title: Wnt/β-catenin regulates alloreactive T cells for the treatment of hematological malignancies

abstract: Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is one of the most widely applied forms of adaptive immunotherapy. Both the detrimental graft-versus-host disease (GVHD) and the beneficial graft-versus-leukemia (GVL) effects occurring after allo-HSCT are largely mediated by alloantigen-reactive donor T cells in the graft. Separating GVHD from GVL effects is a formidable challenge, and a greater understanding of donor T cell biology is required to accomplish the uncoupling of GVHD from GVL. Here, we tested a novel mouse model of {beta}-catenin (Cat-Tg) in an allo-HSCT model. Our data show that T cells from Cat-Tg mice did not cause GVHD. Surprisingly, Cat-Tg T cells maintained the GVL effect. Donor T cells from Cat-Tg mice exhibited significantly lower inflammatory cytokine production and reduced donor T cell proliferation, while upregulating cytotoxic mediators that resulted in enhanced cytotoxicity. RNA sequencing data revealed changes in the expression of over 150 genes for CD4, and over 250 genes for CD8+T cells involved in essential aspects of immune response and GVHD pathophysiology. Transgenic over-expression of human {beta}-catenin primarily affects CD8+ T cell phenotype. Altogether, our data suggest that {beta}-catenin is a druggable target for developing therapeutic strategies to reduce GVHD while preserving the beneficial GVL effects following allo-HSCT treatment.

category: immunology


title: The Seroprevalence of SARS-CoV-2 in Europe: A Systematic Review

abstract:
BackgroundA year following the onset of the COVID-19 pandemic, new infections and deaths continue to increase in Europe. Serological studies, through providing evidence of past infection, can aid understanding of the population dynamics of SARS-CoV-2 infection.

ObjectivesThis systematic review of SARS-CoV-2 seroprevalence studies in Europe was undertaken to inform public health strategies including vaccination, that aim to accelerate population immunity.

MethodsWe searched the databases Web of Science, MEDLINE, EMBASE, SCOPUS, Cochrane Database of Systematic Reviews and grey literature sources for studies reporting seroprevalence of SARS-CoV-2 antibodies in Europe published between 01/12/2019 - 30/09/20. We provide a narrative synthesis of included studies. Studies were categorized into subgroups including healthcare workers (HCWs), community, outbreaks, pregnancy and children/school. Due to heterogeneity in other subgroups, we only performed a random effects meta-analysis of the seroprevalence amongst HCWs stratified by their country.

Results109 studies were included spanning 17 European countries, that estimated the seroprevalence of SAR-CoV2 from samples obtained between November 2019 - August 2020. A total of 53/109 studies included HCWs with a reported seroprevalence among HCWs ranging from 0.7% to 45.3%, which did not differ significantly by country. In community studies significant heterogeneity was reported in the seroprevalence among different age groups and the majority of studies reported there was no significant difference by gender.

ConclusionThis review demonstrates a wide heterogeneity in reported seroprevalence of SARS-CoV-2 antibodies between populations. Continued evaluation of seroprevalence is required to understand the impact of public health measures and inform interventions including vaccination programmes.

category: microbiology


title: How basic components of information-updating interact to encourage variation in the results of empirical studies of within and transgenerational plasticity

abstract: Experiences of parents and/or offspring are often assumed to affect the development of trait values in offspring because they provide information about the external environment, but it is currently unclear how information from different sources and times might combine to affect the information-states that provide the foundation for the patterns observed in empirical studies of developmental plasticity in response to environmental cues. We analyze Bayesian models designed to mimic fully-factorial experimental studies of within and transgenerational plasticity (TWP), in which parents, offspring, neither or both are exposed to cues from predators, to determine how different durations of cue exposure for parents and offspring, the devaluation of information from parents or the degradation of information from parents would affect offspring estimates of environmental states related to risk of predation at the end of such experiments. We show that the effects of different cue durations, the devaluation of information from parents, and the degradation of information from parents on offspring estimates are all expected to vary as a function of interactions with two other key parameters of information-based models of TWP: parental priors and the relative cue reliability in the different treatments. Our results suggest empiricists should expect to observe considerable variation across experimental studies of TWP based on simple principles of information-updating, without needing to invoke additional assumptions about costs, tradeoffs, development constraints, the fitness consequences of different trait values, or other factors.

category: animal behavior and cognition


title: Heterogeneity in Proline Hydroxylation of Fibrillar Collagens Observed by Mass Spectrometry

abstract:
Collagen is the major protein in the extracellular matrix and plays vital roles in tissue development and function. Collagen is also one of the most processed proteins in its biosynthesis. The most prominent post-translational modification (PTM) of collagen is the hydroxylation of Pro residues in the Y-position of the characteristic (Gly-Xaa-Yaa) repeating amino acid sequence of a collagen triple helix. Recent studies using mass-spectrometry (MS) and tandem MS sequencing (MS/MS) have revealed unexpected hydroxylation of Pro residues in the X-positions (X-Hyp). The newly identified X-Hyp residues appear to be highly heterogeneous in location and percent occupancy. In order to understand the dynamic nature of the new X-Hyps and their potential impact on applications of MS and MS/MS for collagen research, we sampled four different collagen samples using standard MS and MS/MS techniques. We found considerable variations in the degree of PTMs of the same collagen from different organisms and/or tissues. The rat tail tendon type I collagen is particularly variable in terms of both over-hydroxylation of Pro in the X-position and under-hydroxylation of Pro in the Y-position. In contrast, only a few unexpected PTMs in collagens type I and type III from human placenta were observed. The reproducibility of the different sequencing efforts of the same sample is also limited especially when the modified species are present at a low population, presumably due to the unpredictable nature of the ionization process. Additionally, despite the heterogeneous preparation and sourcing, collagen samples from commercial sources do not show elevated variations in PTMs compared to samples prepared from a single tissue and/or organism. These findings will contribute to the growing body of information regarding the PTMs of collagen by MS technology, and culminate to a more comprehensive understanding of the extent and the functional roles of the PTMs of collagen.

Abbreviations pageBoth the single letter and the three letter abbreviations of an amino acid will be used with the following additions: Hyp or O stands for 4R-hydroxylated proline and 3Hyp stands for 3- hydroxylated proline. When needed for clarity, the lower case single letter abbreviation will be used to represent the genomic DNA sequence, and upper case ones the sequence seen in the peptides.

category: biochemistry


title: Optical bench simulation for intraocular lenses using field-tracing technology

abstract:
PurposeTo evaluate the image quality of intraocular lenses (IOLs) using field-tracing optical simulation and then compare it with the image quality using conventional ray-tracing simulation.

MethodsWe simulated aspheric IOLs with a decenter, tilt, and no misalignment using an aspheric corneal eye model with a positive spherical aberration. The retinal image, Strehl ratio, and modulation transfer function (MTF) were compared between the ray-tracing and field-tracing optical simulation and confirmed by the results reported in an in vitro experiment using the same eye model.

ResultsThe retinal image showed interference fringes from target due to diffraction from the object in a field-tracing simulation. When compared with the experimental results, the field tracing represented the experimental results more precisely than ray tracing after passing over 400 {micro}m of the decentration and 4 degrees of the tilt of the IOLs. The MTF values showed similar results for the case of no IOL misalignment in both the field tracing and ray tracing. In the case of the 200-{micro}m decentration or 8-degree tilt of IOL, the field-traced MTF shows lower values than the ray-traced one.

ConclusionsThe field-tracing optical bench simulation is a reliable method to evaluate IOL performance according to the IOL misalignment. It can provide retinal image quality close to real by taking into account the wave nature of light, interference and diffraction to explain to patients having the IOL misalignment.

category: bioinformatics


title: Renalase is a novel tissue and serological biomarker in pancreatic ductal adenocarcinoma

abstract: Dysregulated expression of the secretory protein renalase (RNLS) can promote pancreatic ductal adenocarcinoma (PDAC) growth in animal models. We characterized RNLS expression in premalignant and malignant PDAC tissue and investigated whether plasma RNLS levels corresponded to clinical PDAC characteristics. RNLS immunohistochemistry was used to determine the presence and distribution of RNLS in normal pancreas, chronic pancreatitis, PDAC precursor lesions, and PDAC tissues. Associations between pretreatment plasma RNLS and PDAC clinical status were assessed in patients with varied clinical stages of PDAC and included tumor characteristics, surgical resection in locally advanced/borderline resectable PDAC, and overall survival. Data were retrospectively obtained and correlated using non-parametric analysis. Mild to no RNLS was detected by histochemistry in the normal pancreas in the absence of abdominal trauma. In chronic pancreatitis, RNLS immunoreactivity localized to peri-acinar spindle-shaped cells in some samples. It was also widely present in PDAC precursor lesions and PDAC tissue. Among 240 patients with PDAC, elevated plasma RNLS levels were associated with worse tumor characteristics, including greater angiolymphatic invasion (80.0% vs. 58.1%, p = 0.012) and greater node positive disease (76.5% vs. 56.5%, p = 0.024). Overall survival was worse in patients with high plasma RNLS levels with median follow-up of 27.70 months vs. 65.03 months (p < 0.001). RNLS levels also predicted whether patients with locally advanced/borderline resectable (LA/BR) PDAC underwent resection (AUC 0.674; 95%CI 0.42-0.82, p = 0.04). Overall tissue RNLS was increased in both premalignant and malignant PDAC tissues compared to normal pancreas. Elevated plasma RNLS levels were associated with advanced tumor characteristics, decreased overall survival, and reduced resectability in patients with LA/BR PDAC. These studies show that RNLS levels are increased in premalignant pancreatic tissues and that its levels in plasma correspond to the clinical behavior of PDAC.

category: cancer biology


title: Preservation of collagen in the soft tissues of frozen mammoths

abstract: We investigated the characteristics of extracellular matrix (ECM) in the soft tissue of two frozen baby woolly mammoths (Mammuthus primigenius) that died and were buried in Siberian permafrost approximately 40,000 years ago. Morphological and biochemical analyses of mammoth lung and liver demonstrated that those soft tissues were preserved at the gross anatomical and histological levels. The ultrastructure of ECM components, namely a fibrillar structure with a collagen-characteristic pattern of cross-striation, was clearly visible with transmission and scanning electron microscopy. Type I and type IV collagens were detected by immunohistochemical observation. Quantitative amino acid analysis of liver and lung tissues of the baby mammoths indicated that collagenous protein is selectively preserved in these tissues as a main protein. Type I and type III collagens were detected as major components by means of liquid chromatography-mass spectrometry analysis after digestion with trypsin. These results indicate that the triple helical collagen molecule, which is resistant to proteinase digestion, has been preserved in the soft tissues of these frozen mammoths for 40,000 years.

category: paleontology


title: First description of Lotmaria passim and Crithidia mellificae haptomonad stage in the honeybee hindgut

abstract:
The remodelling of flagella into attachment structures is a common and important event in the insect stages of the trypanosomatid life cycle. Among their hymenopteran hosts, Lotmaria passim and Crithidia mellificae can parasitize Apis mellifera, and as a result they might have a significant impact on honeybee health. However, there are details of their life cycle and the mechanisms underlying their pathogenicity in this host that remain unclear. Here we show that both L. passim promastigotes and C. mellificae choanomastigotes differentiate into haptomonad stage covering the ileum and rectum of honeybees. These haptomonad cells remain attached to the host surface via zonular hemidesmosome-like structures, as revealed by Transmission Electron Microscopy. Hence, for the first time this work describes the haptomonad morphotype of these species and their hemidesmosome-like attachment in Apis mellifera, a key trait exploited by other trypanosomatid species to proliferate in the insect host hindgut.

Author summaryIn recent years, the mortality of European Honeybees (Apis mellifera) has risen worldwide due to a variety of factors, including their infection by parasites. Former studies have linked the presence of several trypanosomatids species, being Lotmaria passim and Crithidia mellificae the most prevalent ones, with this increase in mortality. Although previous studies have shown that trypanosomatid infection reduces the lifespan of bees, there is little information regarding their development in the gut when honeybees become infected. Here, for the first time we describe the haptomonad morphotype of these two trypanosomatid species in A. mellifera. The most characteristic feature of haptomonads is the extensive remodelling of the flagellum and the formation of junctional complexes at the host gut wall. The presence of this morphotype in the honeybee hindgut increases our understanding of the life cycle of these species and their possible pathogenic mechanisms. We found that they can multiply while attached and that their disposition, covering the hindgut walls, could hinder host nutrient uptake and consequently, represent a pathogenic mechanism itself. This attachment could also be a key stage in the life-cycle to prevent the trypanosomatids leaving the host prematurely, ensuring transmission through infective morphotypes.

category: pathology


title: Developmental changes in story-evoked responses in the neocortex and hippocampus

abstract: How does the representation of naturalistic life events change with age? Here we analyzed fMRI data from 415 children and adolescents (5 - 19 years) as they watched a narrative movie. In addition to changes in the degree of inter-subject correlation (ISC) with age in sensory and medial parietal regions, we used a novel measure (between-groups ISC) to reveal age-related shifts in the responses across the majority of the neocortex. Over the course of development, brain responses became more discretized into stable and coherent events, and shifted earlier in time to anticipate upcoming event transitions. However, hippocampal responses to event boundaries actually decreased with age, suggesting a shifting division of labor between episodic encoding processes and schematic event representations between the ages of 5 and 19.

category: neuroscience


title: Midbrain organoids with an SNCA gene triplication model key features of synucleinopathy

abstract: SNCA, the first gene associated with Parkinsons disease, encodes the -synuclein (-syn) protein, the predominant component within pathological inclusions termed Lewy bodies (LBs). The presence of LBs is one of the classical hallmarks found in the brain of patients with Parkinsons disease, and LBs have also been observed in patients with other synucleinopathies. However, the study of -syn pathology in cells has relied largely on two-dimensional culture models, which typically lack the cellular diversity and complex spatial environment found in the brain. Here, to address this gap, we use 3D midbrain organoids (hMOs), differentiated from human induced pluripotent stem cells derived from patients carrying a triplication of the SNCA gene and from CRISPR/Cas9 corrected isogenic control iPSCs. These hMOs recapitulate key features of -syn pathology observed in the brains of patients with synucleinopathies. In particular, we find that SNCA triplication hMOs express elevated levels of -syn and exhibit an age-dependent increase in -syn aggregation, manifested by the presence of both oligomeric and phosphorylated forms of -syn. These phosphorylated -syn aggregates were found in both neurons and glial cells and their time-dependent accumulation correlated with a selective reduction in dopaminergic neuron numbers. Thus, hMOs from patients carrying SNCA gene multiplication can reliably model key pathological features of Parkinsons disease and provide a powerful system to study the pathogenesis of synucleinopathies.


title: The world's hotspot of linguistic and biocultural diversity under threat

abstract:
Papua New Guinea is home to >10% of the worlds languages and rich and varied biocultural knowledge, but the future of this diversity remains unclear. We measured language skills of 6,190 students speaking 392 languages (5.5% of the global total) and modelled their future trends, using individual-level variables characterizing family language use, socio-economic conditions, students skills, and language traits. This approach showed that only 58% of the students, compared to 91% of their parents, were fluent in indigenous languages, while the trends in key drivers of language skills (language use at home, proportion of mixed-language families, urbanization, students traditional skills) predicted accelerating decline of fluency, to an estimated 26% in the next generation of students. Ethnobiological knowledge declined in close parallel with language skills. Varied medicinal plant uses known to the students speaking indigenous languages are replaced by a few, mostly non-native species for the students speaking English or Tok Pisin, the national lingua franca. Most (88%) students want to teach indigenous language to their children. While crucial for keeping languages alive, this intention faces powerful external pressures as key factors (education, cash economy, road networks, urbanization) associated with language attrition are valued in contemporary society.

Significance StatementAround the world, more than 7,000 languages are spoken, most of them by small populations of speakers in the tropics. Globalization puts small languages at a disadvantage, but our understanding of the drivers and rate of language loss remains incomplete. When we tested key factors causing language attrition among Papua New Guinean students speaking 392 different indigenous languages, we found an unexpectedly rapid decline in their language skills compared to their parents and predicted further acceleration of language loss in the next generation. Language attrition was accompanied by decline in the traditional knowledge of nature among the students, pointing to an uncertain future for languages and biocultural knowledge in the most linguistically diverse place on Earth.

category: ecology


title: The Akt-mTOR pathway drives myelin sheath growth by regulating cap-dependent translation

abstract:
In the vertebrate central nervous system, oligodendrocytes produce myelin, a specialized proteolipid rich membrane, to insulate and support axons. Individual oligodendrocytes wrap multiple axons with myelin sheaths of variable lengths and thicknesses. Myelin grows at the distal ends of oligodendrocyte processes and multiple lines of work have provided evidence that mRNAs and RNA binding proteins localize to myelin, together supporting a model where local translation controls myelin sheath growth. What signal transduction mechanisms could control this? One strong candidate is the Akt-mTOR pathway, a major cellular signaling hub that coordinates transcription, translation, metabolism, and cytoskeletal organization. Here, using zebrafish as a model system, we found that Akt-mTOR signaling promotes myelin sheath growth and stability during development. Through cell-specific manipulations to oligodendrocytes, we show that the Akt-mTOR pathway drives cap-dependent translation to promote myelination and that restoration of cap-dependent translation is sufficient to rescue myelin deficits in mTOR loss-of-function animals. Moreover, an mTOR-dependent translational regulator co-localized with mRNA encoding a canonically myelin-translated protein in vivo and bioinformatic investigation revealed numerous putative translational targets in the myelin transcriptome. Together, these data raise the possibility that Akt-mTOR signaling in nascent myelin sheaths promotes sheath growth via translation of myelin-resident mRNAs during development.

SIGNIFICANCE STATEMENTIn the brain and spinal cord oligodendrocytes extend processes that tightly wrap axons with myelin, a protein and lipid rich membrane that increases electrical impulses and provides trophic support. Myelin membrane grows dramatically following initial axon wrapping in a process that demands protein and lipid synthesis. How protein and lipid synthesis is coordinated with the need for myelin to be generated in certain locations remains unknown. Our study reveals that the Akt-mTOR signaling pathway promotes myelin sheath growth by regulating protein translation. Because we found translational regulators of the Akt-mTOR pathway in myelin, our data raise the possibility Akt-mTOR activity regulates translation in myelin sheaths to deliver myelin on demand to the places it is needed.


title: Cortical tracking of a background speaker modulates the comprehension of a foreground speech signal

abstract:
Understanding speech in background noise is a difficult task. The tracking of speech rhythms such as the rate of syllables and words by cortical activity has emerged as a key neural mechanism for speech-in-noise comprehension. In particular, recent investigations have used transcranial alternating current stimulation (tACS) with the envelope of a speech signal to influence the cortical speech tracking, demonstrating that this type of stimulation modulates comprehension and therefore evidencing a functional role of the cortical tracking in speech processing. Cortical activity has been found to track the rhythms of a background speaker as well, but the functional significance of this neural response remains unclear. Here we employ a speech-comprehension task with a target speaker in the presence of a distractor voice to show that tACS with the speech envelope of the target voice as well as tACS with the envelope of the distractor speaker both modulate the comprehension of the target speech.

Because the envelope of the distractor speech does not carry information about the target speech stream, the modulation of speech comprehension through tACS with this envelope evidences that the cortical tracking of the background speaker affects the comprehension of the foreground speech signal. The phase dependency of the resulting modulation of speech comprehension is, however, opposite to that obtained from tACS with the envelope of the target speech signal. This suggests that the cortical tracking of the ignored speech stream and that of the attended speech stream may compete for neural resources.

Significance StatementLoud environments such as busy pubs or restaurants can make conversation difficult. However, they also allow us to eavesdrop into other conversations that occur in the background. In particular, we often notice when somebody else mentions our name, even if we have not been listening to that person. However, the neural mechanisms by which background speech is processed remain poorly understood. Here we employ transcranial alternating current stimulation, a technique through which neural activity in the cerebral cortex can be influenced, to show that cortical responses to rhythms in the distractor speech modulate the comprehension of the target speaker. Our results evidence that the cortical tracking of background speech rhythms plays a functional role in speech processing.


title: DDX21 is a p38-MAPK sensitive nucleolar protein necessary for mouse preimplantation embryo development and cell-fate specification.

abstract: Successful navigation of the mouse preimplantation stages of development, during which three distinct blastocyst lineages are derived, represents a prerequisite for continued development. We previously identified a role for p38-mitogen-activated kinases (p38-MAPK) regulating blastocyst inner cell mass (ICM) cell-fate, specifically primitive endoderm (PrE) differentiation, that is intimately linked to rRNA precursor processing, polysome formation and protein translation regulation. Here, we develop this work by assaying the role of DEAD-box RNA helicase 21 (Ddx21), a known regulator of rRNA processing, in the context of p38-MAPK regulation of preimplantation mouse embryo development. We show nuclear Ddx21 protein is robustly expressed from the 16-cell stage, becoming exclusively nucleolar during blastocyst maturation; a localisation dependent on active p38-MAPK. Efficient siRNA mediated clonal Ddx21 knockdown within developing embryos is associated with profound cell autonomous and non-autonomous proliferation defects and reduced blastocyst volume, by the equivalent peri-implantation blastocyst stage. Moreover, ICM residing Ddx21 knockdown clones express the EPI marker NANOG but rarely express the PrE differentiation marker GATA4. These data contribute extra significance to emerging importance of lineage specific translation regulation, as identified for p38-MAPK, during mouse preimplantation development.

category: developmental biology


title: Biphasic roles of hedgehog signaling in the production and self-renewal of outerradial glia in the ferret cerebral cortex

abstract: The neocortex, the center for higher brain function, emerged in mammals and expanded in the course of evolution. The expansion of outer radial glia (oRGs) and intermediate progenitor cells (IPCs) plays key roles in the expansion and consequential folding of the neocortex. Therefore, understanding the mechanisms of oRG and IPC expansion is important for understanding neocortical development and evolution. By using mice and human cerebral organoids, we previously revealed that hedgehog (HH) signaling expands oRGs and IPCs. Nevertheless, it remained to be determined whether HH signaling expanded oRGs and IPCs in vivo in gyrencephalic species, in which oRGs and IPCs are naturally expanded. Here, we show that HH signaling is necessary and sufficient to expand oRGs and IPCs in ferrets, a gyrencephalic species, through conserved cellular mechanisms. HH signaling increases oRG-producing division modes of ventricular radial glia (vRGs), oRG self-renewal, and IPC proliferation. Notably, HH signaling affects vRG division modes only in an early restricted phase before superficial-layer neuron production peaks. Beyond this restricted phase, HH signaling promotes oRG self-renewal. Thus, HH signaling expands oRGs and IPCs in two distinct but continuous phases during cortical development.


title: Comprehensive analysis of prime editing outcomes in human embryonic stem cells

abstract: Prime editing is a versatile and precise genome editing technique that can directly copy desired genetic modifications into target DNA sites without the need for donor DNA. This technique holds great promise for the analysis of gene function, disease modeling, and the correction of pathogenic mutations in clinically relevant cells such as human pluripotent stem cells (hPSCs). Here we comprehensively tested prime editing in hPSCs by generating a doxycycline-inducible prime editing platform. Prime editing successfully induced all types of nucleotide substitutions and small insertions and deletions, similar to observations in other human cell types. Moreover, we compared prime editing and base editing for correcting a disease-related mutation in induced pluripotent stem cells derived form a patient with  1-antitrypsin (A1AT) deficiency. Finally, whole-genome sequencing showed that, unlike the cytidine deaminase domain of cytosine base editors, the reverse transcriptase domain of a prime editor does not lead to guide RNA-independent off-target mutations in the genome. Our results demonstrate that prime editing in hPSCs has great potential for complementing previously developed CRISPR genome editing tools.

category: genomics


title: Long-read whole genome analysis of human single cells

abstract: With long-read sequencing we have entered an era where individual genomes are routinely assembled to near-completion and where complex genetic variation can efficiently be resolved. Here we demonstrate that long reads can be applied also to study the genomic architecture of individual human cells. Clonally expanded CD8+ T-cells from a human donor were used as starting material for a droplet-based multiple displacement amplification (dMDA) method designed to ensure long molecule lengths and minimal amplification bias. Sequencing of two single cells was performed on the PacBio Sequel II system, generating over 2.5 million reads and ~20Gb HiFi data (>QV20) per cell, achieving up to 40% genome coverage. This data allowed for single nucleotide variant (SNV) detection, including in genomic regions inaccessible by short reads. Over 1000 high-confidence structural variants (SVs) per cell were discovered in the PacBio data, which is four times more than the number of SVs detected in Illumina dMDA data from clonally related cells. In addition, several putative clone-specific somatic SV events could be identified. Single-cell de novo assembly resulted in 454-598 Mb assembly sizes and 35-42 kb contig N50 values. 1762 (12.8%) of expected gene models were found to be complete in the best single-cell assembly. The de novo constructed mitochondrial genomes were 100% identical for the two single cells subjected to PacBio sequencing, although mitochondrial heteroplasmy was also observed. In summary, the work presented here demonstrates the utility of long-read sequencing towards understanding the extent and distribution of complex genetic variation at the single cell level.


title: Mediodorsal thalamus is critical for updating during extra-dimensional shifts but not reversals in the attentional set-shifting task

abstract:
Cognitive flexibility, attributed to frontal cortex, is vital for navigating the complexities of everyday life. The mediodorsal thalamus (MD), interconnected to frontal cortex, may influence cognitive flexibility. Here male rats performed an attentional set-shifting task measuring intra-dimensional and extra-dimensional shifts in sensory discriminations. MD lesion rats needed more trials to learn the rewarded sensory dimension. However, once the choice response strategy was established, learning further two-choice discriminations in the same sensory dimension, and reversals of the reward contingencies in the same dimension, were unimpaired. Critically though, MD lesion rats were impaired during the extra-dimensional shift, when they must rapidly update the optimal choice response strategy. Behavioral analyses showed MD lesion rats had significantly reduced correct within trial second choice responses. This evidence shows transfer of information via the MD is critical when monitoring and rapid within trial updates in established choice response strategies are required after there is a rule change.

Significance statementWe demonstrate for the first time that rodent mediodorsal (MD) thalamus is a critical node when choice response strategies need to change rapidly after a within session rule change but not after reversals of reward contingencies during reward guided learning. MD interactions with orbitofrontal cortex are critical for value based learning, while MD interactions with medial prefrontal cortex are critical for rapid within trial updating of optimal choice response rules. MD interactions with the orbitofrontal cortex are not always necessary for reversal learning.


title: Mutations affecting the N-terminal domains of SHANK3 point to different pathomechanisms in neurodevelopmental disorders.

abstract: Shank proteins are major scaffolds of the postsynaptic density of excitatory synapses. Mutations in SHANK genes are associated with autism and intellectual disability. The relevance of missense mutations for these pathologies is unclear. Several missense mutations in SHANK3 affect the N-terminal region, consisting of the Shank/ProSAP N-terminal (SPN) domain and a set of Ankyrin (Ank) repeats. Here we identify a novel SHANK3 missense mutation (p.L270M) in the Ankyrin repeats in patients with an ADHD-like phenotype. We functionally analysed this and a series of other mutations, using biochemical and biophysical techniques. We observe two major effects: (i) a loss of binding to {delta}-catenin (e.g. in the p.L270M variant), and (ii) interference with the intramolecular interaction between N-terminal SPN domain and the Ank repeats. This also interferes with binding to the -subunit of the calcium-/calmodulin dependent kinase II (CaMKII), and appears to be associated with a more severe neurodevelopmental pathology.


title: Epitope classification and RBD binding properties of neutralizing antibodies against SARS-CoV-2 variants of concern

abstract: Severe acute respiratory syndrome coronavirus-2 (SAR-CoV-2) causes coronavirus disease 2019 (COVID19) that is responsible for short and long-term disease, as well as death, in susceptible hosts. The receptor binding domain (RBD) of the SARS-CoV-2 Spike (S) protein binds to cell surface angiotensin converting enzyme type-II (ACE2) to initiate viral attachment and ultimately viral pathogenesis. The SARS-CoV-2 S RBD is a major target of neutralizing antibodies (NAbs) that block RBD - ACE2 interactions. In this report, NAb-RBD binding epitopes in the protein databank were classified as C1, C1D, C2, C3, or C4, using a RBD binding profile (BP), based on NAb-specific RBD buried surface area and used to predict the binding epitopes of a series of uncharacterized NAbs. Naturally occurring SARS-CoV-2 RBD sequence variation was also quantified to predict NAb binding sensitivities to the RBD-variants. NAb and ACE2 binding studies confirmed the NAb classifications and determined whether the RBD variants enhanced ACE2 binding to promote viral infectivity, and/or disrupted NAb binding to evade the host immune response. Of 9 single RBD mutants evaluated, K417T, E484K, and N501Y disrupted binding of 65% of the NAbs evaluated, consistent with the assignment of the SARS-CoV-2 P.1 Japan/Brazil strain as a variant of concern (VoC). RBD variants E484K and N501Y exhibited ACE2 binding equivalent to a Wuhan-1 reference SARS-CoV-2 RBD. While slightly less disruptive to NAb binding, L452R enhanced ACE2 binding affinity. Thus, the L452R mutant, associated with the SARS-CoV-2 California VoC (B.1.427/B.1.429-California), has evolved to enhance ACE2 binding, while simultaneously disrupting C1 and C2 NAb classes. The analysis also identified a non-overlapping antibody pair (1213H7 and 1215D1) that bound to all SARS-CoV-2 RBD variants evaluated, representing an excellent therapeutic option for treatment of SARS-CoV-2 WT and VoC strains.


title: Variant SARS-CoV-2 mRNA vaccines confer broad neutralization as primary or booster series in mice

abstract: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of a global pandemic. Safe and effective COVID-19 vaccines are now available, including mRNA-1273, which has shown 94% efficacy in prevention of symptomatic COVID-19 disease. However, the emergence of SARS-CoV-2 variants has led to concerns of viral escape from vaccine-induced immunity. Several variants have shown decreased susceptibility to neutralization by vaccine-induced immunity, most notably B.1.351 (Beta), although the overall impact on vaccine efficacy remains to be determined. Here, we present the initial evaluation in mice of 2 updated mRNA vaccines designed to target SARS-CoV-2 variants: (1) monovalent mRNA-1273.351 encodes for the spike protein found in B.1.351 and (2) mRNA-1273.211 comprising a 1:1 mix of mRNA-1273 and mRNA-1273.351. Both vaccines were evaluated as a 2-dose primary series in mice; mRNA-1273.351 was also evaluated as a booster dose in animals previously vaccinated with mRNA-1273. The results demonstrated that a primary vaccination series of mRNA-1273.351 was effective at increasing neutralizing antibody titers against B.1.351, while mRNA-1273.211 was effective at providing broad cross-variant neutralization. A third (booster) dose of mRNA-1273.351 significantly increased both wild-type and B.1.351-specific neutralization titers. Both mRNA-1273.351 and mRNA-1273.211 are being evaluated in pre-clinical challenge and clinical studies.


title: Multiscale interactome analysis coupled with off-target drug predictions reveals drug repurposing candidates for human coronavirus disease

abstract: The COVID-19 pandemic has led to an urgent need for the identification of new antiviral drug therapies that can be rapidly deployed to treat patients with this disease. COVID-19 is caused by infection with the human coronavirus SARS-CoV-2. We developed a computational approach to identify new antiviral drug targets and repurpose clinically-relevant drug compounds for the treatment of COVID-19. Our approach is based on graph convolutional networks (GCN) and involves multiscale host-virus interactome analysis coupled to off-target drug predictions. Cellbased experimental assessment reveals several clinically-relevant repurposing drug candidates predicted by the in silico analyses to have antiviral activity against human coronavirus infection. In particular, we identify the MET inhibitor capmatinib as having potent and broad antiviral activity against several coronaviruses in a MET-independent manner, as well as novel roles for host cell proteins such as IRAK1/4 in supporting human coronavirus infection, which can inform further drug discovery studies.

category: cell biology


title: From partial to whole genome imputation of SARS-CoV-2 for epidemiological surveillance

abstract:
Backgroundthe current SARS-CoV-2 pandemic has emphasized the utility of viral whole genome sequencing in the surveillance and control of the pathogen. An unprecedented ongoing global initiative is increasingly producing hundreds of thousands of sequences worldwide. However, the complex circumstances in which viruses are sequenced, along with the demand of urgent results, causes a high rate of incomplete and therefore useless, sequences. However, viral sequences evolve in the context of a complex phylogeny and therefore different positions along the genome are in linkage disequilibrium. Therefore, an imputation method would be able to predict missing positions from the available sequencing data.

ResultsWe developed impuSARS, an application that includes Minimac, the most widely used strategy for genomic data imputation and, taking advantage of the enormous amount of SARS-CoV-2 whole genome sequences available, a reference panel containing 239,301 sequences was built. The impuSARS application was tested in a wide range of conditions (continuous fragments, amplicons or sparse individual positions missing) showing great fidelity when reconstructing the original sequences. The impuSARS application is also able to impute whole genomes from commercial kits covering less than 20% of the genome or only from the Spike protein with a precision of 0.96. It also recovers the lineage with a 100% precision for almost all the lineages, even in very poorly covered genomes (< 20%)

Conclusionsimputation can improve the pace of SARS-CoV-2 sequencing production by recovering many incomplete or low-quality sequences that would be otherwise discarded. impuSARS can be incorporated in any primary data processing pipeline for SARS-CoV-2 whole genome sequencing.


title: Elevated anti-SARS-CoV-2 antibodies and IL-6, IL-8, MIP-1β, early predictors of severe COVID-19.

abstract:
BackgroundViral and host immune kinetics during acute COVID-19 and after remission of acute symptoms need better characterization.

MethodsSARS-CoV-2 RNA, anti-SARS-CoV-2 IgA, IgM, and IgG antibodies, and pro-inflammatory cytokines were measured in sequential samples among hospitalized COVID-19 patients during acute infection and 6 months following diagnosis.

Results24 laboratory-confirmed COVID-19 patients with mild/moderate and severe COVID-19 were included. Most were males 83%, median age of 61 years. 21% were admitted to the ICU and 8 of them (33.3%) met criteria for severe COVID-19 disease. A delay in SARS-CoV-2 levels decline during the first 6 days of follow-up and viral load persistence until month 3 were related with severe COVID-19, but not viral load levels at the diagnosis. Higher levels of anti-SARS-CoV-2 IgA, IgM, IgG and the cytokines IL-6, IL-8 and MIP-1{beta} at the diagnosis time were related with severe COVID-19 outcome. Higher levels of MIP-1{beta}, IL-1{beta}, MIP-1 and IFN-{gamma} were observed at month 1/3 during mild/moderate disease compared to severe COVID-19. IgG persisted at low levels after 6 months of diagnosis.

ConclusionsHigher concentrations of IgA, IgM, and IgG, and IL-6, IL-8 and MIP-1{beta} are identified as early predictors of COVID-19 severity, but not SARS-CoV-2 RNA levels at diagnosis.


title: Contrasting patterns and co-occurrence network of soil bacterial and fungal community along depth profiles in cold temperate montane forests of China

abstract: Soil bacterial and fungal communities with different key ecological functions play an important role in the boreal forest ecosystem. Despite several studies have reported the microbial altitudinal distribution patterns, our understanding about the characteristics of the microbial community and the core composition of the microbiome in cold-temperate mountain forests is still limited. In this study, Illumina MiSeq sequencing was used to investigate the changes in soil bacterial and fungal communities in surface and subsurface soils along at an altitudinal gradient (from 830 m to 1300 m) on Oakley Mountain in the northern Greater Khingan Mountains. Altitude and soil depth had significant impacts on the relative abundance of Proteobacteria, Acidobacteria and Actinobacteria (dominant phylum for bacteria), and altitude had significant impacts on the Ascomycota, Basidiomycota and Mucoromycota (dominant phylum for fungi). The diversity of bacterial and fungal communities showed a monotonous decrease and increase with altitude. The influence of altitude on bacterial and fungal community composition was greater than that of soil depth. The variation of pH and dissolved organic nitrogen (DON) content in different altitudes were the main factors driving the bacterial and fungal community structure, respectively. There is no obvious difference between the network structure of surface and subsurface soil fungal communities, while the network of subsurface soil bacterial communities was more complex and compact than the surface layer. The network nodes mainly belonging to Proteobacteria and Actinobacteria are the key species in the two soil layers. Our results demonstrated that the altitude had a stronger influence on soil bacterial and fungal communities than soil depth, and bacterial and fungal communities showed divergent patterns along the altitudes and soil profiles.


title: Rapid diversification of the Australian Amitermes group during late Cenozoic climate change

abstract: Late Cenozoic climate change led to the progressive aridification of Australia over the past 15 million years. This gradual biome turnover fundamentally changed Australias ecosystems, opening new niches and prompting diversification of plants and animals. One example is the Australian Amitermes Group (AAG), consisting of the Australian Amitermes and affiliated genera. Although it represents the most speciose and diverse higher termite group in Australia, little is known about its evolutionary history. We used ancestral range reconstruction and diversification analyses to illuminate 1) the origin and phylogenetic relationships of the AAG, 2) biogeographical processes leading to the current continent-wide distribution, and 3) timing and pattern of diversification in the context of late Cenozoic climate change. By estimating the first time-calibrated phylogeny, we show that the AAG is a monophyletic group, whose ancestor arrived ~11-10 million years ago from Southeast Asia. Ancestral range reconstruction indicates that Australias monsoon region was the launching point for a continental radiation that has been shaped by range expansions and within-area speciation rather than vicariance. We found that multiple arid species diversified from mesic and tropical ancestors in the Plio-Pleistocene, but also observed diversification in the opposite direction. Finally, we show that two pulses of rapid diversification coincided with past climate change during the late Miocene and early Pliocene. Consistent with rapid diversification, species accumulation slowed, likely caused by progressive niche saturation. This study provides a stepping stone for predicting the future response of Australias termite fauna in the face of human-mediated climate change.

category: evolutionary biology


title: A memetic algorithm enables global all-atom protein-protein docking with sidechain flexibility

abstract: Protein complex formation is encoded by specific interactions at the atomic scale, but the computational cost of modeling proteins at this level often requires the use of simplified energy models and limited conformational flexibility. In particular, the use of all-atom energy functions, backbone and sidechain flexibility results in rugged energy landscapes that are difficult to explore. In this study we develop a protein-protein docking algorithm, EvoDOCK, that combine the strength of a differential evolution algorithm for efficient exploration of the global search space with the benefits of a local optimization method to refine detailed atomic interactions. EvoDOCK enabled accurate and fast local and global protein-protein docking using an all-atom energy function with side-chain flexibility. Comparison with a standard method built on Monte Carlo optimization demonstrated improved accuracy and with increases in computational speed of up to 35 times. The evolutionary algorithm also enabled efficient atomistic docking with backbone flexibility.


title: A memetic algorithm enables efficient local and global all-atom protein-protein docking with backbone and sidechain flexibility


title: Imatinib mesylate induces necroptotic cell death and impairs autophagic flux in human cardiac progenitor cells

abstract: The receptor tyrosine kinase inhibitor imatinib mesylate has improved patient cancer survival rates but has been linked to long-term cardiotoxicity. This study investigated the effects of imatinib on cell viability, apoptosis, autophagy and necroptosis in human cardiac progenitor cells in vitro. After 24 hours, imatinib significantly reduced cell viability (75.9{+/-}2.7% vs._100.0{+/-}0.0%, n=5, p<0.05) at concentrations comparable to peak plasma levels (10 {micro}M). Further investigation showed no increase in caspase 3 or 7 activation. Imatinib also significantly reduced the fluorescence of cells stained with TMRM (74.6{+/-}6.5% vs. 100.0{+/-}0.0%, n=5, p<0.05), consistent with mitochondrial depolarization. Imatinib increased lysosome and autophagosome content relative to the control, as indicated by changes in acridine orange fluorescence (46.0{+/-}5.4% vs. 9.0{+/-}3.0, n=7, p<0.001) and expression of LAMP2 (2.4{+/-}0.3 fold, n=3, p<0.05) after 24 hours treatment. Although imatinib increased the expression of proteins associated with autophagy, it also impaired the autophagic flux, as demonstrated by the proximity ligation assay staining for LAMP2 (lysosome marker) and LC3II (autophagosome marker), with control cells showing 11.3{+/-}2.1 puncta per cell and 48 hours of imatinib treatment reducing the visible puncta to 2.7{+/-}0.7 per cell (n=10, p<0.05). Cell viability was partially recovered by autophagosome inhibition by wortmannin, with a 91.8{+/-}8.2% (n=5, p>0.05) increase in viability after imatinib and wortmannin co-treatment. Imatinib-induced necroptosis was associated with an 8.5{+/-}2.5-fold increase in activation of mixed lineage kinase domain-like pseudokinase. Imatinib-induced toxicity was rescued by RIP1 inhibition relative to the control; 88.6{+/-}3.0% vs. 100.0{+/-}0.0% (n=4, p>0.05). In summary, imatinib applied to human cardiac progenitor cells depolarizes mitochondria and induces cell death through necroptosis, which can be recovered by inhibition of RIP1, with an additional partial role for autophagy in the cell death pathway. These data provide two possible targets for co-therapies to address imatinib-induced long-term cardiotoxicity.

category: pharmacology and toxicology


title: In-depth comparative toxicogenomics of glyphosate and Roundup herbicides: histopathology, transcriptome and epigenome signatures, and DNA damage

abstract: Whether glyphosate-based herbicides (GBHs) are more potent than glyphosate alone at activating cellular mechanisms, which drive carcinogenesis remains controversial. As GBHs are more cytotoxic that glyphosate, we reasoned they may also be more capable of activating carcinogenic pathways. We tested this hypothesis by comparing the effects of glyphosate with Roundup GBHs both in vitro and in vivo. First, glyphosate was compared with representative GBHs namely MON 52276 (EU), MON 76473 (UK) and MON 76207 (USA) using the mammalian stem cell-based ToxTracker system. Here, MON 52276 and MON 76473, but not glyphosate and MON 76207, activated oxidative stress and unfolded protein responses. Second, molecular profiling of liver was performed in female Sprague-Dawley rats exposed to glyphosate or MON 52276 (both at 0.5, 50, 175 mg/kg bw/day glyphosate) for 90 days. MON 52276 but not glyphosate increased hepatic steatosis and necrosis. MON 52276 and glyphosate altered the expression of genes in liver reflecting TP53 activation by DNA damage and circadian rhythm regulation. Genes most affected in liver were similarly altered in kidneys. Small RNA profiling in liver showed decreased amounts of miR-22 and miR-17 from MON 52276 ingestion. Glyphosate decreased mir-30 while miR-10 levels were increased. DNA methylation profiling of liver revealed 5,727 and 4,496 differentially methylated CpG sites between the control and glyphosate and MON 52276 exposed animals respectively. Apurinic/apyrimidinic DNA damage formation in liver was increased with glyphosate exposure. Altogether, our results show that Roundup formulations cause more biological changes linked with carcinogenesis than glyphosate.


title: Comparative toxicogenomics of glyphosate and Roundup herbicides by mammalian stem cell-based genotoxicity assays and molecular profiling in Sprague-Dawley rats


title: Cold-triggered induction of ROS- and raffinose metabolism in freezing-sensitive taproot tissue of sugar beet

abstract: Sugar beet (Beta vulgaris subsp. vulgaris) is the exclusive source of sugar in the form of sucrose in temperate climate zones. There, sugar beet is grown as an annual crop from spring to autumn because of the damaging effect of freezing temperatures to taproot tissue. Natural and breeded varieties display variance in the degree of tolerance to freezing temperatures and genotypes with elevated tolerance to freezing have been isolated. Here we compare initial responses to frost between genotypes with either low and high winter survival rates. The selected genotypes differed in the severity of frost injury. We combined transcriptomic and metabolite analyses of leaf- and taproot tissues from such genotypes to elucidate mechanisms of the early freezing response and to dissect genotype- and tissue-dependent responses. Freezing temperatures induced drastic downregulation of photosynthesis-related genes in leaves but upregulation of genes related to minor carbohydrate metabolism, particularly of genes involved in raffinose metabolism in both, leaf and taproot tissue. In agreement with this, it has been revealed that raffinose and the corresponding intermediates, inositol and galactinol, increased markedly in these tissues. We found that genotypes with improved tolerance to freezing, showed higher accumulation of raffinose in a defined interior region within the upper part of the taproot, the pith, representing the tissue most susceptible to freeze damages. This accumulation was accompanied by specific upregulation of raffinose synthesizing enzymes in taproots, suggesting a protective role for raffinose and its precursors for freezing damage in sugar beet.

category: plant biology


title: Network dynamics scale with levels of awareness

abstract: Small world topologies are thought to provide a valuable insight into human brain organisation and consciousness. However, functional magnetic resonance imaging studies in consciousness have not yielded consistent results. Given the importance of dynamics for both consciousness and cognition, here we investigate how the diversity of brain dynamics pertaining to small world topology (quantified by sample entropy; dSW-E) scales with decreasing levels of awareness (i.e., sedation and disorders of consciousness). Paying particular attention to result reproducibility, we show that dSW-E is a consistent predictor of levels of awareness even when controlling for the underlying functional connectivity dynamics. We find that dSW-E of subcortical and cortical areas are predictive, with the former showing higher and more robust effect sizes across analyses. Consequently, we propose that the dynamic reorganisation of the functional information architecture, in particular of the subcortex, is a characteristic that emerges with awareness and has explanatory power beyond that of the complexity of dynamic functional connectivity.


title: Early memory deficits and extensive brain network disorganization in the AppNL-F/MAPT double knock-in mouse model of familial Alzheimer's disease.

abstract: A critical challenge in current research on AD is to clarify the relationship between early neuropathology and network dysfunction associated to the emergence of subtle memory alterations which announce disease onset. In the present work, the new generation AppNL-F/MAPT double knock in (dKI) model was used to evaluate early stages of AD. The initial step of tau pathology was restricted to the perirhinal-entorhinal region, sparing the hippocampus. This discrete neuropathological sign was associated with deficits in the object-place associative memory, one of the earliest recognition memories affected in individuals at risk for developing AD. Analyses of task-dependent c-Fos activation was carried out in 22 brain regions across the medial prefrontal cortex, claustrum, retrosplenial cortex, and medial temporal lobe. Initial hyperactivity was detected in the entorhinal cortex and the claustrum of dKI mice. The retention phase was associated to reduced network efficiency especially across cingulate cortical regions, which may be caused by a disruption of information flow through the retrosplenial cortex. Moreover, the relationship between network global efficiency and memory performance in the WT could predict memory loss in the dKI, further linking reduced network efficiency to memory dysfunction. Our results suggest that early perirhinal-entorhinal pathology is associated with local hyperactivity which spreads towards connected regions such as the claustrum, the medial prefrontal cortex and ultimately the key retrosplenial hub which is needed to relay information flow from frontal to temporal lobes. The similarity between our findings and those reported in the earliest stages of AD suggests that the AppNL-F/MAPT dKI model has a high potential for generating key information on the initial stage of the disease.


title: Spontaneous visual preference for face-like stimuli is impaired in newly-hatched domestic chicks exposed to valproic acid during embryogenesis

abstract: One of the most fascinating properties of the human brain is the capacity of newborn babies to recognize and orient to faces and simple schematic face-like patterns since the first hours of life. A striking feature of these social orienting mechanisms is their transversal appearance in remarkably diverse vertebrate species. Similar to newborn babies, also non-human primates and domestic chicks have been shown to express orienting responses to faces and schematic face-like patterns. More importantly, existing studies have hypothesized that early disturbances of these mechanisms represent one of the earliest biomarkers of social deficits in autism spectrum disorders (ASD). Recent data suggest that newborns at high risk for the disorder express altered responses to schematic face-like configurations. Here we modeled ASD in domestic chicks using the anticonvulsant valproic acid (VPA), and tested the animals for their predisposed preference towards schematic face-like configuration stimuli. We found that VPA impairs the chicks preference responses to the social stimuli. Based on the results shown here and in previous studies, we propose the domestic chicks as elective animal models to study early-emerging neurobehavioural markers and to investigate the biological mechanisms underlying face processing deficits in ASD.


title: Selection in a growing bacterial/yeast colony biases results of mutation accumulation experiments.

abstract: Mutations provide the raw material for natural selection to act. Therefore, understanding the variety and relative frequency of different type of mutations is critical to understanding the nature of genetic diversity in a population. Mutation accumulation (MA) experiments have been used in this context to estimate parameters defining mutation rates, distribution of fitness effects (DFE), and spectrum of mutations. MA experiments performed with organisms such as Drosophila have an effective population size of one. However, in MA experiments with bacteria and yeast, a single founder is allowed to grow to a size of a colony (~108). The effective population size in these experiments is of the order of 10. In this scenario, while it is assumed that natural selection plays a minimal role in dictating the dynamics of colony growth and therefore, the MA experiment; this effect has not been tested explicitly. In this work, we simulate colony growth and perform an MA experiment, and demonstrate that selection ensures that, in an MA experiment, fraction of all mutations that are beneficial is over represented by a factor greater than two. The DFE of beneficial and deleterious mutations are accurately captured in an MA experiment. We show that the effect of selection in a growing colony varies non-monotonically and that, in the face of natural selection dictating an MA experiment, estimates of mutation rate of an organism is not trivial. We perform experiments with 160 MA lines of E. coli, and demonstrate that rate of change of mean fitness is a non-monotonic function of the colony size, and that selection acts differently in different sectors of a growing colony. Overall, we demonstrate that the results of MA experiments need to be revisited taking into account the action of selection in a growing colony.


title: Revealing intraosseous blood flow in the human tibia with ultrasound

abstract: Intraosseous blood circulation is thought to have a critical role in bone growth and remodeling, fracture healing, and bone disorders. However, it is rarely considered in clinical practice due to the absence of a suitable non-invasive in vivo measurement technique. In this work, we assessed blood perfusion in tibial cortical bone simultaneously with blood flow in the superficial femoral artery with ultrasound imaging in 5 healthy volunteers. After suppression of stationary signal with Singular-Value-Decomposition, pulsatile blood flow in cortical bone tissue is revealed, following the heart rate measured in the femoral artery. Using a method combining transverse oscillations and phase-based motion estimation, two-dimensional vector flow was obtained in the cortex of the tibia. After spatial averaging over the cortex, the peak blood velocity along the long axis of the tibia was measured four times larger than the peak blood velocity across the bone cortex. This suggests that blood flow in central (Haversian) canals is larger than in perforating (Volkmanns) canals, as expected from the intracortical vascular organization in humans. The peak blood velocity indicates a flow from the endosteum to the periosteum and from the heart to the foot for all subjects. Because aging and the development of bone disorders are thought to modify the direction and velocity of intra-cortical blood flow, their quantification is crucial. This work reports for the first time an in vivo quantification of the direction and velocity of blood flow in human cortical bone.

category: bioengineering


title: Recombinant binder of sperm protein1 (rec-BSP1) as a potent fertility factor mediates its effect on embryo production

abstract:
ObjectiveTo understand the effect of recombinant BSP1 (rec-BSP1) on in vitro capacitation of sperm and fertilization study

Method(s)Articles were screened for reports including rec-BSP1, Capacitation, in vitro fertilization

InterventionNone

Main Outcome Measure(s)Reproductive outcomes, effect on gametes and embryos

Result(s)Here we report an optimization of condition for rec-BSP1 production which was used for in vitro capacitation and enhancement of buffalo embryo production. The sequence of the protein was used for multiple sequence alignment which has 99% similarity with PDC 109 protein. The expression of rec-BSP1 was carried out successfully with 1 mM IPTG at 160 C for 22 hrs and purified it in soluble form. The structure of rec-BSP1 was generated using 3D modelling and analysed its mode of binding with heparin and PC by molecular docking and the structural stability of rec-BSP1-PC and rec-BSP1-heparin complexes by using molecular dynamic (MD) simulation. The effect of rec-BSP1 was observed on in vitro capacitation of spermatozoa and buffalo blastocyst production. It was found that the rec-BSP1 enhanced the sperm motility at a concentration of 50 g/ml for 1 h of incubation without having any detrimental effect on the sperm morphology and a significant increase in blastocyst production at concentration of 50 g/ml rec-BSP1. Hence this finding represents a new insight and advance the prospective approach to develop a potential fertility factor in reproduction.

Conclusion(s)The purified rec-BSP1 may enhance on male fertility and mediated its effect on in vitro blastocyst production in buffalo.


title: RNA labelling in live plants reveals single cell transcriptional dynamics: application to phosphate signaling

abstract: Plants are sessile organisms constantly adapting to ambient fluctuations through spatial and temporal transcriptional responses. Here, we implemented the latest generation RNA imaging system and combined it with microfluidics to visualize transcriptional regulation in living Arabidopsis plants. This enabled quantitative measurements of the transcriptional activity of single loci in single cells, real time and changing environmental conditions. Using phosphate responsive genes as model, we found that active genes displayed high transcription initiation rates ([~]3s) and frequently clustered together in endoreplicated cells. We observed gene bursting and large allelic differences in single cells, revealing that at steady-state, intrinsic noise dominated extrinsic variations. Moreover, we established that transcriptional repression triggered in roots by phosphate, a crucial macronutrient limiting plant development, occurred with unexpected fast kinetics ([~]minutes) and striking heterogeneity between neighboring cells. Access to single cell RNA polymerase II dynamics within live plants will benefit future studies of signaling processes.


title: Feature selectivity explains mismatch signals in mouse visual cortex

abstract: Sensory experience is often dependent on ones own actions, including self-motion. Theories of predictive coding postulate that actions are regulated by calculating prediction error, which is the difference between sensory experience and expectation based on self-generated actions. Signals consistent with prediction error have been reported in mouse visual cortex (V1) when visual flow coupled to running is unexpectedly perturbed. Here, we show that such signals can be elicited by visual stimuli uncoupled with the animals running. We recorded the activity of mouse V1 neurons while presenting drifting gratings that unexpectedly stopped. We found strong responses to visual perturbations, which were enhanced during running. If these perturbation responses are signals about sensorimotor mismatch, they should be largest for front-to-back visual flow expected from the animals running. Responses, however, did not show a bias for front-to-back visual flow. Instead, perturbation responses were strongest in the preferred orientation of individual neurons and perturbation responsive neurons were more likely to prefer slow visual speeds. Our results therefore indicate that prediction error signals can be explained by the convergence of known motor and sensory signals in visual cortex, providing a purely sensory and motor explanation for purported mismatch signals.


title: A distinctive neural nexus in blind individuals supports Braille reading

abstract:
Natural Braille reading, a demanding cognitive skill, poses a huge challenge for the brain network of the blind. Here, with behavioral measurement and functional MRI imaging data, we pinpointed the neural pathway and investigated the neural mechanisms of individual differences in Braille reading in late blindness. Using resting state fMRI, we identified a distinct neural link between the higher-tier  visual cortex--the lateral occipital cortex (LOC), and the inferior frontal cortex (IFC) in the late blind brain, which is significantly stronger than sighted controls. Individual Braille reading proficiency positively correlated with the left-lateralized LOC-IFC functional connectivity. In a natural Braille reading task, we found an enhanced bidirectional information flow with a stronger top-down modulation of the IFC-to-LOC effective connectivity. Greater top-down modulation contributed to higher Braille reading proficiency via a broader area of task-engaged LOC. Together, we established a model to predict Braille reading proficiency, considering both functional and effective connectivity of the LOC-IFC pathway. This  two-tale model suggests that developing the underpinning neural circuit and the top-down cognitive strategy contributes uniquely to superior Braille reading performance.

SIGNIFICANCE STATEMENTFor late blind humans, one of the most challenging cognitive skills is natural Braille reading. However, little is known about the neural mechanisms of significant differences in individual Braille reading performance. Using functional imaging data, we identified a distinct neural link between the left lateral occipital cortex (LOC) and the left inferior frontal cortex (IFC) for natural Braille reading in the late blind brain. To better predict individual Braille reading proficiency, we proposed a linear model with two variables of the LOC-IFC link: the resting-state functional connectivity and the task-engaged top-down effective connectivity. These findings suggest that developing the underpinning neural pathway and the top-down cognitive strategy contributes uniquely to superior Braille reading performance.


title: A distinctive neural nexus predicts Braille reading proficiency in blind individuals


title: Two tales of one neural link predict blind individual's Braille reading proficiency


title: MLL1 Methyltransferase Activity is Regulated by Distinct Nucleosome Binding Modes

abstract: Here we solve the single particle cryoEM structure for the MLL1 complex with nucleosome core particle (NCP) carrying histone H3 lysine 4 to methionine mutation. The MLL1 complex displays significant rotational dynamics on the NCP, a feature distinct from the yeast SET1 complex. We identified two major binding modes of the MLL1 complex on the NCP. Both binding modes anchor on the NCP through ASH2L, but they differ drastically with regard to where the MLL1 SET domain and RbBP5 bind. We show that one of the binding modes is catalytically inactive since disrupting interactions unique to this binding mode does not affect overall MLL1 activity in an NCP-specific manner. Interestingly, the inactive binding mode is in a configuration similar to that of the ySET1- NCP complex, which is intrinsically inactive on an unmodified NCP. The high rotational dynamics of the MLL1 complex as well as distinction between MLL and yeast SET1 complexes may reflect the necessity for loci-specific regulation of H3K4 methylation states in higher eukaryotes.


title: Kinetic and structural characterization of the self-labeling protein tags HaloTag7, SNAP-tag and CLIP-tag

abstract: The self-labeling protein tags (SLPs) HaloTag7, SNAP-tag and CLIP-tag allow the covalent labeling of fusion proteins with synthetic molecules for applications in bioimaging and biotechnology. To guide the selection of an SLP-substrate pair and provide guidelines for the design of substrates, we report a systematic and comparative study on the labeling kinetics and substrate specificities of HaloTag7, SNAP-tag and CLIP-tag. HaloTag7 reaches almost diffusion-limited labeling rates with certain rhodamine substrates, which are more than two orders of magnitude higher than those of SNAP-tag for the corresponding substrates. SNAP-tag labeling rates however are less affected by the structure of the label than those of HaloTag7, which vary over six orders of magnitude for commonly employed substrates. Solving the crystal structures of HaloTag7 and SNAP-tag labeled with fluorescent substrates allowed us to rationalize their substrate preferences. We also demonstrate how these insights can be exploited to design substrates with improved labeling kinetics.


title: Synthetic biology meets proteomics: Construction of a la carte QconCATs for absolute protein quantification

abstract: We report a new approach to the assembly and construction of QconCATs, quantitative concatamers for proteomic applications that yield stoichiometric quantities of sets of stable isotope-labelled internal standards. The new approach is based on synthetic biology precepts of biobricks, making use of loop assembly to construct larger entities from individual biobricks. It offers a major gain in flexibility of QconCAT implementation and enables rapid and efficient editability that permits, for example, substitution of one peptide for another. The basic building block (a Qbrick) is a segment of DNA that encodes two or more quantification peptides for a single protein, readily held in a repository as a library resource. These Qbricks are then assembled in a one tube ligation reaction that enforces the order of assembly, to yield short QconCATs that are useable for small quantification products. However, the DNA context of the short also allows a second cycle of assembly such that five different short QconCATs can be assembled into a longer QconCAT in a second, single tube ligation. From a library of Qbricks, a bespoke QconCAT can be assembled quickly and efficiently in a form suitable for expression and labelling in vivo or in vitro. We refer to this approach as the ALACAT strategy as it permits a la carte design of quantification standards.


title: In vitro reconstitution of Sgk3 activation by phosphatidylinositol-3-phosphate

abstract: Serum- and glucocorticoid-regulated kinase 3 (Sgk3) is activated by the phospholipid phosphatidylinositol-3-phosphate (PI3P) downstream of growth factor signaling and by Vps34-mediated PI3P production on endosomes. Upregulation of Sgk3 activity has recently been linked to a number of human cancers. Here, we show that Sgk3 is regulated by a combination of phosphorylation and allosteric activation by PI3P. We demonstrate that PI3P binding induces large conformational changes in Sgk3 associated with its activation, and that the PI3P binding pocket of the PX domain of Sgk3 is sequestered in its inactive conformation. Finally, we reconstituted Sgk3 activation via Vps34-mediated PI3P synthesis on phosphatidylinositol liposomes in vitro. In addition to defining the mechanism of Sgk3 activation by PI3P, our findings open up potential therapeutic avenues in allosteric inhibitor development to target Sgk3 in cancer.


title: Chemoproteomic study uncovers HemK2/KMT9 as a new target for NTMT1 bisubstrate inhibitors

abstract:
Understanding the selectivity of methyltransferase inhibitors is important to dissect the functions of each methyltransferase target. From this perspective, here we report a chemoproteomic study to profile the selectivity of a potent protein N-terminal methyltransferase 1 (NTMT1) bisubstrate inhibitor NAH-C3-GPKK (Ki, app = 7{+/-}1 nM) in endogenous proteomes. First, we describe the rational design, synthesis, and biochemical characterization of a new chemical probe 6, a biotinylated analogue of NAH-C3-GPKK. Next, we systematically analyze protein networks that may selectively interact with the biotinylated probe 6 in concert with the competitor NAH-C3-GPKK. Besides NTMT1, the designated NTMT1 bisubstrate inhibitor NAH-C3-GPKK was found to also potently inhibit a methyltransferase complex HemK2-Trm112 (also known as KMT9-Trm112), highlighting the importance of systematic selectivity profiling. Furthermore, this is the first potent inhibitor for HemK2/KMT9 reported to date. Thus, our studies lay the foundation for future efforts towards the development of selective inhibitors for NTMT1 and HemK2/KMT9.

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title: Targeted Neuronal Activation of the Gastrointestinal Tract Shapes the Environment of the Gut in Mice

abstract: The enteric nervous system (ENS) coordinates responses that shape the intestinal milieu, and participates in bi-directional communication with various organs including the brain. Tools to study, and knowledge of, ENS development and function lag behind brain research. Herein, we deploy recombinant adeno-associated viral (rAAV) vectors with enhanced tropism for the gut to map and activate gut associated neurons in mice with spatial and temporal resolution. rAAV-mediated fluorescent labelling coupled with whole-tissue clearing methods in the small intestine and colon reveal novel neuronal architectures in the gut of mice. Viral delivery of genetically-encoded calcium indicators enable in vivo measurement of ENS activity. Further, we employed chemogentics to specifically activate gut neurons that express choline acetyltransferase (ChAT+) or tyrosine hydroxylase (TH+). Targeted activation of ChAT+ or TH+ neuronal populations associated with the gastrointestinal (GI) tract alters the intestinal transcriptome, mouse and microbial proteomes, metagenome, and fecal metabolome. We reveal previously unknown and broad roles for enteric neurons in modulating GI physiology, mucosal immunity, and microbiome structure, and provide evidence for novel functions by the ENS such as regulating fungal colonization and shaping of bile acid profiles in the gut. These advanced experimental tools and rich datasets with multi-parameter characterization of the gut ecosystem may facilitate expanded understanding and study of the ENS.

category: physiology


title: Neuronal Activation of the Gastrointestinal Tract Shapes the Gut Environment in Mice


title: Genetic architecture of floral traits in bee- and hummingbird-pollinated sister species of Aquilegia (columbine)

abstract: Interactions with animal pollinators have helped shape the stunning diversity of flower morphologies across the angiosperms. A common evolutionary consequence of these interactions is that some flowers have converged on suites of traits, or pollination syndromes, that attract and reward specific pollinator groups. Determining the genetic basis of these floral pollination syndromes can help us understand the processes that contributed to the diversification of the angiosperms. Here, we characterize the genetic architecture of a bee-to-hummingbird pollination shift in Aquilegia (columbine) using QTL mapping of 17 floral traits encompassing color, nectar composition, and organ morphology. In this system, we find that the genetic architectures underlying differences in floral color are quite complex, and we identify several likely candidate genes involved in anthocyanin and carotenoid floral pigmentation. Most morphological and nectar traits also have complex genetic underpinnings; however, one of the key floral morphological phenotypes, nectar spur curvature, is shaped by a single locus of large effect.


title: Nisin- and ripcin-derived hybrid lanthipeptides display selective antimicrobial activity against Staphylococcus aureus

abstract:
Lanthipeptides are (methyl)lanthionine ring-containing ribosomally synthesized and post-translationally modified peptides (RiPPs). Many lanthipeptides show strong antimicrobial activity against bacterial pathogens, including antibiotic-resistant bacterial pathogens. The group of disulfide bond-containing antimicrobial peptides (AMPs) is well known in nature and forms a rich source of templates for the production of novel peptides with corresponding (methyl)lanthionine analogues instead of disulfides. Here, we show that novel macrocyclic lanthipeptides (termed thanacin and ripcin) can be synthesized using the known antimicrobials thanatin and rip-thanatin as templates. Notably, the synthesized nisin(1-20)-ripcin hybrid lanthipeptides (ripcin B-G) showed selective antimicrobial activity against S. aureus, including an antibiotic-resistant MRSA strain. Interestingly, ripcin B-G, which are hybrid peptides of nisin(1-20) and ripcin, respectively, that are each inactive against Gram-negative pathogens, showed substantial antimicrobial activity against the tested Gram-negative pathogens. Moreover, ripcin B-G was highly resistant against the nisin resistance protein (NSR; a protease could cleave nisin and strongly reduce its activity), opposed to nisin itself. Mode of action studies show that ripcin C exerts its antimicrobial activity against Gram-positive pathogens by binding to the cell wall synthesis precursor lipid II and thereafter arrests cell growth. In addition, ripcin C exerts its antimicrobial activity against Gram-negative pathogens by binding to LPS and the cell wall synthesis precursor lipid II. This study provides an example of converting disulfide bond-based AMPs into (methyl)lanthionine-based macrocyclic hybrid lanthipeptides and can yield antimicrobial peptides with selective antimicrobial activity against S. aureus.

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category: synthetic biology


title: Dual-function RNA-binding proteins influence mRNA abundance and translational efficiency of distinct sets of target genes.

abstract: RNA-binding proteins (RBPs) can regulate more than a single aspect of RNA metabolism. We searched for such previously undiscovered multifunctionality within a set of 143 RBPs, by defining the predictive value of RBP abundance for the transcription and translation levels of known RBP target genes across 80 human hearts. This led us to newly associate 27 RBPs with cardiac translational regulation in vivo. Of these, 21 impacted both RNA expression and translation, albeit for virtually independent sets of target genes. We highlight a subset of these, including G3BP1, PUM1, UCHL5, and DDX3X, where dual regulation is achieved through differential affinity for target length, by which separate biological processes are controlled. Similar to the RNA helicase DDX3X, the known splicing factors EFTUD2 and PRPF8 - all identified as multifunctional RBPs by our analysis - selectively influence target translation rates depending on 5 UTR structure. Our analyses identify dozens of RBPs as being multifunctional and pinpoint potential novel regulators of translation, postulating unanticipated complexity of protein-RNA interactions at consecutive stages of gene expression.

category: systems biology


title: Multifunctional RNA-binding proteins influence mRNA abundance and translational efficiency of distinct sets of target genes


title: Self-regulation of stress-related large-scale brain network balance using real-time fMRI Neurofeedback

abstract:
It has recently been shown that acute stress affects the allocation of neural resources between large-scale brain networks, and the balance between the executive control network and the salience network in particular. Maladaptation of this dynamic resource reallocation process is thought to play a major role in stress-related psychopathology, suggesting that stress resilience may be determined by the retained ability to adaptively reallocate neural resources between these two networks. Actively training this ability could hence be a potentially promising way to increase resilience in individuals at risk for developing stress-related symptomatology. Using real-time functional Magnetic Resonance Imaging, the current study investigated whether individuals can learn to self-regulate stress-related large-scale network balance. Participants were engaged in a bidirectional and implicit real-time fMRI neurofeedback paradigm in which they were intermittently provided with a visual representation of the difference signal between the average activation of the salience and executive control networks, and tasked with attempting to self-regulate this signal. Our results show that, given feedback about their performance over three training sessions, participants were able to (1) learn strategies to differentially control the balance between SN and ECN activation on demand, as well as (2) successfully transfer this newly learned skill to a situation where they (a) did not receive any feedback anymore, and (b) were exposed to an acute stressor in form of the prospect of a mild electric stimulation. The current study hence constitutes an important first successful demonstration of neurofeedback training based on stress-related large-scale network balance - a novel approach that has the potential to train control over the central response to stressors in real-life and could build the foundation for future clinical interventions that aim at increasing resilience.

HighlightsO_LIAcute stress affects the allocation of neural resources between large-scale brain networks
C_LIO_LIWe provide a first successful demonstration of neurofeedback training based on stress-related large-scale brain networks
C_LIO_LINovel approach has the potential to train control over central response to stressors in real-life
C_LIO_LICould build foundation for future clinical interventions to increase resilience
C_LI


title: Longitudinal characterization of neuroanatomical changes in the Fischer 344 rat brain during normal aging and between sexes

abstract:
Animal models are widely used to study the pathophysiology of disease and to evaluate the efficacy of novel interventions, crucial steps towards improving disease outcomes in humans. The Fischer 344 (F344) wildtype rat is a common experimental background strain for transgenic models of disease and is also one of the most frequently used models in aging research. Despite frequency of use, characterization of neuroanatomical change with age has not been performed in the F344 rat. To this end, we present a comprehensive longitudinal examination of morphometric change in 73 brain regions and at a voxel-wise level during normative aging in a mixed-sex cohort of F344 rats. We identified age- and sex-related changes in regions such as the cortex, hippocampus, cingulum, caudoputamen, and nucleus accumbens, which are implicated in memory and motor control circuits frequently affected by aging and neurodegenerative disease. These findings provide a baseline for neuroanatomical changes associated with aging in male and female F344 rats, to which data from transgenic models or other background strains can be compared.

HIGHLIGHTSO_LIIn vivo magnetic resonance imaging reveals altered neuroanatomy in aging Fischer rats
C_LIO_LILinear and curvilinear age effects exist in both grey and white matter structures
C_LIO_LISex differences are primarily seen in grey matter structures
C_LIO_LIThis study clarifies normal aging trajectories across 73 brain regions in both sexes
C_LIO_LIImproved understanding of normal aging will inform future pathological aging studies
C_LI


title: A species-specific signature residue in the PB2 subunit of the bat influenza virus polymerase restricts viral RNA synthesis.

abstract:
Bat influenza A viruses (H17N10 and H18N11) are genetically distant from conventional influenza A viruses and replicates poorly in non-bat hosts species. However, the reason behind the lower replication fitness of these viruses are yet to be elucidated. In this work, we have identified species-specific signature residues, present in viral PB2 protein, which is a major determinant of polymerase fitness in human, avian and bat cell lines. Through extensive sequence and structural comparison between the bat and non-bat influenza virus RNA polymerases, we have identified a previously uncharacterized PB2-282 residue, which is serine in bat virus PB2 protein but harbours highly conserved glutamic acid in conventional influenza A viruses. Introduction of these bat specific signatures in the polymerase of a human adapted strain of influenza A/H1N1 virus drastically reduces its polymerase activity and replication efficiency in cell lines of human, bat and canine origin. In contrast, introduction of the human specific signatures in bat virus PB2 (H17N10), significantly enhances its function in the context of a chimeric RNA polymerase. Interestingly, the PB2-282 resides within an evolutionary conserved "S-E-S" motif present across different genera of influenza viruses but is replaced with a "S-S-T" motif in bat influenza viruses, indicating that this E to S transition may serve as a species-specific adaptation signature that modulates the activity of bat virus polymerase in other host species.

ImportanceRecent isolation of influenza A like viruses (H17N10 and H18N11) in bats raised concerns about their potential of zoonotic transmission in human. Here we present species-specific signature residues present in the bat influenza virus polymerase, which may act as critical modulators of bat virus propagation in non-bat host species. We utilize bioinformatics based comparative analysis followed by functional screening in order to identify the PB2-282nd position, which harbors a highly conserved glutamic acid in conventional influenza A viruses, but contains an unusual serine in case of bat influenza viruses. Human adapted polymerase, harboring bat specific signature (PB2-S282) performs poorly, while bat PB2 protein harboring human specific signature (PB2-E282) shows increased fitness in human cells. Together, our data identifies novel species-specific signatures present within the influenza virus polymerase that may serve as a key factor in the adaptation of influenza viruses from bat to non-bat host species and vice versa.

category: microbiology


title: Dynamics of Salmonella inoculated during rearing ofblack soldier fly larvae (Hermetia illucens) on chicken feed

abstract: The black soldier fly is currently the most produced edible insect on industrial scale, with its larval stage being processed into animal feed as the main application. As this insect species enters the feed and food chain, good hygiene and monitoring practices are needed to avoid the entrance of foodborne pathogens via the larvae. However, insufficient data on the risk of such introductions via industrial larvae production are available. To address this gap, a range of rearing trials were conducted in which the substrate, chicken feed, was inoculated with different levels of Salmonella and in which total viable counts and Salmonella counts were determined during the following days. The outgrowth of Salmonella was slower in those experiments with a lower initial contamination level than in experiments with a higher level. No significant reducing effect originating from the larvae on the substrate Salmonella counts was observed, in contrast to previous studies using other substrates. Our study also revealed that airborne transmission of Salmonella is possible under rearing conditions corresponding to those applied at industrial production sites. Based on our results, we recommend insect producers to use substrate ingredients free of Salmonella, and not to count on the antimicrobial activities that BSFL may exert in some situations towards food pathogens. More inoculation studies using other Salmonella serotypes, other zoonotic bacteria, other substrates, larvae of other ages and including variations on rearing protocols are needed in order to obtain a general view on the dynamics of food pathogens in this insect species and to support comprehensive risk assessments.


title: pUL21 is a viral phosphatase adaptor that promotes herpes simplex virus replication and spread

abstract: The herpes simplex virus (HSV)-1 protein pUL21 is essential for efficient virus replication and dissemination. While pUL21 has been shown to promote multiple steps of virus assembly and spread, the molecular basis of its function remained unclear. Here we identify that pUL21 is a virus-encoded adaptor of protein phosphatase 1 (PP1). pUL21 directs the dephosphorylation of cellular and virus proteins, including components of the viral nuclear egress complex, and we define a conserved non-canonical linear motif in pUL21 that is essential for PP1 recruitment. In vitro evolution experiments reveal that pUL21 directly antagonises the activity of the virus-encoded kinase pUS3, with growth and spread of pUL21 PP1-binding mutant viruses being restored when pUS3 activity is disrupted. This study shows that virus-directed phosphatase activity is essential for efficient herpesvirus assembly and spread, highlighting the fine balance between kinase and phosphatase activity required for optimal virus replication.


title: Divergent Acyl Carrier Protein Mediates Mitochondrial Fe-S Cluster Biosynthesis in Malaria Parasites

abstract:
Plasmodium falciparum malaria parasites are early-diverging eukaryotes with many unusual metabolic adaptations. Understanding these adaptations will give insight into parasite evolution and unveil new parasite-specific drug targets. Most eukaryotic cells retain a mitochondrial fatty acid synthesis (FASII) pathway whose acyl carrier protein (mACP) and 4-phosphopantetheine (Ppant) prosthetic group provide a soluble scaffold for acyl chain synthesis. In yeast and humans, mACP also functions to biochemically couple FASII activity to electron transport chain (ETC) assembly and Fe-S cluster biogenesis. In contrast to most eukaryotes, the Plasmodium mitochondrion lacks FASII enzymes yet curiously retains a divergent mACP lacking a Ppant group. We report that ligand-dependent knockdown of mACP is lethal to parasites, indicating an essential FASII-independent function. Decyl-ubiquinone rescues parasites temporarily from death, suggesting a dominant dysfunction of the mitochondrial ETC followed by broader cellular defects. Biochemical studies reveal that Plasmodium mACP binds and stabilizes the Isd11-Nfs1 complex required for Fe-S cluster biosynthesis, despite lacking the Ppant group required for this association in other eukaryotes, and knockdown of parasite mACP causes loss of both Nfs1 and the Rieske Fe-S protein in ETC Complex III. This work reveals that Plasmodium parasites have evolved to decouple mitochondrial Fe-S cluster biogenesis from FASII activity, and this adaptation is a shared metabolic feature of other Apicomplexan pathogens, including Toxoplasma and Babesia. This discovery also highlights the ancient, fundamental role of ACP in mitochondrial Fe-S cluster biogenesis and unveils an evolutionary driving force to retain this interaction with ACP independent of its eponymous function in FASII.

Significance StatementPlasmodium malaria parasites are single-celled eukaryotes that evolved unusual metabolic adaptations. Parasites require a mitochondrion for blood-stage viability, but essential functions beyond the electron transport chain are sparsely understood. Unlike yeast and human cells, the Plasmodium mitochondrion lacks fatty acid synthesis enzymes but retains a divergent acyl carrier protein (mACP) incapable of tethering acyl groups. Nevertheless, mACP is essential for parasite viability by binding and stabilizing the core mitochondrial Fe-S cluster biogenesis complex via a divergent molecular interface lacking an acyl-pantetheine group that contrasts with other eukaryotes. This discovery unveils an essential metabolic adaptation in Plasmodium and other human parasites that decouples mitochondrial Fe-S cluster biogenesis from fatty acid synthesis and evolved at or near the emergence of Apicomplexan parasitism.


title: Divergent Acyl Carrier Protein Decouples Mitochondrial Fe-S Cluster Biogenesis from Fatty Acid Synthesis in Malaria Parasites


title: One Health Investigation of SARS-CoV-2 Infection and Seropositivity among Pets in Households with Confirmed Human COVID-19 Cases - Utah and Wisconsin, 2020

abstract:
BackgroundApproximately 67% of U.S. households have pets. Limited data are available on SARS-CoV-2 in pets. We assessed SARS-CoV-2 infection in pet cohabitants as a sub-study of an ongoing COVID-19 household transmission investigation.

MethodsMammalian pets from households with [≥]1 person with laboratory-confirmed COVID-19 were eligible for inclusion from April-May 2020. Demographic/exposure information, oropharyngeal, nasal, rectal, and fur swabs, feces, and blood were collected from enrolled pets and tested by rRT-PCR and virus neutralization assays.

FindingsWe enrolled 37 dogs and 19 cats from 34 of 41 eligible households. All oropharyngeal, nasal, and rectal swabs tested negative by rRT-PCR; one dogs fur swabs (2%) tested positive by rRT-PCR at the first animal sampling. Among 47 pets with serological results from 30 households, eight (17%) pets (4 dogs, 4 cats) from 6 (20%) households had detectable SARS-CoV-2 neutralizing antibodies. In households with a seropositive pet, the proportion of people with laboratory-confirmed COVID-19 was greater (median 79%; range: 40-100%) compared to households with no seropositive pet (median 37%; range: 13-100%) (p=0.01). Thirty-three pets with serologic results had frequent daily contact ([≥]1 hour) with the human index patient before the persons COVID-19 diagnosis. Of these 33 pets, 14 (42%) had decreased contact with the human index patient after diagnosis and none (0%) were seropositive; of the 19 (58%) pets with continued contact, 4 (21%) were seropositive.

InterpretationsSeropositive pets likely acquired infection from humans, which may occur more frequently than previously recognized. People with COVID-19 should restrict contact with animals.

FundingCenters for Disease Control and Prevention, U.S. Department of Agriculture