities to B7H3-PBD-ADC exposure were observed as multiple imperfectly overlapping classes of biomarkers, unrelated to androgen receptor dependence and substantially expanded beyond current homologous repair deficiencies. RB1 deficiency and/or replication stress, particularly prominent in small cell neuroendocrine prostate cancer (SCNPC), predicted responsiveness. The presence of SLFN11 in SCNPC and adenocarcinoma models correlated perfectly with vulnerability, although an absence of SLFN11 was not predictive. Of particular interest, an elevated interferon signature in adenocarcinomas, associated with alteration of wild type TP53, was quantitatively correlated with responsiveness. Finally, we also observed that susceptible models uniquely associated with the loss of select downstream DNA repair proteins. The above biomarkers are expressed in toto in about 70% of mPC clinical samples, suggesting broad potential for mechanistically complementary treatment of tumors escaping current regimens.

One Sentence SummaryB7H3-PBD-ADC sensitivity in metastatic prostate cancer is associated with replication stress, SLFN11 expression related to interferon signaling, and alterations in specific DNA repair factors.

category: cancer biology


title: mTORC1 activity is essential for disease progression in chronic lymphocytic leukemia

abstract: The precise role of mechanistic target of rapamycin complex 1 (mTORC1) during chronic lymphocytic leukemia (CLL) pathogenesis remains to be elucidated. Targeted deletion of mTORC1 component Raptor in adult mice reveals that mTORC1 function is essential for initiation and maintenance of CLL. Raptor-deficient bone marrow-derived PKC-KR transduced haemopoietic progenitors failed to generate a CLL-like disease in vitro, due to an inability to overcome the mTORC1-mediated block in B cell lineage commitment. Induction of Raptor-deficiency in NSG mice transplanted with Mx1-Raptor BM-derived PKC-KR transduced cells after disease was established, revealed a reduced CLL-like disease load and a significant increase in survival in the mice. Interestingly in mice transplanted with an aggressive CLL-like disease, rapamycin treatment reduced disease burden more effectively than AZD2014 (dual mTORC1/2 inhibitor), indicating a skew towards mTORC1 sensitivity with more aggressive leukemic disease. Rapamycin efficiently targeted the translation elongation axis eEF2/eEF2K downstream of mTORC1, resulting in eEF2 inactivation through induction of eEF2T56 phosphorylation. Rapamycin treatment of primary CLL cells halted proliferation, modulated eEF2K/eEF2 phosphorylation and inhibited MCL1 expression. Our studies demonstrate that mTORC1 plays an essential role in leukemia progression in vitro and in vivo in our CLL mouse model, with evidence for increased rapamycin sensitivity in aggressive secondary CLL transplants. Furthermore, the suppression of translation elongation through inactivation of eEF2 may offer a novel therapeutic target for blocking CLL progression.


title: Statistical evaluation of metaproteomics and 16s rRNA amplicon sequencing techniques for the study of the gut microbiota establishment of infants with cystic fibrosis

abstract: The newborn screening for cystic fibrosis (CF) can identify affected but still asymptomatic infants. The selection of omic technique for gut microbiota study is therefore crucial due to both the small amount of feces available and the low microorganism load. Our aim was to compare the agreement between 16S rRNA amplicon sequencing and metaproteomics by a robust statistical analysis including McNemar (taxa presence) test and Bland-Altman (taxa abundance) data plotting for studying the sequential establishment of the gut microbiota during the first year of life in 8 infants with CF (28 fecal samples). The taxonomic assignation was similar by the two techniques, whereas certain discrepancies were observed in the abundance detection, mostly the underrepresentation of Bifidobacterium and the overestimation of certain Firmicutes and Proteobacteria by amplicon sequencing. During the first months of life, the CF gut microbiota is characterized by poor alpha diversity, a significant enrichment of Ruminococcus gnavus, the expression of certain virulent bacterial traits, and the detection of human inflammation-related proteins. Our study provides an extended comparative analysis with robust statistical support that could optimize the use of both approaches for gut microbiota research. Metaproteomics provides information on composition and functionality, as well as data on host-microbiome interactions. Its strength is the identification and quantification of Actinobacteria and certain classes of Firmicutes. Both techniques detected an aberrant microbiota in infants with CF during their first year of life, dominated by the enrichment of R. gnavus within a human inflammatory environment.


title: SUMOylation controls the rapid transcriptional reprogramming induced by anthracyclines in Acute Myeloid Leukemias

abstract: Genotoxicants have been used for decades as front-line therapies against cancer on the basis of their DNA-damaging actions. However, some of their non-DNA-damaging effects are also instrumental for killing dividing cells. We report here that the anthracycline Daunorubicin (DNR), one of the main drugs used to treat Acute Myeloid Leukemia (AML), induces broad transcriptional changes in AML cells before cell death induction. The regulated genes are particularly enriched in genes controlling cell proliferation and death, as well as inflammation and immunity. These transcriptional changes are preceded by DNR-dependent deSUMOylation of chromatin proteins, which limits both the positive and negative effects of DNR on transcription. Quantitative proteomics shows that proteins that are deSUMOylated in response to DNR are mostly transcription factors, transcriptional co-regulators and chromatin organizers. Among them, the CCCTC-binding factor CTCF is highly enriched at SUMO-binding sites found in cis-regulatory regions. This is notably the case at the promoter of the DNR-induced NFKB2 gene. Its induction is preceded by a SUMO-dependent reconfiguration of chromatin loops engaging its CTCF- and SUMO-bound promoter with distal cis-regulatory regions. Altogether, our work suggests that one of the earliest effects of DNR in AML cells is a SUMO-dependent transcriptional reprogramming.


title: Nanostructure of the desmosomal plaque

abstract: Desmosomes are considered one of the most important intercellular junctions with respect to mechanical strength. Therefore, their spatial distribution and structure is of interest with respect to understanding both healthy and diseased tissue. Previous studies have imaged desmosomes in tissue slices using transmission electron microscopy, or low-resolution confocal images, but both these techniques lack the ability to resolve the 3-dimensional structure of the desmosomes. In this work it was possible to determine the 3D-nanostructure of single desmosomal complexes in both mouse and human epidermis, by 3D stimulated emission depletion (STED) microscopy. STED images of desmoplakin and the desmosomal cadherin, desmoglein revealed that desmosomes form ring-like structures, distributed over the cell surface, with diameters of around 1 m. STED images of the tight junction plaque protein ZO1 also displayed ring formations, suggesting a common structure for intercellular junctions. Measurements of the desmosomal plaque protein, desmoplakin showed an increased intercellular plaque distance during the stratum basale (0.23{+/-}0.027{micro}m) to stratum spinosum (0.28{+/-}0.039 {micro}m) transition.


title: Polio Virotherapy of Malignant Glioma Engages the Tumor Myeloid Infiltrate and Induces Diffuse Microglia Activation

abstract: Malignant gliomas commandeer abundant inflammatory infiltrates with glioma-associated macrophages and microglia (GAMM) actively promoting tumor progression. Like all cells in the mononuclear phagocytic system, macrophages and microglia constitutively express the poliovirus receptor, CD155. Besides myeloid cells, CD155 is widely upregulated ectopically in the neoplastic compartment of malignant gliomas (and solid cancers in general). Intratumor treatment with the highly attenuated rhino:poliovirus chimera, PVSRIPO, yielded long-term survival with durable radiographic responses in patients with recurrent glioblastoma (Desjardins et al. New England Journal of Medicine, 2018). Here, we studied mechanisms of PVSRIPO immunotherapy in mouse brain tumor models to decipher contributions of myeloid vs. malignant cells to antitumor efficacy. PVSRIPO treatment caused intense engagement of the GAMM infiltrate associated with substantial, but transient tumor regression. This was accompanied by diffuse microglia activation and proliferation in the normal central nervous system (CNS) surrounding the tumor, extending to the ipsilateral and even the contralateral hemispheres. PVSRIPO-instigated microglia activation occurred against a backdrop of sustained innate antiviral inflammation, associated with induction of the PD-L1 immune checkpoint on GAMM. Combining PVSRIPO with PD1/PD-L1 blockade led to durable remissions. Our work implicates GAMM as active drivers of PVSRIPO-induced antitumor inflammation and reveals profound and widespread neuroinflammatory activation of the CNS-resident myeloid compartment by PVSRIPO.


title: pH as an eco-evolutionary driver of priority effects

abstract: Priority effects, where arrival order and initial relative abundance modulate local species interactions, can exert taxonomic, functional, and evolutionary influences on ecological communities by driving them to alternative states. It remains unclear if these wide-ranging consequences of priority effects can be explained systematically by a common underlying factor. Here, we identify such a factor in an empirical system. In a series of field and laboratory studies, we focus on how pH affects nectar-colonizing microbes and their interactions with plants and pollinators. In a field survey, we found that nectar microbial communities in a hummingbird-pollinated shrub, Diplacus aurantiacus, exhibited patterns indicative of alternative stable states through domination by either bacteria or yeasts within individual flowers. In laboratory experiments, Acinetobacter nectaris, the bacterium most commonly found in D. aurantiacus nectar, exerted a strongly negative priority effect against Metschnikowia reukaufii, the most common nectar-specialist yeast, by reducing nectar pH. This priority effect likely explains the mutually exclusive pattern of dominance found in the field survey. Furthermore, experimental evolution simulating hummingbird-assisted dispersal between flowers revealed that M. reukaufii could evolve rapidly to improve resistance against the priority effect if constantly exposed to A. nectaris-induced pH reduction. Finally, in a field experiment, we found that low nectar pH could reduce nectar consumption by hummingbirds, suggesting functional consequences of the pH-driven priority effect for plant reproduction. Taken together, these results show that it is possible to identify an overarching factor that governs the eco-evolutionary dynamics of priority effects across multiple levels of biological organization.


title: Functional characterization of putative ecdysone transporters in lepidopteran pests

abstract:
The insect steroid hormone ecdysone plays a critical role in insect development. Several recent studies have shown that ecdysone is transported through Organic Anion Transporting Polypeptides (OATPs) in insects such as flies and mosquitoes. However, the conservation of this mechanism across other arthropods and the role of this transporter in canonical ecdysone pathways are less well studied. Herein we functionally characterized the putative ecdysone transporter OATP74D from two major agricultural moth pests: Helicoverpa armigera (cotton bollworm) and Spodoptera frugiperda (fall armyworm). Phylogenetic analysis of OATP transporters across the superphylum Ecdysozoa revealed that Oatp74D is well represented among arthropod species and appeared only at the root of the arthropod lineage. Partial disruption of Oatp74D in S. frugiperda decreased embryo hatching rate and larval survival, suggesting that this gene is essential for development in vivo. Depletion and re-expression of OatP74D in the lepidoptera cell line RP-HzGUT-AW1(MG) confirmed the genes role in ecdysone import and demonstrated that OATP74D is essential for the transcriptional activation of ecdysone responsive genes including caspase-3, implicating this transporter in cell death pathways. Establishment of a simple and robust luciferase assay using the RP-HzGUT- AW1(MG) cell line demonstrated that both HaOATP74D and SfOATP74D are inhibited by rifampicin, a well-known organic anion transporter inhibitor. Overall, this work sheds more light on ecdysone uptake mechanisms across insect species and broadens our knowledge of the physiological roles of OATPs in the transportation of endogenous substrates.

Author SummaryThe insect steroid hormone ecdysone is critical in regulating many aspects of insects life, including development and reproduction. A passive diffusion model was never functionally resolved, but was strongly supported until an organic anion transporting polypeptide was identified to mediate the transport of the hormone. The OATP74D, belonging to the Solute carrier superfamily, has been identified and functionally characterized for the first time in Drosophila melanogaster. Although phylogenetic analysis suggests that the Drosophila Oatp74D is probably conserved among several insect species, the theory for transporter mediated ecdysone uptake cannot be generalized to all insects without concrete proof. In here we provide functional evidence that the Oatp74D of two lepidopteran pest species: Helicoverpa armigera and Spodoptera frugiperda, is highly required for insect survival and development. Furthermore, we reveal that the OATP74D is necessary to regulate the expression of several ecdysone response genes, including caspase-3 which is involved in programmed cell death. In addition, we have developed a cell-based platform for screening chemical compounds against the lepidopteran orthologs of Oat74D and rifampicin was functionally shown to inhibit ecdysone uptake. Taken all together, our study reveals that Oatp74D is conserved among several arthropod species in the ecdysone pathway and given the high necessity for an effective control of these two lepidopteran species, we hypothesized that OATP74D could serve as a possible drug target in those two species.


title: The estrogen receptor α cistrome in human endometrium and epithelial organoids

abstract:
ContextEndometrial health is impacted by molecular processes that underlie estrogen responses.

ObjectiveTo define estrogen regulation of endometrial function by integrating the estrogen receptor alpha (ESR1) cistrome and transcriptome of endometrial biopsies taken from the proliferative and midsecretory phases of the menstrual cycle and hormonally stimulated endometrial epithelial organoids.

DesignESR1 ChIPseq and RNAseq were performed on proliferative or mid-secretory endometrial biopsies and on hormone treated organoid cultures.

SettingEndometrial samples were obtained from volunteers at outpatient research clinics for ChIPseq and for organoid culture.

Patients or Other ParticipantsParticipants were fertile, reproductive aged women with normal cycle length, and without any history of infertility or irregular cycles. In total, 5 new endometrial biopsies obtained from 5 women were used in this study and were analyzed together with previously published cycle stage endometrial RNAseq data.

Intervention(s)There were no interventions in this study.

Main Outcome Measure(s)The cycle stage specific ESR1 binding sites and gene expression identification of human endometrium and organoid cultures were integrated with changes in gene expression.

ResultsGenes with ESR1 binding in whole endometrium were enriched for chromatin modification and regulation of cell proliferation. The distribution of ESR1 binding sites in organoids was more distal to the gene promoter when compared to primary endometrium. Organoid estrogen/ESR1 candidate target genes impacted formation of cellular protrusions, and chromatin modification,

ConclusionsAnalysis of the ESR1 cistromes and transcriptomes from endometrium and organoids provides important resources for understanding how estrogen impacts endometrial health and function.


title: Dynamics and Energetics of Bottlenose Dolphins (Tursiops truncatus) Fluke-and-Glide Gait

abstract: Intermittent locomotion composed of periods of active flapping/stroking followed by inactive gliding has been observed with species that inhabit both aerial and marine environments. However, studies on the energetic benefits of a fluke-and-glide (FG) gait during horizontal locomotion are limited for dolphins. This work presents a physics-based model of FG gait and analysis of the associated costs of transport of bottlenose dolphins (Tursiops truncatus). New estimates of gliding drag coefficients for the model were estimated experimentally from free-swimming bottlenose dolphins. The data-driven approach used kinematic measurement from 84 hours of biologging tag data collected from 3 animals to estimate the coefficients. A set of 532 qualified gliding events were automatically extracted for gliding drag coefficient estimation, and an additional 783 FG bouts were parameterized and used to inform the model-based dynamic analysis. Experimental results indicate that FG gait was preferred at speeds around 2.2 - 2.7 m/s. Observed FG bouts had an average duty factor of 0.45 and gliding duration of 5 sec. The average associated metabolic cost of transport (COT) and mechanical cost of transport (MECOT) of FG gait are 2.53 and 0.35 J {middle dot} m-1 {middle dot} kg-1 at the preferred speeds. This corresponded to an 18.9% and 27.1% reduction in cost when compared to modeled continuous fluking gait at the same reference speed. Average thrust was positively correlated with fluking frequency and amplitude as animals accelerated during the FG bouts. While fluking frequency and amplitude were negatively correlated for a given thrust range. These results support the supposition that FG gait enhances the horizontal swimming efficiency of bottlenose dolphins and provides new dynamical insights into the gait of these animals.

category: animal behavior and cognition


title: Lagrange-NG: The next generation of Lagrange

abstract: AO_SCPLOWBSTRACTC_SCPLOWComputing ancestral ranges via the Dispersion Extinction and Cladogensis (DEC) model of biogeography is characterized by an exponential number of states relative to the number of regions considered. This is because the DEC model requires computing a large matrix exponential, which typically accounts for up to 80% of overall runtime. Therefore, the kinds of biogeographical analyses that can be conducted under the DEC model are limited by the number of regions under consideration. In this work, we present a completely redesigned efficient version of the popular tool Lagrange which is up to 2.5 times faster, which we call Lagrange-NG (Next Generation). We further reduce time-to-completion by introducing a multi-grained parallelization approach, achieving a total parallel speedup of 8.5 over Lagrange on a machine with 8 cores. In order to validate the correctness of Lagrange-NG, we also introduce a novel metric on range distributions for trees in order to assess the difference between any two range inferences. Finally, Lagrange-NG exhibits substantially higher adherence to coding quality standards. It improves a respective software quality indicator as implemented in the SoftWipe tool from average (5.5; Lagrange) to high (7.8; Lagrange-NG). Lagrange-NG is freely available under GPL2.


title: Integrated molecular and pharmacological characterization of patient-derived xenografts from bladder and ureteral cancers identifies new potential therapies.

abstract:
BackgroundMuscle-invasive bladder cancer (MIBC) and upper urinary tract urothelial carcinoma (UTUC) are molecularly heterogeneous. Despite chemotherapies, immunotherapies or anti-FGFR treatments, these tumors are still of poor outcome. Our objective was to develop a bank of patient-derived xenografts (PDXs) recapitulating molecular heterogeneity of MIBC and UTUC, to facilitate preclinical identification of therapies.

MethodsFresh tumors were obtained from patients and subcutaneously engrafted into immune-compromised mice. Patient tumors and matched PDXs were compared regarding histopathology, transcriptomic (microarrays) and genomic profiles (targeted-NGS). Several PDXs were treated with chemotherapy (cisplatin/gemcitabine) or targeted therapies (FGFR and EGFR inhibitors).

Results31 PDXs were established from 1 non-MIBC, 25 MIBC, 5 upper urinary tract tumors, including 28 urothelial (UCC) and 3 squamous-cell carcinomas (SCC). Integrated genomic and transcriptomic profiling identified PDXs of 3 different consensus molecular subtypes (Basal/Squamous, Luminal papillary and Luminal unstable), and included FGFR3-mutated PDXs. High histological and genomic concordance was found between matched patient tumor/PDX. Discordance in molecular subtypes, such as a basal/squamous patient tumor giving rise to a luminal papillary PDX, was observed (n=5) at molecular and histological levels. Ten models were treated with cisplatin-based chemotherapy and we did not observe association between subtypes and response. Of the 3 basal/squamous models treated with anti-EGFR therapy, two models were sensitive and one model, of sarcomatoid variant, was resistant. Treatment of 3 FGFR3-mutant PDXs with combined FGFR/EGFR inhibitors was more efficient than anti-FGFR3 treatment alone.

ConclusionsWe developed preclinical PDX models that recapitulate the molecular heterogeneity of MIBCs and UTUC, including actionable mutations, which will represent an essential tool in therapy development. Pharmacological characterization of the PDXs suggested that upper urinary tract and MIBCs, UCC but also SCC, with similar molecular characteristics could benefit from the same treatments including anti-FGFR for FGFR3-mutated tumors and anti-EGFR for basal ones and showed a benefit for combined FGFR/EGFR inhibition in FGFR3-mutant PDXs, compared to FGFR inhibition alone.


title: Mechanosensitive hormone signaling promotes mammary progenitor expansion and breast cancer progression

abstract:
Tissue stem-progenitor cell frequency has been implicated in tumor risk and progression. Tissue-specific factors linking stem-progenitor cell frequency to cancer risk and progression remain ill defined. Using a genetically engineered mouse model that promotes integrin mechanosignaling with syngeneic manipulations, spheroid models, and patient-derived xenografts we determined that a stiff extracellular matrix and high integrin mechanosignaling increase stem-progenitor cell frequency to enhance breast tumor risk and progression. Studies revealed that high integrin-mechanosignaling expands breast epithelial stem-progenitor cell number by potentiating progesterone receptor-dependent RANK signaling. Consistently, we observed that the stiff breast tissue from women with high mammographic density, who exhibit an increased lifetime risk for breast cancer, also have elevated RANK signaling and a high frequency of stem-progenitor epithelial cells. The findings link tissue fibrosis and integrin mechanosignaling to stem-progenitor cell frequency and causally implicate hormone signaling in this phenotype. Accordingly, inhibiting RANK signaling could temper the tumor promoting impact of fibrosis on breast cancer and reduce the elevated breast cancer risk exhibited by women with high mammographic density.

SummaryElevated mechano-signaling and matrix stiffness promote progesterone and RANK mediated expansion of mammary progenitors and breast cancer risk and progression.


title: Restriction of dietary fat, but not carbohydrate, alters brain reward circuitry in adults with obesity

abstract:
SUMMARYWeight loss diets often restrict either fat or carbohydrate, macronutrients that are sensed via distinct gut-brain pathways and differentially affect peripheral hormones and metabolism. To investigate whether reductions in dietary fat versus carbohydrate alter brain reward circuitry, we measured dopamine D2/3 receptor binding potential (D2BP) using PET and neural activity in response to visual food cues using fMRI in 17 inpatient adults with obesity during a eucaloric baseline diet and on the fifth day of isocaloric diets selectively reduced in either dietary fat or carbohydrate, in random order. Reduction of dietary fat, but not carbohydrate, decreased D2BP and decreased neural activity to food cues in brain reward regions. After the reduced fat diet, ad libitum intake shifted towards foods high in both fat and carbohydrates. These results suggest that dietary fat restriction increases tonic dopamine in brain reward regions thereby affecting food choice in ways that may hamper diet adherence.

IN BRIEFDarcey et al. found that selective reduction of dietary fat, but not carbohydrate, resulted in changes in brain dopamine receptor binding potential and neural activity in response to food cues, consistent with increased tonic dopamine and resulting in greater subsequent consumption of rewarding foods high in both fat and carbohydrate.

HIGHLIGHTSO_LICompared to a eucaloric baseline diet, selective reduction of dietary fat, but not carbohydrate, significantly decreased both dopamine D2/3 receptor binding potential and neural activity to visual food cues in brain reward regions.
C_LIO_LIMultimodal neuroimaging suggests that reduction of dietary fat, but not carbohydrate, increased tonic brain dopamine and influenced food choices in ways that may hamper diet adherence.
C_LI


title: Cortical facilitation of somatosensory inputs using gravity-related tactile information in patients with bilateral vestibular loss

abstract:
A few years after their bilateral vestibular loss, individuals usually show a motor repertoire that is almost back to normal. This recovery is thought to involve an up-regulation of the visual and proprioceptive information that compensates for the lack of vestibular information. Here, we investigated whether plantar tactile inputs, which provide body information relative to the ground and to the Earth-vertical, contribute to this compensation. More specifically, we tested the hypothesis that somatosensory cortex response to electric stimulation of the plantar sole in standing adults will be greater in patients (n = 10) with bilateral vestibular loss than in an aged-matched healthy group (n = 10). Showing significant greater somatosensory evoked potentials (i.e., P1N1) in patients than in controls, the electroencephalographic recordings supported this hypothesis. Furthermore, we found evidence that increasing the differential pressure between both feet, by adding a 1 kg mass at each pending wrist, enhanced the internal representation of body orientation and motion relative to a gravitational reference frame. The large decreased in alpha/beta power in the right posterior parietal cortex (and not in the left) is in line with this assumption. Finally, our behavioral analyses showed smaller body sway oscillations for patients, likely originated from a tactile-based control strategy. Conversely, healthy subjects showed smaller head oscillations suggesting a vestibular-based control strategy, the head serving as a reference for balance control.

HighlightsO_LISomatosensory cortex excitability is greater in patients with bilateral vestibular loss than in aged-matched healthy individuals
C_LIO_LITo control balance, healthy individuals "locked" the head while vestibular patients "locked" their pelvis
C_LIO_LIFor vestibular patients, increasing loading/unloading mechanism enhances the internal representation of body state in the posterior parietal cortex
C_LI


title: Microglial inflammatory activation paradoxically improves glucose tolerance during diet-induced obesity

abstract: Hypothalamic gliosis associated with high fat diet (HFD) feeding increases susceptibility to hyperphagia and weight gain, and is therefore presumed to promote obesity-associated consequences such as glucose intolerance as well. Nevertheless, the body weight-independent contribution of microglial activation to glucose regulation has not been determined. Here we show that reducing microglial NF-{kappa}B signaling via cell-specific IKK{beta} deletion exacerbates HFD-induced glucose intolerance and insulin resistance despite reducing body weight and adiposity. This effect was associated with reduced activity of hypothalamic glucose sensing neurons. Conversely, a genetic approach to increase microglial inflammatory activity improved glucose tolerance independently of diet in lean rodents. To avoid confounding effects due to chronic alterations to microglial signaling pathways from dietary or genetic interventions, we developed an inducible model of microglial activation using DREADD-based chemogenetics. Gq-coupled GPCR activation rapidly increased microglial calcium levels, cytokine gene expression, and morphological hallmarks of inflammatory activation. In both lean and obese rodents, chemogenetic microglial activation caused a marked improvement in glucose tolerance along with increased activation of hypothalamic glucose sensing neurons, effects abrogated by central blockade of TNF signaling. Thus, while diet-induced microglial activation promotes weight gain, it may also serve an adaptive function--to prevent the deterioration of glucose tolerance associated with obesity, an important consideration for immune-modulating metabolic therapies.


title: Prolonged dexamethasone exposure enhances zebrafish lateral-line regeneration but disrupts mitochondrial homeostasis and hair cell function

abstract: The synthetic glucocorticoid dexamethasone is commonly used to treat inner ear disorders. Previous work in larval zebrafish has shown that dexamethasone treatment enhances hair cell regeneration, yet dexamethasone has also been shown to inhibit regeneration of peripheral nerves after lesion. We therefore used the zebrafish model to determine the impact of dexamethasone treatment on lateral-line hair cells and primary afferents. To explore dexamethasone in the context of regeneration, we used copper sulfate (CuSO4) to induce hair cell loss and retraction of nerve terminals, and then allowed animals to recover in dexamethasone for 48 hours. Consistent with previously work, we observed significantly more regenerated hair cells in dexamethasone-treated larvae. Importantly, we found that the afferent processes beneath neuromasts also regenerated in the presence of dexamethasone and formed an appropriate number of synapses, indicating that innervation of hair cells was not inhibited by dexamethasone. In addition to regeneration, we also explored the effects of prolonged dexamethasone exposure on lateral-line homeostasis and function. Following dexamethasone treatment, we observed more negatively charged mitochondrial membrane potentials ({Delta}{Psi}m) in neuromast hair cells and supporting cells. Hair cells exposed to dexamethasone were also more vulnerable to neomycin-induced cell death. In response to a fluid-jet delivered saturating stimulus, calcium influx through hair cell mechanotransduction channels was significantly reduced, yet presynaptic calcium influx was unchanged. Cumulatively, these observations indicate that dexamethasone enhances hair cell regeneration in lateral-line neuromasts, yet also disrupts mitochondrial homeostasis, making hair cells more vulnerable to ototoxic insults and possibly impacting hair cell function.


title: TubULAR: Tracking deformations of dynamic tissues and interfaces in 3D

abstract: A common motif in biology is the arrangement of cells into tube-like sheets, which further transform into more complex forms. Traditionally, analysis of tissues dynamic surfaces has relied on inspecting static snapshots, live imaging of cross-sections, or tracking isolated cells in 3D. However, capturing the interplay between in-plane and out-of-plane behaviors requires following the full surface as it deforms and integrating cell-scale motions into collective, tissue-scale deformations. Here, we introduce an approach to build in toto maps of surface deformation, following tissue parcels in the material frame of reference. The Tube-like sUrface Lagrangian Analysis Resource ( TubULAR) provides an open-source MATLAB implementation whose functionality is accessible either as a standalone toolkit or as an extension of the ImSAnE package used in the developmental biology community. TubULAR provides a framework for linking in-plane and out-of-plane behaviors and decomposing complex deformation maps into elementary contributions defining physically meaningful signatures of motion. We underscore the power of our approach by analyzing shape change in the embryonic Drosophila midgut and beating zebrafish heart. In the midgut, our results link asymmetric constrictions to the increasing organ length and to patterned vorticity in the organ chambers. We then extract the dynamics of the beating zebrafish heart, constructing a mode-based description for the organs beat cycle. The method naturally generalizes to in vitro and synthetic systems, such as stem cell models of tube development and deforming phase-separated interfaces.

category: developmental biology