title: A novel fully-spliced accessory gene in equine lentivirus with distinct Rev-responsive element

abstract:
All lentiviruses encode an accessory protein Rev, whose main biological function is to mediate the nuclear export of unspliced and incompletely spliced viral transcripts by binding to a viral cis-acting element (termed the Rev-responsive element, RRE) that is located within env-encoding region. Equine infectious anemia virus (EIAV) is a member of the Lentivirus genus in the Retroviridae family, and is considered an important model for the study of lentivirus pathogenesis. Here, we identified a novel transcript from the EIAV genome that encodes a viral protein, named Mat, with unknown function. The transcript mat is fully spliced and comprises parts of the coding regions of MA and TM. Interestingly, the expression of Mat depends on Rev and the CRM1 pathway. Rev could specifically bind to Mat mRNA to promote its nuclear export. We further identified that the first exon of Mat mRNA, which is located within the Gag-encoding region, acts as an unreported RRE. Altogether, we identified a novel fully spliced transcript mat with an unusual RRE, which interacts with Rev for nuclear export through the CRM1 pathway. Our findings may help to expend the understanding of gene transcription and expression of lentivirus.

Author summaryIn lentiviruses, the nuclear export of viral transcripts is an important step in controlling viral gene expression. Generally, the unspliced and incompletely spliced (that is, intron-containing) transcripts are exported via the CRM1-dependent export pathway in a process mediated by the viral Rev protein by binding to the Rev-responsive element (RRE) located within the Env-coding region. However, the completely spliced (intronless) transcripts are exported via an endogenous cellular pathway which is Rev independent. Here we identified a novel intronless transcript from EIAV and demonstrated that it encodes a viral protein, termed Mat. Interestingly, we have determined that the expression of Mat depends on Rev, and identified that the first exon of Mat mRNA could specifically bind to Rev and be exported to the cytoplasm, which suggests that first exon of Mat mRNA is a second RRE of EIAV. These findings update the EIAV genome structure and highlight the diversification of post-transcriptional regulation patterns in EIAV, and provides important insights into the Rev-dependent nuclear export of completely spliced transcripts in lentiviruses.


title: Peptide location fingerprinting identifies species- and tissue-conserved structural remodelling of proteins as a consequence of ageing and disease

abstract: Extracellular matrix (ECM) in the intervertebral disc (IVD), lung and artery are thought to undergo the age-dependant accumulation of damage by chronic exposure to mechanisms such as reactive oxygen species, proteases and glycation. It is unknown whether this damage accumulation is species-dependant (via differing lifespans and hence cumulative exposures) or whether it can influence the progression of age-related diseases such as atherosclerosis. Peptide location fingerprinting (PLF) is a new proteomic analysis method, capable of the non-targeted identification of structure-associated changes within proteins. Here we applied PLF to publicly available ageing human IVD (outer annulus fibrosus), ageing mouse lung and human arterial atherosclerosis datasets and identified novel target proteins alongside common age-associated differences within protein structures which were conserved between tissue regions, organs, sexes and species and in age-related disease. We identify peptide yield differences across protein structures which coincide with biological regions, potentially reflecting the functional consequences of ageing or atherosclerosis for macromolecular assemblies (collagen VI and fibrin), enzyme/inhibitor activity (cathepsin B and alpha-2 macroglobulin), activation states (complement C3 and thrombin) and interaction states (laminins, perlecan, fibronectin, filamin-A, collagen XIV and apolipoprotein-B). Furthermore, we show that alpha-2 macroglobulin, prothrombin, collagen XIV and apolipoprotein-B all exhibit possible shared structural consequences in IVD ageing and arterial atherosclerosis, providing novel links between an age-related disease and intrinsic ageing. Crucially, we also demonstrate that fibronectin, laminin beta chains and filamin-A all exhibit conserved age-associated structural differences between mouse lung and human IVD, providing evidence that ECM, and their associating proteins, may be subjected to potentially similar mechanisms or consequences of ageing across species, irrespective of differences in lifespan and tissue function.

category: molecular biology


title: A Novel Cytoskeletal Action of Xylosides

abstract: Proteoglycan glycosaminoglycan (GAG) chains are attached to a serine residue in the protein through a linkage series of sugars, the first of which is xylose. Xylosides are chemicals which compete with the xylose at the enzyme xylosyl transferase to prevent the attachment of GAG chains to proteins. These compounds have been employed at concentrations in the millimolar range as tools to study the role of GAG chains in proteoglycan function. In the course of our studies with xylosides, we conducted a dose-response curve for xyloside actions on neural cells. To our surprise, we found that concentrations of xylosides in the nanomolar to micromolar range had major effects on cell morphology. These effects are due to changes in cytoskeletal dynamics. Concentrations of xylosides which were effective in altering morphology did not alter GAG chain synthesis rates, nor did they produce any changes in gene expression as determined by RNAseq of treated cells. These observations support a novel action of xylosides on neuronal cells.

category: cell biology


title: A subset of gut leukocytes have telomerase-dependent 'hyper-long' telomeres and require telomerase for function in zebrafish

abstract:
Telomerase, the enzyme capable of elongating telomeres, is usually restricted in human somatic cells, which contributes to progressive telomere shortening with cell-division and ageing. Using the zebrafish model, we show that subsets gut immune cells have telomerase-dependent "hyper-long" telomeres, which we identified as being predominantly macrophages and dendritics (mpeg1.1+ and cd45+mhcII+). Notably, mpeg+ macrophages have much longer telomeres in the gut than in their haematopoietic tissue of origin, suggesting that there is a gut-specific modulation of telomerase in these cells.

Moreover, we show that a subset of gut mpeg+ cells express telomerase (tert) in young WT zebrafish, but that the relative proportion of these cells decreases with ageing. Importantly, this is accompanied by telomere shortening and DNA damage responses with ageing and a telomerase-dependent decrease in expression of autophagy and immune activation markers. Finally, these telomerase-dependent molecular alterations are accompanied by impaired phagocytosis of E.coli and increased gut permeability in vivo. Together, our data show that limiting levels of telomerase lead to changes in gut immunity and gut permeability, which, together, are likely contributors to local and systemic tissue degeneration and increased susceptibility to infection with ageing.

GRAPHICAL ABSTRACT

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=143 SRC="FIGDIR/small/478480v1_ufig1.gif" ALT="Figure 1">
View larger version (21K):
org.highwire.dtl.DTLVardef@141a6caorg.highwire.dtl.DTLVardef@12d24baorg.highwire.dtl.DTLVardef@c64267org.highwire.dtl.DTLVardef@319449_HPS_FORMAT_FIGEXP  M_FIG C_FIG Our data show that limiting levels of telomerase contribute to alterations in gut immunity, namely increased apoptosis, decreased autophagy and immune activation of mpeg+ macrophages in the gut. This is accompanied by a decreased ability to clear pathogens and increased gut permeability. Together, these are likely contributors to local and systemic tissue degeneration and susceptibility to infection with ageing.


title: Circadian disruption enhances HSF1 signaling and tumorigenesis in Kras-driven lung cancer

abstract: Disrupted circadian rhythmicity is a prominent feature of modern society and has been designated as a probable carcinogen by the World Health Organization. However, the biological mechanisms that connect circadian disruption and cancer risk remain largely undefined. We demonstrate that exposure to chronic circadian disruption (chronic jetlag, CJL) increases tumor burden in a mouse model of KRAS-driven lung cancer. Molecular characterization of tumors and tumor-bearing lung tissues revealed that CJL enhances the expression of heat shock factor 1 (HSF1) target genes. Consistently, exposure to CJL disrupted the highly rhythmic nuclear trafficking of HSF1 in the lung, resulting in an enhanced accumulation of HSF1 in the nucleus. HSF1 has been shown to promote tumorigenesis in other systems, and we find that pharmacological inhibition of HSF1 reduces the growth of KRAS-mutant human lung cancer cells. These findings implicate HSF1 as a molecular link between circadian disruption and enhanced tumorigenesis.


title: Exploring the thermal limits of malaria transmission in the western Himalaya

abstract: Environmental temperature is a key driver of malaria transmission dynamics. Using detailed temperature records from four sites (1800-3200m) in the western Himalaya, we model how temperature regulates parasite development rate (the inverse of the extrinsic incubation period, EIP) in the wild. Using a Briere parametrization of the EIP, combined with Bayesian parameter inference, we study the thermal limits of transmission for avian (P. relictum) and human Plasmodium parasites (P. vivax and P. falciparum) as well as for two malaria-like avian parasites, Haemoproteus and Leucocytozoon. We demonstrate that temperature conditions can substantially alter the incubation period of parasites at high elevation sites (2600-3200m) leading to restricted parasite development or long transmission windows. We then compare estimates of EIP based on measures of mean temperature versus hourly temperatures to show that EIP days vary in cold versus warm environments. We found that human Plasmodium parasites experience a limited transmission window at 2600m. In contrast, for avian Plasmodium transmission was not possible between September to March at 2600m. In addition, temperature conditions suitable for both Haemoproteus and Leucocytozoon transmission were obtained from June to August and in April, at 2600m. Finally, we use temperature projections from a suite of climate models to predict that by 2040, high elevation sites (~ 2600 m) will have a temperature range conducive for malaria transmission, albeit with a limited transmission window. Our study highlights the importance of accounting for fine-scale thermal effects in the expansion of the range of the malaria parasite with global climate change.

category: ecology


title: Reproductive strategies in a complex of simultaneously hermaphroditic species, the Allolobophora chlorotica case study

abstract: An in-depth knowledge of reproductive strategies is essential to understand the evolutionary history of species and their resulting patterns of taxonomic diversity. In particular, the study of life history traits related to reproduction could help to resolve the speciation patterns in the cryptic species complexes recently found in earthworms. Here, we carried out a parentage analysis in such a complex, the Allolobophora chlorotica aggregate. Using four nuclear microsatellite markers and a fragment of the cytochrome c oxidase subunit I mitochondrial gene we investigated (i) the mating strategies between individuals belonging to two divergent mitochondrial lineages (L2 and L3) that cannot be distinguished with nuclear markers and (ii) the reproductive isolation between lineages that are differentiated both at the mitochondrial and nuclear level (L2/L3 and L1). Among the 157 field collected individuals, 66 adults were used in cross-breeding experiments to form 22 trios based on their assignment to a mitochondrial lineage, and 453 obtained juveniles were genotyped. We showed that adults that mated with both their potential mates in the trio produced significantly more juveniles. In L2 and L3 crosses, a sex-specific pattern of reproduction characteristic to each lineage was observed, suggesting a possible conflict of interest between mating partners. In L2/L3 and L1 crosses, a high production of cocoons was counterbalanced by a low hatching rate, suggesting a post-zygotic reproductive isolation. Reproductive strategies are thus likely to contributed to the diversification of this complex of species and we advocate further studies on sexual selection and sex allocation in earthworms.

category: evolutionary biology


title: Flagellar energy costs across the Tree of Life

abstract: Flagellar-driven motility grants unicellular organisms the ability to gather more food and avoid predators, but the energetic costs of construction and operation of flagella are considerable. Paths of flagellar evolution depend on the deviations between fitness gains and energy costs. Using structural data available for all three major flagellar types (bacterial, archaeal, and eukaryotic), flagellar construction costs were determined for Escherichia coli, Pyrococcus furiosus, and Chlamydomonas reinhardtii. Estimates of cell volumes, flagella numbers, and flagellum lengths from the literature, yield flagellar costs for another [~]200 species. The benefits of flagellar investment were analysed in terms of swimming speed, nutrient collection, and growth rate; showing, among other things, that the cost-effectiveness of bacterial and eukaryotic flagella follows a common trend. However, a comparison of whole cell costs and flagellum costs across the Tree of Life reveals that only cells with larger cell volumes than the typical bacterium could evolve the more expensive eukaryotic flagellum. These findings provide insight into the unsolved evolutionary question of why the three domains of life each carry their own type of flagellum.


title: Expanded Analysis of Pigmentation Genetics in UK Biobank

abstract: The genetics of pigmentation is an excellent model for understanding gene interactions in a trait almost entirely unaffected by environment. We have analysed pigmentation phenotypes in UK Biobank using DISSECT, a tool which enables genome-wide association studies (GWAS) whilst accounting for relatedness between individuals, and thus allows a much larger cohort to be studied. We have increased the number of candidate genes associated with red and blonde hair colour, basal skin colour and tanning response to UV radiation. As previously described, we find almost all red hair individuals have two variant MC1R alleles; exome sequence data expands the number of associated coding variants. Rare red-headed individuals with only a single MC1R variant are enriched for an associated eQTL at the ASIP gene. We find that females are most likely to self-report red or blonde hair, paler skin and less tanning ability than men, and that variants at KITLG, MC1R, OCA2 and IRF4 show significant sex differences in effect. After taking sex into account, pigmentation phenotypes are not correlated with sex hormone levels, except for tanning ability, which shows a positive correlation with testosterone in men. Across the UK there is a correlation between place of birth and hair colour; red hair being more common in the north and west, whilst blonde hair is more common in the east. Combining GWAS with transcriptome data to generate a transcriptome wide association study identifies candidate genes whose expression in skin or melanocytes shows association with pigmentation phenotypes. A comparison of candidates associated with different pigmentation phenotypes finds that candidates for blonde hair, but not skin colour, are enriched for skin and hair genes suggesting that it may be hair shape and structure that impacts hair colour, rather than the melanocyte/keratinocyte interaction.

category: genetics


title: The evolution of coordination: A phylogenetic meta-analysis and systematic review

abstract: To solve many cooperative problems, humans must have evolved the ability to solve physical problems in their environment by coordinating their actions. There have been many studies conducted across multiple different species regarding coordinating abilities. These studies aim to provide data which will help illuminate the evolutionary origins of cooperative problem solving and coordination. However, it is impossible to make firm conclusions about the evolutionary origins of coordinating abilities without a thorough comparative analysis of the existing data. Furthermore, there may be certain aspects of the literature that make it very difficult to confidently address evolutionary and meta-analytic questions. This study aimed to rectify this by using meta-analysis, phylogenetic analysis, and systematic review to analyse the data already obtained across multiple studies, and to assess the reliability of this data. We found that many studies did not provide the information necessary for meta analysis, or were not comparable enough to other studies to be included in analyses, meaning meta analyses were underpowered or could not be conducted due to low samples of both studies and different species. Overall, we found that many studies reported small positive effects across studies, but the standard errors of these effects frequently traversed zero.

category: animal behavior and cognition


title: Pseudotime analysis reveals novel regulatory factors for multigenic onset and monogenic transition of odorant receptor expression

abstract: During their maturation from horizontal basal stem cells, olfactory sensory neurons (OSNs) are known to select exactly one out of hundreds of olfactory receptors (ORs) and express it on their surface, a process called monogenic selection. Monogenic expression is preceded by a multigenic phase during which several OR genes are expressed in a single OSN. Here, we perform pseudotime analysis of a single cell RNA-Seq dataset of murine olfactory epithelium to precisely align the multigenic and monogenic expression phases with the cell types occurring during OSN differentiation. In combination with motif analysis of OR gene cluster-associated enhancer regions, we identify known and novel transcription (co-)factors (Ebf1, Lhx2, Ldb1, Fos and Sspp2) and chromatin remodelers (Kdm1a, Eed and Zmynd8) associated with OR expression. The inferred temporal order of their activity suggests novel mechanisms contributing to multigenic OR expression and monogenic selection.

category: bioinformatics


title: Evaluation of PRKACA as a Therapeutic Target for Fibrolamellar Carcinoma

abstract:
Background & AimsFibrolamellar carcinoma (FLC) is a rare, difficult-to-treat liver cancer primarily affecting pediatric and adolescent patients, and for which precision medicine approaches have historically not been possible. The DNAJB1-PRKACA gene fusion was identified as a driver of FLC pathogenesis. We aimed to assess whether FLC tumors maintain dependency on this gene fusion and determine if PRKACA is a viable therapeutic target.

MethodsFLC patient-derived xenograft (PDX) shRNA cell lines were implanted subcutaneously into female NOD-SCID mice and tumors were allowed to develop prior to randomization to doxycycline (to induce knockdown) or control groups. Tumor development was assessed every 2 days. To assess the effect of treatment with novel selective PRKACA small molecule kinase inhibitors, BLU0588 and BLU2864, FLC PDX tumor cells were implanted subcutaneously into NOD-SCID mice and tumors allowed to develop. Mice were randomized to treatment (BLU0588 and BLU2864, orally, once daily) or control groups and tumor size determined as above.

ResultsKnockdown of DNAJB1-PRKACA reversed a FLC-specific gene signature and reduced PDX tumor growth in mice compared to the control group. Furthermore, FLC PDX tumor growth was significantly reduced with BLU0588 and BLU2864 treatment versus control (P = 0.003 and P = 0.0005, respectively).

ConclusionsWe demonstrated, using an inducible knockdown and small molecule approaches, that FLC PDX tumors were dependent upon DNAJB1-PRKACA fusion activity. In addition, this study serves as a proof-of-concept that PRKACA is a viable therapeutic target for FLC and warrants further investigation.

category: cancer biology


title: In vitro direct regeneration and Agrobacterium tumefaciens mediated in planta transformation of Ocimum sanctum L.

abstract: Ocimum sanctum is a multipurpose herb with highly significant medicinal properties. An in vitro direct regeneration protocol for propagation of a valuable medicinal plant Ocimum sanctum, using petiole explants has been successfully developed. The protocol employed regeneration of shoots directly, without any intervening callus using Murashige and Skoog (MS) medium fortified with 3 mg L-1 BAP + 1 mg L-1 NAA. The maximum regeneration frequency of 98% with 9.6 shoots per explants was achieved. Agrobacterium tumefaciens mediated genetic transformation (ATMT) protocol (transient and stable) was established using LBA4404 strain harboring pBI121 with uid-A reporter gene and neomycin phosphotransferase (npt-II) as selection marker. The putative transformants were screened on MS with 50 mg L-1 kanamycin and subsequently rooted on the half-strength MS medium. The confirmation was done via polymerase chain reaction (PCR) using npt-II and gus-A gene-specific primers. The maximum stable transformation frequency 70% {+/-} 0.35. Thus, it is apparent that the established in vitro direct regeneration and ATMT method was suitable for integrating novel genes and modulating the metabolic flux for obtaining desired agronomic trait in planta.

category: plant biology


title: CRISPR/Cas9 gene editing uncovers the role of CTR1 and ROS1 in melon fruit ripening and epigenetic regulation

abstract: Melon (Cucumis melo L.) has emerged as an alternative model to study fruit ripening due to the coexistence of climacteric and non-climacteric varieties. The previous characterization of a major QTL ETHQV8.1 sufficient to trigger climacteric ripening in a non-climacteric background allowed the identification within the QTL interval of a negative regulator of ripening CmCTR1-like (MELO3C024518), and a putative DNA demethylase CmROS1 (MELO3C024516), the orthologue of DML2, a DNA demethylase regulating fruit ripening in tomato. To understand the role of these genes in climacteric ripening, we generated homozygous CRISPR knockout mutants of CmCTR1-like and CmROS1 in a climacteric genetic background. The climacteric behavior was altered in both loss-of-function mutants in two summer seasons with an advanced ethylene production profile compared to the climacteric wild type, suggesting a role of both genes in climacteric ripening in melon. Single cytosine methylome analyses of the CmROS1 knockout mutant revealed DNA methylation changes in the promoter regions of key ripening genes as ACS1, ETR1 and ACO1, and ripening associated-transcription factors as NAC-NOR, RIN and CNR, suggesting the importance of CmROS1-mediated DNA demethylation for triggering fruit ripening in melon.


title: Common signatures of differential microRNA expression in Parkinson's and Alzheimer's disease brains

abstract:
BackgroundDysregulation of microRNA (miRNA)-mediated gene expression has been implicated in the pathogenesis and course of many neurodegenerative diseases including Parkinsons disease (PD). However, the functionally relevant miRNAs remain largely unknown. Previous meta-analyses on differential miRNA expression data in post-mortem PD brains have highlighted several miRNAs showing consistent and statistically significant effects. However, these meta-analyses were based on exceedingly small sample sizes.

MethodsIn this study, we quantified the expression of the four most compelling PD candidate miRNAs from these meta-analyses in the superior temporal gyrus (STG) of one of the largest case-control post-mortem brain datasets available (261 samples), thereby quadruplicating previously investigated sample sizes. Furthermore, we probed for common differential miRNA expression signatures with Alzheimers disease (AD) by also analyzing these miRNAs in post-mortem STG of 190 AD patients and controls and by testing six top AD miRNAs in the PD brains.

ResultsOf all ten analyzed miRNAs, PD candidate miRNA homo sapiens (hsa-) miR-132-3p showed evidence for differential expression in both PD (p=4.89E-06) and AD (p=3.20E-24), and AD miRNAs hsa-miR-132-5p (p=4.52E-06) and hsa-miR-129-5p (p=0.0379) showed evidence for differential expression in PD. Combining these novel data with previously published data substantially improved the statistical support (=3.85E-03 using Bonferroni correction) of the corresponding meta-analyses clearly and compellingly implicating these miRNAs in both PD and AD. Furthermore, hsa-miR-132-3p/-5p (but not hsa-miR-129-5p) showed association with neuropathological Braak PD staging (p=3.51E-03/p=0.0117), suggesting that these miRNAs may play a role in -synuclein aggregation beyond the early disease phase.

ConclusionsOur study represents the largest independent assessment of recently highlighted candidate brain miRNAs in PD and AD post-mortem brain samples, to date. Our results implicate hsa-miR-132-3p/-5p and hsa-miR-129-5p to be differentially expressed in both PD and AD brains, potentially pinpointing shared pathogenic mechanisms across these neurodegenerative diseases.

category: neuroscience


title: Orally active mGluR2/3 metabotropic antagonist pro-drug mimics the beneficial effects of physical exercise on neurogenesis, behavior, and exercise-related molecular pathways in an Alzheimer's disease mouse model

abstract: Modulation of physical activity represents an important intervention that may delay, slow, or prevent mild cognitive impairment (MCI) or dementia due to Alzheimers disease (AD). One mechanism proposed to underlie the beneficial effect of physical exercise involves the apparent stimulation of adult hippocampal neurogenesis (AHN). BCI-838 is a pro-drug whose active metabolite BCI-632 is an antagonist at the group II metabotropic glutamate receptor (mGluR2/3). We previously demonstrated that administration of BCI-838 to a mouse model of cerebrovascular accumulation of oligomeric A{beta}E22Q (APPE693Q = "Dutch APP") reduced learning behavior impairment and anxiety, both of which are associated with the phenotype of the Dutch APP mice. Here we show that (i) administration of BCI-838, physical exercise, or a combination of BCI-838 and physical exercise enhanced AHN in a four-month old mouse model of AD amyloid pathology (APPKM670/671NL/ PSEN1{Delta}exon9 = APP/PS1), (ii) administration of BCI-838 alone or associated with physical exercise led to stimulation of AHN and improvement in both spatial and recognition memory, (iii) significantly, the hippocampal dentate gyrus transcriptome of APP/PS1 mice following BCI-838 treatment up-regulated brain-derived neurotrophic factor (BDNF), PIK3C2A of the PI3K-MTOR pathway, and metabotropic glutamate receptors, and down-regulated EIF5A of ketamine-modulating mTOR activity, and finally (iv) qPCR findings validate a significantly strong association between increased BDNF levels and BCI-838 treatment. Our study points to BCI-838 as a safe and orally active compound capable of mimicking the beneficial effect of exercise on AHN, learning behavior, and anxiety in a mouse model of AD neuropathology.


title: Orexin action on the dopaminergic system modulates theta during REM sleep and wakefulness

abstract: Both dopaminergic (DA) and orexinergic (OX) systems establish brain-wide neuromodulatory circuits that profoundly influence brain states and behavioral outputs. To unravel their interactions, we inactivated OX-to-DA neurotransmission by selective disruption of HcrtR1/OxR1, or HcrtR2/OxR2, or both receptors, in DA neurons. Chronic loss of OXR2 in DA neurons (OxR2Dat-CKO mice) dramatically increased electrocorticographic (EcoG) theta rhythms in wakefulness and REM sleep. Episode duration and total times spent in  active wakefulness and REMS were prolonged, and theta/fast-gamma wave coupling was enhanced in both states. Increased theta in OxR2DatCKO mice baseline wake was accompanied by diminished infra-theta and increased fast-gamma activities, i.e. the mice exhibited signs of constitutive electrocortical hyperarousal, albeit uncoupled with locomotor activity. These effects were not seen in OxR1-ablated dopaminergic mutants, which tended to show opposite phenotypes, resembling those caused by the loss of both receptors. Our data establish a clear, genetically-defined link between monosynaptic orexin-to-dopaminergic connectivity and the power of theta oscillations, with a differential role of OXR2 in cross-frequency wave coupling and attentional processes.


title: Intervertebral disc cell chondroptosis elicits neutrophil response in Staphylococcus aureus spondylodiscitis

abstract:
ObjectiveTo understand the pathophysiology of spondylodiscitis due to Staphylococcus aureus, an emerging infectious disease of the intervertebral disc (IVD) and vertebral body with a high complication rate, by combining clinical insights and experimental approaches.

DesignClinical data and histological material of nine patients suffering from S. aureus spondylodiscitis were retrospectively collected at a single center. To mirror the clinical findings experimentally, we developed a novel porcine ex vivo model mimicking acute S. aureus spondylodiscitis and assessed the interaction between S. aureus and IVD cells within their native environment. In addition, the inflammatory features underlying this interaction were assessed in primary human IVD cells. Finally, mirroring the clinical findings, we assessed primary human neutrophils for their ability to respond to secreted inflammatory modulators of IVD cells upon S. aureus challenge.

ResultsAcute S. aureus spondylodiscitis in patients was characterized by tissue necrosis and neutrophil infiltration. Additionally, the presence of empty IVD cells lacunae was observed. This was mirrored in the ex vivo porcine model, where S. aureus induced extensive IVD cell death, leading to empty lacunae. Concomitant engagement of the apoptotic and pyroptotic cell death pathways was observed in primary human IVD cells, resulting in cytokine release. Among the released cytokines, functionally intact neutrophil-priming as well as broad pro- and anti-inflammatory cytokines known for their involvement in IVD degeneration were found.

ConclusionsIn patients as well as ex vivo in a novel porcine model, S. aureus spondylodiscitis infection caused IVD cell death, resulting in empty lacunae, which was accompanied by release of inflammation markers and recruitment of neutrophils. These findings offer valuable insights into the important role of inflammatory IVD cell death during the onset of spondylodiscitis and potential future therapeutic approaches.

category: immunology


title: Neuronal RNA granules are ribosome complexes stalled at the pre-translocation state

abstract: The polarized cell morphology of neurons dictates many neuronal processes, including the axodendridic transport of specific mRNAs and subsequent translation. mRNAs together with ribosomes and RNA-binding proteins form RNA granules that are targeted to axodendrites for localized translation in neurons. It has been established that localized protein synthesis in neurons is essential for long-term memory formation, synaptic plasticity, and neurodegeneration. We have used proteomics and electron microscopy to characterize neuronal RNA granules (nRNAg) isolated from rat brain tissues or human neuroblastoma. We show that ribosome containing RNA granules are morula-like structures when visualized by electron microscopy. Crosslinking-coupled mass-spectrometry identified potential G3BP2 binding site on the ribosome near the eIF3d-binding site on the 40S ribosomal subunit. We used cryo-EM to resolve the structure of the ribosome-component of nRNAg. The cryo-EM reveals that ribosomes in the nRNAg are stalled at the elongation state where tRNAs are in the hybrid A/P and P/E site, and resemble the pre-translocation state ribosomes. We also describe a new kind of principal motion of the ribosome, which we call the rocking motion.


title: Domain expansion and functional diversification in vertebrate reproductive proteins

abstract: The rapid evolution of fertilization proteins can result in remarkable diversity in their structure and function. Many of the proteins in vertebrate egg coats contain copies (1-6) of the ZP-N domain. These ZP-N domains can facilitate multiple reproduction functions, including species-specific sperm recognition. We integrated phylogenetics and machine learning to investigate how ZP-N domains diversified in structure and function. The most C-terminal ZP-N domain of each paralog is associated with another domain type (ZP-C) which together form a "ZP module." All modular ZP-N domains were phylogenetically distinct from non-modular or free ZP-N domains. Machine learning-based classification identified 8 residues that form a stabilizing network in modular ZP-N domains that is absent in free domains. Positive selection was identified in some free ZP-N domains. Our findings suggest that purifying selection has conserved an essential structural core in modular ZP-N domains, while free N-terminal domains have been able to experience functionally diversify.


title: Selective inhibition of protein secretion by abrogating receptor-coat interactions during ER export.

abstract: Protein secretion is an essential cellular process that permits cell growth, movement and communication. Traffic of proteins within the eukaryotic secretory pathway is mediated by transport intermediates that bud from one compartment, populated with appropriate cargo proteins, and fuse with a downstream compartment to deliver their contents. Here, we explore the possibility that protein secretion can be selectively inhibited by perturbing protein-protein interactions that drive capture into transport vesicles. Human PCSK9 is a major determinant of cholesterol metabolism, whose secretion is mediated by a specific cargo adaptor of the ER export machinery, SEC24A. We map a series of protein-protein interactions between PCSK9, its ER export receptor, SURF4, and SEC24A, that mediate secretion of PCSK9. We show that the interaction between SURF4 and SEC24A can be inhibited by 4-PBA, a small molecule that occludes a cargo-binding domain of SEC24. This inhibition reduces secretion of PCSK9 and additional SURF4 clients that we identify by mass spectrometry, leaving other secreted cargoes unaffected. We propose that selective small molecule inhibition of cargo recognition by SEC24 is a potential therapeutic intervention for atherosclerosis and other diseases that are modulated by secreted proteins.


title: Revisiting degron motifs in human AURKA required for its targeting by APC/C-FZR1

abstract:
Mitotic kinase Aurora A (AURKA) diverges from other kinases in its multiple active conformations that may explain its interphase roles and association with cancer, and the limited efficacy of drugs targeting the kinase pocket. Regulation of AURKA activity by the cell is critically dependent on destruction mediated by the Anaphase-Promoting Complex (APC/CFZR1) during mitotic exit and G1 phase and requires an atypical N-terminal degron in AURKA called the  A-box in addition to a reported canonical D-box degron in the C-terminus. Here we find that the proposed C-terminal D-box of AURKA does not act as a degron and instead mediates essential structural features of the protein. In living cells, as previously reported in vitro, the N-terminal intrinsically disordered region (IDR) of AURKA containing the A-box is sufficient to confer FZR1-dependent mitotic degradation. Both in silico and in cellulo assays predict the QRVL Short Linear Interacting Motif (SLiM) of the A-box to be a phospho-regulated D-box. We propose that degradation of full-length AURKA additionally depends on an intact C-terminal domain because of critical conformational parameters permissive for both activity and mitotic degradation of AURKA.

Summary blurbAURKA degron motifs are redefined to show that the so-called N-terminal  A-box is in fact a D-box, and the so-called  D-box in the C-terminus is not a degron but a motif critical for the active, degradable conformation of AURKA


title: A cross taxonomic comparison of bird and butterfly communities of Tamhini Wildlife Sanctuary across two decades.

abstract: Human disturbance can alter the structure and function of ecological communities. We studied bird and butterfly communities of Tamhini wildlife sanctuary to understand the effects two decades of changing land use and management. We replicated a previous study conducted between 1998 and 2001, sampling 7 line transects every fortnight between April 2016 and April 2017. We analyzed the diversity and structure of functional groups and compared our findings to previous surveys. Species diversity increased for both taxa, however, community composition was significantly different across studies. Changing land may affect multiple trophic levels and alter community assembly and ecosystem function.


title: Exploring the possible role of hybridization in the evolution of photosynthetic pathways in Flaveria (Asteraceae), the prime model of C4 photosynthesis evolution

abstract: Flaveria (Asteraceae) is the prime model for the study of C4 photosynthesis evolution and seems to support a stepwise acquisition of the pathway through C3-C4 intermediate phenotypes, still existing in Flaveria today. Molecular phylogenies of Flaveria based on concatenated data matrices are currently used to reconstruct the complex sequence of trait shifts during C4 evolution. To assess the possible role of hybridization in C4 evolution in Flaveria, we re-analyzed transcriptome data of 17 Flaveria species to infer the extent of gene tree discordance and possible reticulation events. We found massive gene tree discordance as well as reticulation along the backbone and within clades containing C3-C4 intermediate and C4-like species. An early hybridization event between two C3 species might have triggered C4 evolution in the genus. The clade containing all C4 species plus the C4-like species F. vaginata and F. palmeri is robust but of hybrid origin involving F. angustifolia and F. sonorensis (both C3-C4 intermediate) as parental lineages. Hybridization seems to be a driver of C4 evolution in Flaveria and likely promoted the fast acquisition of C4 traits. This new insight can be used in further exploring C4 evolution and can inform C4 bioengineering efforts.


title: Non-canonical scaffold-type ligase complex mediates protein UFMylation

abstract: Protein UFMylation is emerging as a posttranslational modification essential for endoplasmic reticulum and cellular homeostasis. Despite its biological importance, we have a poor understanding of how UFM1 is conjugated onto substrates. Here, we use a rebuilding approach to define the minimal requirements of protein UFMylation. We find that the reported E3 ligase UFL1 is inactive on its own and identify UFBP1 to bind UFL1 to form an active E3 ligase complex. While UFC1 is an intrinsically Cys-reactive E2, we do not identify any catalytic cysteines on UFL1/UFBP1, suggesting a scaffold-type E3 ligase mechanism. Interestingly, the E3 ligase complex consists of winged-helix (WH) domain repeats that activate UFC1 for aminolysis. We identify the adaptor protein CDK5RAP3 to bind to and regulate E3 ligase activity potentially by preventing off-target UFMylation. In summary, our work identifies the minimal requirements for UFMylation and reveals regulatory principles of this atypical E3 ligase complex.

category: biochemistry


title: Non canonical scaffold-type ligase complex mediates protein UFMylation


title: Extracellular vesicle microRNA cargo engineering reveals critical mechanisms underlying therapeutic efficacy

abstract:
BackgroundExtracellular vesicles (EVs) are key mediators of intercellular communication and function to transfer biological cargo, including microRNA (miR), from donor to recipient cells. EVs isolated from cardiosphere-derived cells (CDCs) have demonstrated therapeutic efficacy in pre-clinical models of ischemic heart disease, highlighting them as promising vectors for the treatment of CVD. Importantly, it has not yet been established whether miR cargo is necessary for the observed therapeutic benefit of CDC-EVs following acute MI (AMI).

MethodsCDCs were transfected with siRNA against Drosha, the initial endonuclease in the miRNA biogenesis pathway, to generate miR depleted DROSHA-EVs. EVs were characterized by size, morphology, and protein/miR expression. The role of EV miRNA on cardiac target cell apoptosis, proliferation and angiogenesis was examined using a series of in vitro assays. Mice with acute MI underwent delivery of human CDC EVs, DROSHA-EVs and placebo in a double-blind study. LVEF was assessed by echo at 1- and 28-days post-MI and tissue samples processed for assessment of histological endpoints. In vitro sufficiency assays were performed using a combinatorial approach with individual candidate miRs to identify clusters exhibiting synergistic efficacy.

ResultsDROSHA-EVs exhibited global downregulation of miRNA cargo but were otherwise indistinguishable from wild-type CDC-EVs. miR cargo was responsible for mediating the beneficial effects of human CDC-EV treatment on cardiomyocyte apoptosis, fibroblast proliferation and angiogenesis in vitro. DROSHA-EVs were unable to promote recovery following AMI on a functional or histological level, highlighting the critical role of EV miRNAs in cardioprotection following ischemic injury. A potentially therapeutic miR cluster, miR-146a-370-126a, was identified which acted synergistically to reduce cardiomyocyte apoptosis and was sufficient to render inert EVs into therapeutic vectors.

ConclusionsThese results demonstrate for the first time that miRNAs are required for the regenerative potential of CDC-EVs following AMI and identify a novel miR cluster with therapeutic implications.

category: bioengineering


title: Constructing Causal Cytokine Networks for Autoimmune and Inflammatory Diseases Target Identification

abstract: Cytokines are cell signaling proteins that act as inducers of physiological responses including the activation and differentiation of innate and adaptive immune cell types. In some cases, this mediates the initiation or progression of inflammatory and auto-immune diseases such as septic shock, rheumatoid arthritis, psoriasis, and inflammatory bowel disease. Many cytokines are pleiotropic, meaning that they can have effects on multiple cell types and play several distinct functions. On the other hand, multiple cytokines might have similar function and be redundant or degenerate. Some cytokines also regulate the expression and functions of other cytokines. Such complex cause-effect relationships can be modeled computationally by building "cytokine networks". Those networks describe intricate interactions between different cytokines in their associated tissue environment and cell types. Here we propose a computational approach to find sensitive and "causal" nodes within a cytokine interaction network by identifying whether persistent positive feedback (self-loop) exists on that cytokine in its mRNA transcription network. Based upon two microarray gene expression datasets from immune stimulation experiments using up to 15 different cytokines, we identified TNF as a "causal" cytokine in the colonic epithelial cell line HCT8, and IL-23, IL-1{beta} and TGF{beta} as "causal" cytokines in primary myofibroblasts. We hypothesize that a cytokine with a persistent positive feedback loop that amplifies its transcription and secretion could be identified as "causal" for the transcriptional regulation of other cytokines and potentially, downstream disease phenotypes. The existence of such positive feedback loop may determine whether targeting suppression of such cytokine will result in clinical efficacy.


title: Adaptive scans allow targeted cell-ablations on curved cell sheets

abstract: Tissue morphogenesis proceeds through the mechanical actuation of cells by active forces from the cytoskeleton. Multicellular laser ablation has emerged as an essential means to probe these active forces by severing the tissue locally and inferring the tension born by the severed domain from the recoil of the surrounding structures. The realm of applications of laser ablations is however limited by the 2D steering of the laser in most instrumental configurations, while embryos and developing tissues are intrinsically 3D structures. Here, we present a flexible near infrared (NIR) fs-pulsed laser ablation system in which ablation trajectories proceed in 3D and adapt to the curved surface of cell sheets, which are prominent structures in embryos. Trajectories are computed through an unsupervised search for the surface of interest. We demonstrate that, depending on the exact experimental setup, the surface estimation can rely on a high content 3D imaging with a combined confocal microscope, or alternatively on a rapid Lissajou scan of the sample space with a NIR stand-alone setup. We apply the developed strategy mapping tensions in the developing Drosophila wing imaginal disc. These targeted, adaptive scans could be applied to other forms of non-linear processes such as two-photon fluorescence imaging or opto-genetics.

category: biophysics


title: Guiding Irregular Nuclear Morphology on Nanopillar Array for Malignancy Differentiation in Tumor cells

abstract:
For more than a century, abnormal nuclei in tumor cells, presenting subnuclear invaginations and folds on the nuclear envelope, have been known to be associated with high malignancy and poor prognosis. However, current nuclear morphology analysis focuses on the features of the entire nucleus, overlooking the malignancy-related subnuclear features in nanometer scale. The main technical challenge is to probe such tiny and randomly distributed features inside cells. We here employ nanopillar arrays to guide subnuclear features into ordered patterns enabling their quantification as a strong indicator of cell malignancy. Both breast and liver cancer cells were validated, as well as the quantification of nuclear abnormality heterogeneity. The alterations of subnuclear patterns were also explored as effective readouts for drug treatment. We envision this nanopillar-enabled quantification of subnuclear abnormal features in tumor cells opens a new angle in characterizing malignant cells and studying the unique nuclear biology in cancer.

TeaserA nanopillar-based assay quantifying the abnormal nuclear morphology in tumor cells at single-cell level.


title: In-vivo egfp expression in the honeybee Apis mellifera induced by electroporation and viral expression vector.

abstract: In this study we describe egfp expression induced by two techniques: in vivo electroporation and viral transduction in several cell types of the adult honeybee brain. Non-neuronal and neuronal cell types were identified and the expression persisted at least during three days. Kenyon cells, optic lobe neurons and protocerebral lobe neurons were electroporated. Astrocyte-like glia cells, fibrous lamellar glia cells and cortex glia cells were identified. Viral transduction targeted one specific type of glia cells that could not be identified. EGFP positive cells were rather variable after electroporation, and viral transduction resulted in more homogenous groups of positive cells. We propose that these techniques remain a good alternative to transgenic animals because they potentially target only somatic cells.


title: Analysis of secondary metabolite gene clusters and chitin biosynthesis pathways of Monascus purpureus with high production of pigment and citrinin based on whole-genome sequencing

abstract: Monascus is a filamentous fungus that is commonly used for producing Monascus pigments in the food industry in Southeast Asia. While the development of bioinformatics has helped elucidate the molecular mechanism underlying secondary metabolite biosynthesis of Monascus, the biological information on the metabolic engineering of Monascus morphology remains unclear. In this study, we sequenced the whole genome of Monascus purpureus CSU-M183 strain by using combined single-molecule real-time DNA sequencing and next-generation sequencing platforms. The length of the genome assembly was 23.75 Mb in size with a GC content of 49.13% and 69 genomic contigs and encoded 7305 putative predicted genes. Furthermore, we identified secondary metabolite biosynthetic gene clusters and chitin synthesis pathways in the genome of the high pigment-producing M. purpureus CSU-M183 strain. And we confirmed that atmospheric room temperature plasma induced significant expression of the genes on Monascus pigments and citrinin biosynthetic gene cluster in M. purpureus CSU-M183 by RT-qPCR. These results provide a basis for understanding the secondary metabolite biosynthesis, the regulatory mechanisms of Monascus morphology, disrupting secondary metabolite biosynthesis in submerged fermentation, and the metabolic engineering of Monascus morphology.

category: microbiology


title: Single-cell transcriptomics of dynamic cell behaviors

abstract: Despite advances in spatial transcriptomics, the molecular profiling of dynamic behaviors of cells in their native environment remains a major challenge. We present a method, termed behavioral transcriptomics, that allows us to couple physiological behaviors of single cells in an intact tissue to deep molecular profiling of individual cells. This method enabled us to establish a novel molecular signature for a striking migratory cellular behavior following tissue injury.


title: Single-cell atlas of epithelial and stromal cell heterogeneity by lobe and strain in the mouse prostate

abstract: Evaluating the complex interplay of cell types in the tissue microenvironment is critical to understanding the origin and progression of diseases in the prostate and potential opportunities for intervention. Mouse models are an essential tool to investigate the molecular and cell-type-specific contributions of prostate disease at an organismal level. While there are well-documented differences in the extent, timing, and nature of disease development in various genetically engineered mouse models in different mouse strains and prostate lobes within each mouse strain, yet, the underlying molecular phenotypic differences in cell types across mouse strains and prostate lobes are incompletely understood. To address this, we examined the single-cell transcriptomes of individual mouse prostate lobes from two commonly used mouse strains, FVB/NJ and C57BL/6J. Data dimensionality reduction and clustering analysis revealed that basal and luminal cells possessed strain-specific transcriptomic differences, with luminal cells also displaying marked lobe-specific differences. Additionally, three rare populations of epithelial cells clustered independently of strain and lobe: one population of luminal cells expressing Foxi1 and components of the vacuolar ATPase proton pump (Atp6v0d2 and Atp6v1g3), another population expressing Psca and other stem cell-associated genes (Ly6a/Sca-1, Tacstd2/Trop-2), and a neuroendocrine population expressing Chga, Chgb, and Syp. In contrast, stromal cell clusters, including fibroblasts, smooth muscle cells, endothelial cells, pericytes, and immune cell types, were conserved across strain and lobe, clustering largely by cell type and not by strain or lobe. One notable exception to this was the identification of two distinct fibroblast populations that we term subglandular fibroblasts and interstitial fibroblasts based on their strikingly distinct spatial distribution in the mouse prostate. Altogether, these data provide a practical reference of the transcriptional profiles of mouse prostate from two commonly used mouse strains and across all four prostate lobes.


title: Three-dimensional analysis and in vivo imaging for sperm release and transport in the murine seminiferous tubule

abstract:
IntroductionSpermatozoa released from Sertoli cells must be transported to the epididymis. However, the contribution of the peristaltic motion in the seminiferous tubule to sperm release and transport remains unclear. We, therefore, investigated luminal flow and movements in the seminiferous tubules by three-dimensional analysis and in vivo imaging.

Materials and MethodsSerial testicular sections were cut in 5-m-thick and 50-m-interval and stained by PAS-hematoxylin. After the three-dimensional reconstruction of the seminiferous tubules, the localization of the flowing spermatozoa and stages observed in the sections were recorded in each reconstructed tubule. The luminal movements in the seminiferous tubule were observed by in vivo imaging using a fluorescent-reporter mouse and two-photon excitation microscopy system.

ResultsFlowing spermatozoa were mainly scattered in the lumina at stage VII/VIII, and clustered spermatozoa were also found in some regions. The clustered spermatozoa were observed at zero to two regions in each seminiferous tubule. Flowing spermatozoa were also found in the opposite direction to the rete testis. The flagellum direction of the spermatozoa attached to the seminiferous epithelium was reversed within a few seconds to a few tens of seconds when observed by in vivo imaging. The epithelium at the inner curve of the seminiferous tubule moved more actively and attached fewer spermatozoa compared to that at the outer curve.

DiscussionThis study revealed the presence of repeatedly reversed luminal flow in the seminiferous tubule. Such movements are suggested to help the sperm release from the Sertoli cells and the following aggregation of the released spermatozoa.


title: Local IL-17 orchestrates skin aging

abstract: Skin aging is characterized by structural and functional changes that lead to slower wound healing and higher rate of infections, which contribute to age-associated frailty. This likely depends on synergy between alterations in the local microenvironment and stem cell-intrinsic changes, underscored by pro-inflammatory microenvironments that drive pleotropic changes. To date, little is known about the precise nature and origin of the proposed age-associated inflammatory cues, or how they affect different tissue resident cell types. Based on deep single-cell RNA-sequencing of the entire dermal compartment, we now provide a comprehensive understanding of the age-associated changes in all skin cell types. We show a previously unreported skew towards an IL-17-expressing phenotype of Th cells, {gamma}{delta} T cells and innate lymphoid cells in aged skin. Aberrant IL-17 signaling is common to many autoimmune (e.g., rheumatoid arthritis and psoriasis) and chronic inflammatory diseases. Importantly, in vivo blockade of IL-17-triggered signaling during the aging process reduces the pro-inflammatory state by affecting immune and non-immune skin cells of both dermis and epidermis. Strikingly, IL-17 neutralization significantly delays the appearance of age-related traits, such as decreased epidermal thickness, increased cornified layer thickness and ameliorated hair follicle stem cell activation and hair shaft regeneration. Our results indicate that the aged skin shows chronic and persistent signs of inflammation, and that age-associated increased IL-17 signaling could be targeted as a strategy to prevent age-associated skin ailments in elderly.


title: The Growth Hormone Releasing Hormone Signaling Pathway Governs Cardiomyocyte differentiation in human iPS cells.

abstract: Differentiation and expansion of cardiomyocytes from human induced pluripotent stem cells is a poorly understood process, and the signals promoting differentiation of cardiomyocytes toward specific cell fates are not well defined. Here we show that the growth hormone-releasing hormone receptor (GHRH-R) signaling pathway controls this differentiation process and cell fate. Stimulation of hPSCs with GHRH promotes {beta}-catenin stabilization through the GH/IGF1 axis. The presence of cell-surface GHRH-R differentiates between atrio-ventricular cardiomyocytes and non-cardiomyogenic cardiac cells, including coronary vascular, neuronal and pacemaker cells, such that selecting for GHRH-R allows for the elimination of pacemaker cells and derivation of a pure population of atrio-ventricular cardiomyocytes. Unlike SIRPA, cardiac GHRH-R identifies murine and human NKX2-5+ myocardial derivatives of both mesodermal and neuroectodermal lineages. Using RNA-seq to comprehensively characterize the GHRH-R signaling pathway, we identified that NKX2-5, a transcription factor involved in progenitor specification and differentiation into atrio-ventricular cardiomyocytes, is upregulated in response to GHRH. GHRH-R signaling also upregulated HIF-1 via the PI3k/Akt/mTOR pathway to enhance NKX2-5 transcription and stimulate a Warburg-like effect in developing cardiomyocytes. Together these findings provide crucial insights into human stem cell differentiation and offer a translationally useful tool to optimize the development of pure cardiomyocyte preparations derived from induced pluripotent stem cells.


title: Oo-site: A dashboard to visualize gene expression during Drosophila oogenesis reveals meiotic entry is regulated post-transcriptionally

abstract: Determining how stem cell differentiation is controlled has important implications for understanding the etiology of degenerative disease and designing regenerative therapies. In vivo analyses of stem cell model systems have revealed regulatory paradigms for stem cell self-renewal and differentiation. The germarium of the female Drosophila gonad, which houses both germline and somatic stem cells, is one such model system. Bulk mRNA sequencing (RNA-seq), single-cell (sc) RNA-seq, and bulk translation efficiency of mRNAs are available for stem cells and their differentiating progeny within the Drosophila germarium. However, visualizing those data is hampered by the lack of a tool to spatially map gene expression and translational data in the germarium. Here, we have developed Oo-site (https://www.ranganlab.com/Oo-site), a tool for visualizing bulk RNA-seq, scRNA-seq, and translational efficiency data during different stages of germline differentiation, that makes these data accessible to non-bioinformaticians. Using this tool, we recapitulated previously reported expression patterns of developmentally regulated genes and discovered that meiotic genes, such as those that regulate the synaptonemal complex, are regulated at the level of translation.

category: developmental biology


title: Fungal community dynamics and carbon mineralization in coarse woody debris across decay stage, tree species, and stand development stage in northern boreal forests

abstract: Fungi are primary agents of coarse woody debris (CWD) decay in forests, playing an essential role in nutrient cycling and carbon storage. Characterizing fungal communities within CWD will promote further understanding of the fungal controls on CWD decomposition. We compared fungal community assemblages using alpha and beta diversity metrics, carbon mineralization, and physical and chemical properties of CWD across 3 tree species (trembling aspen [Populus tremuloides], black spruce [Picea mariana], and jack pine [Pinus banksiana]), 5 decay classes, and 2 stand development stages, differing in time-since-stand replacing disturbance (i.e., young/self-thinning and mature/steady-state) in Ontarios boreal forest region. In total, we sampled 180 individual CWD logs from 6 independent stands, with 3 replicates per each species x decay class combination at each site. We found that fungal community structure significantly differed across tree species, decay stage, and stand age. Higher proportions of white rot fungi were found in trembling aspen CWD, whereas higher proportions of brown rot fungi were found in black spruce and jack pine CWD. Proportions of specialized wood decay fungi increased with decay stage and were higher in CWD located in mature forest stands. Fungal diversity was highest in decay class 4 CWD. We found that Mn and K concentrations, total carbon, C/N ratio, carbon mineralization (mg CO2 g dry CWD-1 d-1), and moisture content were important predictors of fungal composition across CWD species and/or decay stage, though how CWD chemistry influences fungal species composition (and vice versa) is unknown. Carbon mineralization was highest in trembling aspen CWD and increased with decay stage, perhaps facilitated by increased N concentrations. This study suggests that forest management guidelines that consider both deadwood quantity and quality will support a broader range of fungal species and communities through post-disturbance stand development, thereby conserving biodiversity over the longer-term in our managed forest systems.


title: Modifications of lipoic arm by reactive nitrogen species regulate α-ketoacid dehydrogenases

abstract: Mitochondrial -ketoacid dehydrogenases, including the pyruvate dehydrogenase complex (PDHC) and the oxoglutarate dehydrogenase complex (OGDC), are a family of multi-subunit enzyme complexes that use a lipoic arm to transfer an acyl group to coenzyme A (CoA). The regulation of -ketoacid dehydrogenases plays crucial roles in mitochondrial metabolism and cellular energy homeostasis. We previously found that PDHC and OGDC become profoundly inhibited in macrophages upon classical activation, causing substantial remodeling of the TCA cycle. This inhibition was driven by the loss of the catalytically active lipoic moiety; however, the molecular mechanism causing this loss was not clear. Here we show that reactive nitrogen species (RNS), which are produced by activated macrophages, can cause a series of thiol-modifications to the lipoic arm that inactivate PDHC and OGDC. CoA-SNO, the non-enzymatic product between RNS and the E2 subunits natural substrate CoA, plays a key role in efficiently delivering RNS mediated modifications onto the lipoic arm. This work reveals a new biochemical mechanism capable of substantially regulating mitochondrial -ketoacid dehydrogenases, which has potential relevance for a range of physiological and pathological conditions.


title: Growth cone macropinocytosis of neurotrophin receptor and neuritogenesis are regulated by Neuron Navigator 1

abstract: Neuron Navigator 1 (Nav1) is a cytoskeleton-associated protein expressed during brain development that is necessary for proper neuritogenesis, but the underlying mechanisms are poorly understood. Here we show that Nav1 is present in elongating axon tracts during mouse brain embryogenesis. We found that depletion of Nav1 in cultured neurons disrupts growth cone morphology and neurotrophin-stimulated neuritogenesis. In addition to regulating both F-actin and microtubule properties, Nav1 promotes actin-rich membrane ruffles in the growth cone, and promotes macropinocytosis at those membrane ruffles, including internalization of the TrkB receptor for the neurotrophin BDNF (brain-derived neurotropic factor). Growth cone macropinocytosis is important for downstream signaling, neurite targeting, and membrane recycling, implicating Nav1 in one or more of these processes. Depletion of Nav1 also induces transient membrane blebbing via disruption of signaling in the Rho GTPase signaling pathway, supporting the novel role of Nav1 in dynamic actin-based membrane regulation at the cell periphery. These data demonstrate that Nav1 works at the interface of microtubules, actin, and plasma membrane to organize the cell periphery and promote uptake of growth and guidance cues to facilitate neural morphogenesis during development.


title: Striatal dopamine encodes the relationship between actions and reward

abstract: Despite the well-described role of dopamine in the striatum for skill learning and movement, its function in encoding actions and their outcomes for goal-directed action is less clear. Here, rats acquired two actions for distinct outcomes while we simultaneously recorded dopamine release in the dorsomedial striatum as these action-outcome associations were incremented and selectively degraded. Goal-directed actions generated a lateralized dopamine signal that reflected the strength of the action-outcome association and tracked increments and decrements in the specific action-outcome relationship, which was sensitive to changes in both contingency and outcome identity. This lateralized signal was updated by a dopamine action value signal that was broadcast bilaterally during the action and updated by a reward prediction error following exposure to the outcome. Our results establish, therefore, a dual role for striatal dopamine signaling during goal-directed actions: bilateral trial-by-trial burst signaling encoding action values, which modifies the long-term action-outcome association, reflected in emergent lateralization of dopamine release during actions.


title: A genomic snapshot of demographic and cultural dynamism in Upper Mesopotamia during the Neolithic Transition

abstract: Upper Mesopotamia played a key role in the Neolithic Transition in Southwest Asia through marked innovations in symbolism, technology, and foodways. We present thirteen ancient genomes (c.8500-7500 calBCE) from Pre-Pottery Neolithic Cayonu in the Tigris basin together with bioarchaeological and material culture data. Our findings reveal that Cayonu was a genetically diverse population, carrying a mixed ancestry from western and eastern Fertile Crescent, and that the community received immigrants. Our results further suggest that the community was organised along biological family lines. We document bodily interventions such as head-shaping and cauterization among the individuals examined, reflecting Cayonus cultural ingenuity. Finally, we identify Upper Mesopotamia as the likely source of eastern gene flow into Neolithic Anatolia, in line with material culture evidence. We hypothesise that Upper Mesopotamias cultural dynamism during the Neolithic Transition was the product not only of its fertile lands but also of its interregional demographic connections.


title: Volumetric development of hippocampal subfields and hippocampal white matter connectivity: Relationship with episodic memory

abstract: The hippocampus is a complex structure composed of several distinct subfields and has been at the center of scientific study examining the neural foundations of episodic memory. To date, there is little consensus regarding the structural development of the hippocampus and its subfields assessed both volumetrically and through anatomical connectivity, nor how the development of the related substructures influences episodic memory. In the current cross-sectional study, using a large sample of 830, 3- to 21-year-olds from a unique and publicly available dataset we examined the following questions: 1) Is there elevated grey matter volume of the hippocampus and its respective subfields in late compared to early development? 2) How does hippocampal volume compare with the rest of the cerebral cortex at different developmental stages? and 3) What is the relation between hippocampal volume, and the connectivity of the hippocampus with cortex as measured by diffusion-weighted imaging, with episodic memory performance? We found hippocampal subfield volumes exhibited a non-linear relation with age. Specifically, hippocampal subfield volumes showed a lag in volumetric change with age when compared to adjacent cortical regions (e.g., entorhinal cortex). We also observed a significant reduction in cortical volume across older cohorts, while hippocampal volume showed the opposite pattern. In addition to age-related differences in grey matter volume, several distinct subfields are significantly related to episodic memory. We did not, however, find any associations with episodic memory performance and connectivity through the uncinate fasciculus, fornix, or cingulum. The results are discussed in the context of current research and theories of hippocampal development and its relation to episodic memory.


title: Novel Association of Lyme Disease, Age, and Atopic Dermatitis

abstract:
Borrelia burgdorferi is a bacterial spirochete that can cause Lyme disease (LD) after infecting a susceptible host. Immune responses to the bacteria are highly variable and host specific. The murine substrain, C3H/HeJ, is a frequently utilized model for LD. Interestingly, we observed dermatitis with flaky lesions of the tail skin on C3H/HeJ after a year of infection with B. burgdorferi. Female C3H/HeJ mice aged 6-8 weeks, 1 year, or 2 years were infected intraperitoneally with 105 B. burgdorferi spirochetes. Mouse tails were evaluated by gross examination and histology either 2 months or 24 months post-infection. Dermatitis worsened over the course of untreated infection, with ulceration, hemorrhaging, flaking, hair loss, and dark lesions as well as spongiosis and acanthosis. These features of atopic dermatitis were present in infected mice after 1 year of age. This relationship among LD, atopic dermatitis, and host age seen in the C3H/HeJ mouse model is consistent with a large pool of human epidemiological data (342,499 individuals) from Finland. We identified 5,248 individuals with LD and 17,233 with atopic dermatitis in FinnGen. Retrospective analysis shows LD is associated with atopic dermatitis (OR = 1.91 [1.68 -2.37], P < 2e-16). Repeat visits for LD complications (3 or more visits versus 1 visit) were associated with atopic dermatitis (OR = 2.19 [1.35-3.55], P = 0.0014) and risk of developing atopic dermatitis over time (HR=2.26 [1.54-3.95], P = 0.0017). Data from mice and humans reveal a novel relationship among LD, age, and atopic dermatitis. Through defined pathological scoring, we demonstrate that the onset of murine Lyme disease associated atopic dermatitis is exacerbated by increased host age at time of B. burgdorferi infection. In humans, a diagnosis of Lyme disease in FinnGen was associated with atopic dermatitis and further research is warranted to establish causation.

ETHICS APPROVAL/CONSENT TO PARTICIPATEAnimal studies were performed at the Stanford School of Medicine Association for Assessment and Accreditation of Laboratory Animal Care International (AAALAC) accredited Rodent Animal Facility (Palo Alto, CA). All procedures and care guidelines were approved by the Stanford University Administrative Panel on Laboratory Animal Care (Protocol #30109).

Patients and control subjects in FinnGen provided informed consent for biobank research, based on the Finnish Biobank Act. Alternatively, separate research cohorts, collected prior the Finnish Biobank Act came into effect (in September 2013) and start of FinnGen (August 2017), were collected based on study-specific consents and later transferred to the Finnish biobanks after approval by Fimea (Finnish Medicines Agency), the National Supervisory Authority for Welfare and Health. Recruitment protocols followed the biobank protocols approved by Fimea. The Coordinating Ethics Committee of the Hospital District of Helsinki and Uusimaa (HUS) statement number for the FinnGen study is Nr HUS/990/2017.


title: Effects of global change on animal biodiversity in boreal forest landscape: an assemblage dissimilarity analysis

abstract: Despite an increasing number of studies highlighting the impacts of climate change on boreal species, the main factors that will drive changes in species assemblages remain ambiguous. We quantify two climate-induced pathways based on direct and indirect effects on species occupancy and assemblage dissimilarity under different harvest management scenarios. The direct climate effects illustrate immediate impact of climate variables while the indirect effects are reflected through the changes in land cover composition. To understand the main causes in assemblage dissimilarity, we analyze the regional and the latitudinal species assemblage dissimilarity by decomposing it into balanced variation in species occupancy and occurrence and occupancy and occurrence gradient. We develop empirical models to predict the distribution of more than 100 bird and beetle species in the Cote-Nord region of Quebec over the next century. Our results show the two pathways that are based on immediate and lagged climate change effects are complementary and alter biodiversity, mainly caused by balanced variation in species occupancy and occurrence. At the regional scale, both effects have an impact on decreasing the number of winning species. Yet, responses are much larger in magnitude under mixed climate effects (a mixture of direct and indirect effects). Regional assemblage dissimilarity reached 0.77 and 0.69 under mixed effects versus 0.09 and 0.10 under indirect effects for beetles and birds, respectively, between RCP8.5 and baseline climate scenarios when considering harvest. Therefore, inclusion of climatic variables considers aspects other than just those related to forest landscapes, such as life-cycle of animal species. Latitudinally, assemblage dissimilarity increased following the climate conditions pattern. Our analysis contributes to the understanding of how climate change alters biodiversity by reshaping community composition and highlights the importance of climate variables in biodiversity prediction.


title: Variation of the Rdr1 gene insertion in wild populations of Nicotiana benthamiana (Solanaceae) and insights into recent species divergence

abstract:
O_LIOne of the most commonly encountered and frequently cited laboratory organisms worldwide is classified taxonomically as Nicotiana benthamiana (Solanaceae), an accession of which, typically referred to as LAB, is renowned for its unique susceptibility to a wide range of plant viruses and hence capacity to be transformed using a variety of methods. However, the origin and age of LAB and the evolution of N. benthamiana across its wide distribution in Australia remains relatively underexplored.
C_LIO_LIHere, we have used multispecies coalescent methods on genome-wide single nuclear polymorphisms (SNPs) to assess species limits, phylogenetic relationships and divergence times within N. benthamiana.
C_LIO_LIOur results show that the previous taxonomic concept of this species in fact comprises five geographically, morphologically and genetically distinct species, one of which includes LAB.
C_LIO_LIRemarkably, we provide clear evidence that LAB is closely related to accessions collected further north in the Northern Territory; this species split much earlier from their common ancestor than the other four in this clade and is morphologically the most distinctive. Furthermore, this long-isolated species typically grows in sheltered sites in subtropical/tropical monsoon areas of northern Australia, contradicting the previously advanced hypothesis that this species is an extremophile that has traded viral resistance for precocious development.
C_LI


title: Electroencephalographic Signatures of Canine Cognitive Dysfunction

abstract: Canine cognitive dysfunction (CCD) is a highly prevalent neurodegenerative disease considered the canine analog of Alzheimers disease (AD). Unfortunately, CCD cannot be cured. However, early therapeutic interventions can slow the progression of cognitive decline and improve quality of life of the patients; therefore, early diagnosis is ideal. In humans, electroencephalogram (EEG) findings specific to AD have been described, and some of them have successfully detect early stages of the disease. In this study we characterized the EEG correlates of CCD, and we compared them with the EEGs of healthy aging dogs and dogs at risk of developing CCD. EEG recordings were performed in 25 senior dogs during wakefulness. Dogs were categorized in normal, at risk of CCD or with CCD according to their score in the Rofina questionnaire. We demonstrated that, quantitative EEG can detect differences between normal dogs and dogs with CCD. Dogs with CCD experience a reduction in beta and gamma interhemispheric coherence, and higher Joint Lempel Ziv complexity. Dogs at risk of developing CCD, had higher alpha power and interhemispheric coherence, making these features potential markers of early stages of the disease. These results demonstrate that EEG could be an additional biomarker that can contribute to the diagnosis of CCD, and reinforce the CCD as a translational model of AD.


title: Group-walk, a rigorous approach to separate FDR analysis by TDC

abstract:
Target-decoy competition (TDC) is a commonly used method for false discovery rate (FDR) control in the analysis of tandem mass spectrometry data. This type of competitionbased FDR control has recently gained significant popularity in other fields after Barber and Candes laid its theoretical foundation in a more general setting that included the feature selection problem. In both cases, the competition is based on a head-to-head comparison between an (observed) target score and a corresponding decoy (knockoff) score. However, the effectiveness of TDC depends on whether the data is homogeneous, which is often not the case: in many settings, the data consists of groups with different score profiles or different proportions of true nulls. In such cases, applying TDC while ignoring the group structure often yields imbalanced lists of discoveries, where some groups might include relatively many false discoveries and other groups include relatively very few. On the other hand, as we show, the alternative approach of applying TDC separately to each group does not rigorously control the FDR.

We developed Group-walk, a procedure that controls the FDR in the target-decoy / knockoff setting while taking into account a given group structure. Group-walk is derived from the recently developed AdaPT -- a general framework for controlling the FDR with sideinformation. We show using simulated and real datasets that when the data naturally divides into groups with different characteristics Group-walk can deliver consistent power gains that in some cases are substantial. These groupings include the precursor charge state (4% more discovered peptides at 1% FDR threshold), the peptide length (3.6% increase) and the mass difference due to modifications (26% increase).

Group-walk is available at https://cran.r-project.org/web/packages/groupwalk/index.html


title: Group-walk, a rigorous approach to group-wise false discovery rate analysis by target-decoy competition


title: HiC-LDNet: A general and robust deep learning framework for accurate chromatin loop detection in genome-wide contact maps

abstract:
MotivationIdentifying chromatin loops from genome-wide interaction matrices like Hi-C data is notoriously difficult. Such kinds of patterns can span through the genome from a hundred kilobases to thousands of kilobases. Most loop patterns are frequently related to biological functions, such as providing contacts between regulatory regions and promoters. They can also affect the cell-specific biological functions of different regulatory regions of DNA, thus leading to disease and tumorigenesis. While most statistical methods failed in the generalization to multiple cell types, recently proposed machine learning-based methods struggled when tested on sparse single-cell Hi-C (scHi-C) contact maps. We notice that there is an urgent need for an algorithm that can handle sparse scHi-C maps, and at the same time, can generate confident loop calls on regular cell lines.

ResultsTherefore, we propose a novel deep learning-based framework for Hi-C chromatin loop detection (HiC-LDNet) and provide corresponding downstream analysis. HiC-LDNet can give relatively more accurate predictions in multiple tissue types and contact technologies. Compared to other loop calling algorithms, such as HiCCUPS, Peakachu, and Chromosight, HiC-LDNet recovers a higher number of loop calls in multiple experimental platforms (Hi-C, ChIA-PET, DNA-SPRITE, and HiChIP), and achieves higher confidence scores in multiple cell types (Human GM12878, K562, HAP1, and H1-hESC). For example, in genome-wide loop detection on the human GM12878 cell line, HiC-LDNet successfully recovered 82.5% of loops within only 5 pixels of 10k bp resolution. Furthermore, in the sparse scHi-C ODC tissue, HiC-LDNet achieves superior performance by recovering 93.5% of ground truth loops with high confidence scores, compared with that of Peakachu (31.5%), Chromosight(69.6%), and HiCCUPS(9.5%). Therefore, our method is a robust and general pipeline for genome-wide chromatin loop detection for both bulk Hi-C and scHi-C data.

Availabilityhttps://github.com/frankchen121212/HiC-LDNet

Contactliyu@cse.cuhk.edu.hk and xin.gao@kaust.edu.sa


title: Composite branch construction by dual autozooidal budding modes in hornerids (Bryozoa: Cyclostomatida)

abstract: Horneridae (Cyclostomata; Cancellata) is a family of marine bryozoans that form tree-like colonies bearing functionally unilaminate branches. Colony development in this clade is not well understood. We used micro-CT and SEM to trace zooidal budding in Hornera from the ancestrula onwards. Results show that hornerid branches are constructed by dual zooidal budding modes occurring synchronously at two separate budding sites at the growing tips. Frontal autozooids bud from a multizooidal budding lamina. Lateral autozooids bud from discrete abfrontal budding loci by  exomural budding, a previously undescribed form of budding centred on hypostegal pores in interzooidal grooves on the colonial body wall. These two budding modes are integrated during primary branch morphogenesis, forming composite, developmentally bilaminate, branches. Tracing development back to the founding zooids shows that the ancestrula and adventitious periancestrular autozooids are the first-formed lateral autozooids. They grow as a vertical stem onto which new laterals bud exomurally, intercalating with earlier autozooids to form an expanding funnel. The inner wall of this funnel becomes a ring-shaped budding lamina, from which the first frontal autozooids bud centripetally. When the branch crown splits, the ring lamina is subdivided, each fragment becoming a paramedial budding lamina in its own branch, while new laterals continue to be budded exomurally onto the frontal walls of existing laterals. Co-occurrence of two rare cyclostome frontal budding types--adventitious budding on the ancestrula and exomural budding on the branches--suggests they may be homologous. Patterns of exomural budding vary among hornerids. Narrow-branched taxa possess a single medial line of budding sites on the abfrontal wall; wide-branched species have up to six parallel lines of loci. Exomural budding also occurs sporadically in Calvetia osheai, a radial-branching hornerid. Future studies of cancellate taxonomy and phylogeny may benefit from morphological concepts presented here.

category: zoology


title: Spatiotemporal dissociation of brain network connectivity in salience processing: A simultaneous pupillometry-EEG-fMRI study

abstract: The processing of salient stimuli involves a wide range of both bottom-up and top-down processes. Previous neuroimaging studies have identified multiple brain areas and networks for salience processing, including the salience network (SN), dorsal attention network (DAN), and the locus coeruleus-norepinephrine (LC-NE) neuromodulatory system. However, interactions among these networks and the cortico-subcortical systems in salience processing remain unclear. Here, we simultaneously recorded pupillometry, electroencephalogram (EEG), and functional magnetic resonance imaging (fMRI) during an auditory oddball paradigm. Using EEG-informed fMRI analysis, we temporally dissociated the target stimulus evoked activation, allowing us to identify the cascades of cortical areas associated with salience processing. Furthermore, functional connectivity analysis uncovered spatiotemporal functional network organizations of these salience processing neural correlates. Using pupillometry as a psychophysiological marker of LC-NE activity, we also assessed brain-pupil relationships. With state-space modeling of target modulated effective connectivity, we found that the target evoked pupillary response is associated with the network causal couplings from late to early subsystems, as well as the network switching initiated by the SN. These findings indicate that the SN might cooperate with pupil-indexed brainstem neuromodulatory systems, such as the LC-NE system, in the reorganization and dynamic switching of cortical networks, and shed light on the implications of their integrative framework in various cognitive processes and neurological diseases.


title: Quantification of phosphonate drugs by 1H-31P HSQC shows that rats are better models of primate drug exposure than mice

abstract: The phosphonate group is a key pharmacophore in many anti-viral, anti-microbial, and anti-neoplastic drugs. Due to its high polarity and short retention time, detecting and quantifying such phosphonate-containing drugs with LC/MS-based methods is challenging and requires derivatization with hazardous reagents. Given the emerging importance of phosphonate-containing drugs, developing a practical, accessible, and safe method for their quantitation in pharmacokinetics (PK) studies is desirable. NMR-based methods are often employed in drug discovery but are seldom used for compound quantitation in PK studies. Here, we show that proton-phosphorous (1H-31P) heteronuclear single quantum correlation (HSQC) NMR allows for quantitation of the phosphonate-containing enolase inhibitor HEX in plasma and tissue at micromolar concentrations. Although mice were shown to rapidly clear HEX from circulation (over 95% in <1 hr), the plasma half-life of HEX was more than 1hr in rats and nonhuman primates. This slower clearance rate affords a significantly higher exposure of HEX in rat models compared to mouse models while maintaining a favorable safety profile. Similar results were observed for the phosphonate-containing antibiotic, fosfomycin. Our study demonstrates the applicability of the 1H-31P HSQC method to quantify phosphonate-containing drugs in complex biological samples and illustrates an important limitation of mice as preclinical model species for phosphonate-containing drugs.

category: pharmacology and toxicology


title: G protein-coupled oestrogen receptor activation by Bisphenol-A disrupts protection from apoptosis conferred by oestrogen receptors ERα and ERβ in pancreatic beta cells

abstract:
17{beta}-estradiol protects pancreatic {beta}-cells from apoptosis via the estrogen receptors ER, ER{beta} and GPER. Conversely, the endocrine disruptor Bisphenol-A (BPA), which exerts multiple effects in this cell type via the same estrogen receptors, increased basal apoptosis. The molecular initiated events that trigger these opposite actions have yet to be identified. We demonstrated that combined genetic downregulation and pharmacological blockade of each estrogen receptor increased apoptosis to a different extent. The increase in apoptosis induced by BPA was diminished by the pharmacological blockade or the genetic silencing of GPER, and it was partially reproduced by the GPER agonist G1. BPA and G1-induced apoptosis were abolished upon pharmacological inhibition, silencing of ER and ER{beta}, or in dispersed islet cells from ER{beta} knockout (BERKO) mice. Yet, the ER and ER{beta} agonists, PPT and DPN, respectively, had no effect on beta cell viability. To exert their biological actions, ER and ER{beta} form homodimers and heterodimers. Molecular dynamic simulations together with proximity ligand assay and coimmunoprecipitation experiments indicated that the interaction of BPA with ER and ER{beta} as well as the GPER activation by G1 decreased ER{beta} heterodimers. We propose that ER{beta} heterodimers play an antiapoptotic role in beta cells and that BPA- and G1-induced decrease in ER{beta} heterodimers leads to beta cell apoptosis. Unveiling how different estrogenic chemicals affect the crosstalk among estrogen receptors should help to identify diabetogenic endocrine disruptors.

HighlightsO_LIPharmacological blockade and gene silencing of estrogen receptors ER, ER{beta} and GPER indicate that they are antiapoptotic in basal conditions.
C_LIO_LIGPER activation by G1 and BPA triggered apoptosis via a crosstalk with ER and ER{beta}.
C_LIO_LIBPA interaction with ER and ER{beta} as well as GPER activation decreased ER{beta} heterodimers, which was associated to increased apoptosis.
C_LIO_LIThis pathway represents a novel molecular initiating event underlying the pro-apoptotic effect of BPA
C_LIO_LIThe EndoC-{beta}H1 cell line may be a valid model of human {beta}-cells for identifying diabetogenic pollutants.
C_LI

category: physiology


title: G protein-coupled estrogen receptor activation by Bisphenol-A disrupts protection from apoptosis conferred by estrogen receptors ERα and ERβ in pancreatic beta cells


title: Discovery of novel therapeutic targets in cancer using patient-specific gene regulatory networks.

abstract: Most cancer types lack effective targeted therapeutic options and in cancers where first-line targeted therapies are available, treatment resistance is a huge challenge. Recent technological advances enable the use of ATAC-seq and RNA-seq on patient biopsies in a high-throughput manner. Here we present a computational approach that leverages these datasets to identify novel drug targets based on tumor lineage. We constructed patient-specific gene regulatory networks for 371 patients of 22 cancer types using machine learning approaches trained using three-dimensional genomic data for enhancer to promoter contacts. Next, we identify the key transcription factors (TFs) in these networks, which are used to identify therapeutic vulnerabilities either by direct targeting of TFs or proteins that they co-operate with. We validate four novel candidates identified for neuroendocrine, liver and renal cancers, which have a dismal prognosis with current therapeutic options. We present a novel approach to use the increasing amounts of functional genomics data from patient biospecimens for identification of novel drug targets.

category: systems biology


title: Amyloid fibrils in frontotemporal lobar degeneration with TDP-43 inclusions are composed of TMEM106B, rather than TDP-43

abstract: FTLD is the third most common neurodegenerative condition, following only Alzheimers and Parkinsons diseases. FTLD typically presents in 45-64-year-olds with behavioral changes or progressive decline of language skills. The subtype FTLD-TDP is characterized by certain clinical symptoms and pathological neuronal inclusions detected by TDP-43 immunoreactivity. Here, we extracted amyloid fibrils from brains of four patients, representing four out of five FTLD-TDP subclasses and determined their near-atomic resolution structures by cryo-EM. Unexpectedly, all amyloid fibrils examined are composed of a 135-residue C-terminal fragment of TMEM106B, a lysosomal membrane protein previously implicated as a genetic risk factor for FTLD-TDP. In addition to TMEM106B fibrils, abundant non-fibrillar aggregated TDP-43 is present, as revealed by immunogold labeling. Our observations confirm that FTLD-TDP is an amyloid-involved disease and suggest that amyloid involvement in FTLD-TDP is of protein TMEM106B, rather than of TDP-43.

category: molecular biology


title: Learning probabilistic protein-DNA recognition codes from DNA-binding specificities using structural mappings

abstract: Knowledge of how proteins interact with DNA is essential for understanding gene regulation. While DNA-binding specificities for thousands of transcription factors (TFs) have been determined, the specific amino acid-base interactions comprising their structural interfaces are largely unknown. This lack of resolution hampers attempts to leverage these data in order to predict specificities for uncharacterized TFs or TFs mutated in disease. Here we introduce rCLAMPS (Recognition Code Learning via Automated Mapping of Protein-DNA Structural interfaces), a probabilistic approach that uses DNA-binding specificities for TFs from the same structural family to simultaneously infer both which nucleotide positions are contacted by particular amino acids within the TF as well as a recognition code that relates each base-contacting amino acid to nucleotide preferences at the DNA positions it contacts. We apply rCLAMPS to homeodomains, the second largest family of TFs in metazoans, and demonstrate that it learns a highly effective recognition code that can predict de novo DNA-binding specificities for TFs. Furthermore, we show that the inferred amino acid-nucleotide contacts reveal whether and how nucleotide preferences at individual binding site positions are altered by mutations within TFs. Our approach is an important step towards automatically uncovering the determinants of protein-DNA specificity from large compendia of DNA-binding specificities, and inferring the altered functionalities of TFs mutated in disease.


title: Keratin 8 is a scaffolding and regulatory protein of ERAD complexes

abstract:
Early recognition and enhanced degradation of misfolded proteins by the endoplasmic reticulum (ER) quality control and ER-associated degradation (ERAD) cause defective protein secretion and membrane targeting, as exemplified for Z-alpha 1 antitrypsin (Z-A1AT), responsible for alpha-1-antitrypsin deficiency (A1ATD) and F508del-CFTR (cystic fibrosis transmembrane conductance regulator) responsible for cystic fibrosis (CF).

Prompted by our previous observation that decreasing Keratin 8 (K8) expression increased trafficking of F508del-CFTR to the plasma membrane, we investigated whether K8 impacts trafficking of soluble misfolded Z-A1AT protein. The subsequent goal of this study was to elucidate the mechanism underlying the K8-dependent regulation of protein trafficking, focusing on the ERAD pathway.

The results show that diminishing K8 concentration in HeLa cells enhances secretion of both Z-A1AT and wild type (WT) A1AT with a 13-fold and 4-fold increase, respectively. K8 down-regulation triggers ER failure and cellular apoptosis when ER stress is jointly elicited by conditional expression of the s heavy chains, as previously shown for Hrd1 knock-out. Simultaneous K8 silencing and Hrd1 knock-out did not show any synergistic effect, consistent with K8 acting in the Hrd1-governed ERAD step. Fractionation experiments reveal that K8 is recruited to ERAD complexes containing Derlin2, Sel1 and Hrd1 proteins upon expression of Z/WT-A1AT and F508del-CFTR. Treatment of the cells with c407, a small molecule inhibiting K8 interaction, decreases K8 and Derlin2 recruitment to high-order ERAD complexes. This was associated with increased Z-A1AT secretion in both HeLa and Z-homozygous A1ATD patients respiratory cells. Overall, we provide evidence that K8 acts as an ERAD modulator. It may play a scaffolding protein role for early-stage ERAD complexes, regulating Hrd1-governed retrotranslocation initiation/ubiquitination processes. Targeting K8-containing ERAD complexes is an attractive strategy for the pharmacotherapy of A1ATD.

category: cell biology


title: CDK11 Promotes Centromeric Transcription to Maintain Centromeric Cohesion during Mitosis

abstract: Actively-transcribing RNA polymerase (RNAP)II is remained on centromeres to maintain centromeric cohesion during mitosis although it is largely released from chromosome arms. This pool of RNAPII plays an important role in centromere functions. However, the mechanism of RNAPII retention on mitotic centromeres is poorly understood. We here demonstrate that Cdk11 depletion-induced centromeric cohesion defects are largely independent of Bub1. We further show that Cdk11 depletion and expression of its kinase-dead version significantly reduce both RNAPII and elongating RNAPII (pSer2) levels on centromeres, and also decrease centromeric transcription without altering the protein expression of cohesin and cohesion-regulators. Interestingly, enhanced centromeric transcription by THZ1 treatment or overexpression of CENP-B DNA-binding domain completely rescues Cdk11-depletion defects. These results suggest that Cdk11 promotes centromeric cohesion through facilitating centromeric transcription. Mechanistically, Cdk11 binds and phosphorylates RNAPII to promote transcription. Furthermore, mitosis-specific degradation of G2/M Cdk11-p58 recapitulates Cdk11-depletion defects. Altogether, our findings establish Cdk11 as an important regulator of centromeric transcription and as part of the mechanism for retaining RNAPII on centromeres during mitosis.