Publications
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(2025) Advanced Science. 2414297. Abstract
Abstract Accumulated evidence of transgenerational inheritance of epigenetic and symbiotic changes raises fundamental questions about the possible types, significance and duration of impacts on the population, as well as whether, and under which conditions, the inheritance of non-genetic changes confers long-term advantage to the population. To address these questions, a population epigenetics model of individuals undergoing stochastic changes and/or induced responses that are transmitted to the offspringis introduced. Potentially adaptive and maladaptive responses are represented, respectively, by environmentally driven changes that reduce and increase the selective pressure. Analytic solutions in a simplified case of populations that are exposed to either periodic or progressively deteriorating environments shows that acquisition and transmission of non-genetic changes that alleviate the selective pressure confer long-term advantage and may facilitate escape from extinction. Systematic analysis of outcomes as a function of population properties further identifies a non-traditional regime of adaptation mediated by stochastic changes that are rapidly acquired within a lifetime. Contrasting model predictions with experimental findings shows that inheritance of dynamically acquired changes enables rapid adaptation to unforeseen challenges and can account for population dynamics that is either unexpected or beyond the scope of traditional models.
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(2025) Advanced Science. 2414297. Abstract
Accumulated evidence of transgenerational inheritance of epigenetic and symbiotic changes raises fundamental questions about the possible types, significance and duration of impacts on the population, as well as whether, and under which conditions, the inheritance of non-genetic changes confers long-term advantage to the population. To address these questions, a population epigenetics model of individuals undergoing stochastic changes and/or induced responses that are transmitted to the offspringis introduced. Potentially adaptive and maladaptive responses are represented, respectively, by environmentally driven changes that reduce and increase the selective pressure. Analytic solutions in a simplified case of populations that are exposed to either periodic or progressively deteriorating environments shows that acquisition and transmission of non-genetic changes that alleviate the selective pressure confer long-term advantage and may facilitate escape from extinction. Systematic analysis of outcomes as a function of population properties further identifies a non-traditional regime of adaptation mediated by stochastic changes that are rapidly acquired within a lifetime. Contrasting model predictions with experimental findings shows that inheritance of dynamically acquired changes enables rapid adaptation to unforeseen challenges and can account for population dynamics that is either unexpected or beyond the scope of traditional models.
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(2024) PLoS Biology. 22, 5, e3002629. Abstract[All authors]
ADUes:pPiteleassigencoifnicfairnmttpharotaglrlehseasdininuglnedveelrssatraenrdepinrgeseepntigedecnoertrieccrtelyp:rogramming of cells, the mechanistic basis of "organ reprogramming"by (epi-)gene-environment interactions remained largely obscure. Here, we use the ether-induced haltere-to-wing transformations in Drosophila as a model for epigenetic "reprogramming"at the whole organism level. Our findings support a mechanistic chain of events explaining why and how brief embryonic exposure to ether leads to haltere-to-wing transformations manifested at the larval stage and on. We show that ether interferes with protein integrity in the egg, leading to altered deployment of Hsp90 and widespread repression of Trithorax-mediated establishment of active H3K4me3 chromatin marks throughout the genome. Despite this global reduction, Ubx targets and wing development genes preferentially retain higher levels of H3K4me3 that predisposes these genes for later up-regulation in the larval haltere disc, hence the wing-like outcome. Consistent with compromised protein integrity during the exposure, the penetrance of bithorax transformations increases by genetic or chemical reduction of Hsp90 function. Moreover, joint reduction in Hsp90 and trx gene dosage can cause bithorax transformations even without exposure to ether, supporting an underlying epistasis between Hsp90 and trx loss-of-functions. These findings implicate environmental disruption of protein integrity at the onset of histone methylation with altered epigenetic regulation of developmental patterning genes. The emerging picture provides a unique example wherein the alleviation of the Hsp90 "capacitor function"by the environment drives a morphogenetic shift towards an ancestral-like body plan. The morphogenetic impact of chaperone response during a major setup of epigenetic patterns may be a general scheme for organ reprogramming by environmental cues.
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(2023) iScience. 26, 9, 107632. Abstract
Microbial enzymes can address diverse challenges such as degradation of toxins. However, if the function of interest does not confer a sufficient fitness effect on the producer, the enzymatic function cannot be improved in the host cells by a conventional selection scheme. To overcome this limitation, we propose an alternative scheme, termed \u201cpartner-assisted artificial selection\u201d (PAAS), wherein the population of enzyme producers is assisted by function-dependent feedback from an accessory population. Simulations investigating the efficiency of toxin degradation reveal that this strategy supports selection of improved degradation performance, which is robust to stochasticity in the model parameters. We observe that conventional considerations still apply in PAAS: more restrictive bottlenecks lead to stronger selection but add uncertainty. Overall, we offer a guideline for successful implementation of PAAS and highlight its potentials and limitations.
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(2022) STAR Protocols. 3, 2, 101253. Abstract
Drosophila gut microbiome in flies has been shown to have a systemic influence on energy production by the host and the energetic investment in growth and reproduction. Here we describe a protocol for studying the mechanisms responsible for this remote regulation by gut bacteria. This protocol enables whole-body and ovary-specific quantification of energy-storing molecules as well as identification of host metabolites and pathways that are regulated by gut microbiome-derived factors. Similar procedures are applicable to additional treatments and genetic manipulations. For complete details on the use and execution of this protocol, please refer to Gnainsky et al. (2021).
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(2021) Frontiers in endocrinology (Lausanne). 12, 635405. Abstract[All authors]
BackgroundCell therapy of diabetes aims at restoring the physiological control of blood glucose by transplantation of functional pancreatic islet cells. A potentially unlimited source of cells for such transplantations would be islet cells derived from an in vitro differentiation of human pluripotent stem cells (hESC/hiPSC). The islet-like clusters (ILC) produced by the known differentiation protocols contain various cell populations. Among these, the β-cells that express both insulin and the transcription factor Nkx6.1 seem to be the most efficient to restore normoglycemia in diabetes animal models. Our aim was to find markers allowing selection of these efficient cells.MethodsFunctional Cell-Capture Screening (FCCS) was used to identify markers that preferentially capture the cells expressing both insulin and Nkx6.1, from hESC-derived ILC cells. In order to test whether selection for such markers could improve cell therapy in diabetic mouse models, we used ILC produced from a clinical-grade line of hESC by a refined differentiation protocol adapted to up-scalable bioreactors. Re-aggregated MACS sorted cells were encapsulated in microspheres made of alginate modified to reduce foreign body reaction. Implantation was done intraperitoneally in STZ-treated C57BL/6 immuno-competent mice.ResultsCD49A (integrin alpha1) was identified by FCCS as a marker for cells that express insulin (or C-peptide) as well as Nkx6.1 in ILC derived by hESC differentiation. The ILC fraction enriched in CD49A+ cells rapidly reduced glycemia when implanted in diabetic mice, whereas mice receiving the CD49A depleted population remained highly diabetic. CD49A-enriched ILC cells also produced higher levels of human C-peptide in the blood of transplanted mice. However, the difference between CD49A-enriched and total ILC cells remained small. Another marker, CD26 (DPP4), was identified by FCCS as binding insulin-expressing cells which are Nkx6.1 negative. Depletion of CD26+ cells followed by enrichment for CD49A+ cells increased insulin+/Nkx6.1+ cells fraction to ~70%. The CD26-/CD49A+ enriched ILC exhibited improved function over non-sorted ILC or CD49A+ cells in diabetic mice and maintain prolonged blood C-peptide levels.ConclusionsRefining the composition of ILC differentiated from hPSC by negative selection to remove cells expressing CD26 and positive selection for CD49A expressing cells could enable more effective cell therapy of diabetes.
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(2021) Cell Reports. 34, 1, 108583. Abstract[All authors]
Gut microbiota have been shown to promote oogenesis and fecundity, but the mechanistic basis of remote influence on oogenesis remained unknown. Here, we report a systemic mechanism of influence mediated by bacterial-derived supply of mitochondrial coenzymes. Removal of microbiota decreased mitochondrial activity and ATP levels in the whole-body and ovary, resulting in repressed oogenesis. Similar repression was caused by RNA-based knockdown of mitochondrial function in ovarian follicle cells. Reduced mitochondrial function in germ-free (GF) females was reversed by bacterial recolonization or supplementation of riboflavin, a precursor of FAD and FMN. Metabolomics analysis of GF females revealed a decrease in oxidative phosphorylation and FAD levels and an increase in metabolites that are degraded by FAD-dependent enzymes (e.g., amino and fatty acids). Riboflavin supplementation opposed this effect, elevating mitochondrial function, ATP, and oogenesis. These findings uncover a bacterial-mitochondrial axis of influence, linking gut bacteria with systemic regulation of host energy and reproduction.
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(2018) Biology Direct. 13, 1, 24. Abstract
Background: The relatively fast selection of symbiotic bacteria within hosts and the potential transmission of these bacteria across generations of hosts raise the question of whether interactions between host and bacteria support emergent adaptive capabilities beyond those of germ-free hosts.Results: To investigate possibilities for emergent adaptations that may distinguish composite host-microbiome systems from germ-free hosts, we introduce a population genetics model of a host-microbiome system with vertical transmission of bacteria. The host and its bacteria are jointly exposed to a toxic agent, creating a toxic stress that can be alleviated by selection of resistant individuals and by secretion of a detoxification agent ("detox"). We show that toxic exposure in one generation of hosts leads to selection of resistant bacteria, which in turn, increases the toxic tolerance of the host's offspring. Prolonged exposure to toxin over many host generations promotes anadditional form of emergent adaptation due to selection of hosts based on detox produced by their bacterial community as a whole (as opposed to properties of individual bacteria).Conclusions: These findings show that interactions between pure Darwinian selections of host and its bacteria can give rise to emergent adaptive capabilities, including Lamarckian-like adaptation of the host-microbiome system.Reviewers: This article was reviewed by Eugene Koonin, Yuri Wolf and Philippe Huneman.
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(2018) BioEssays. 40, 4, 1700018. Abstract
It is becoming increasingly clear that most, if not all, animals and plants are associated with a diverse array of resident gut microbiota. This symbiosis is regulated by host-microbiome interactions which influence the development, homeostasis, adaptation and evolution of the host. Recent evidence indicated that these interactions can also affect the host germline and have a potential of supporting transgenerational effects, including inheritance of acquired characteristics. Taken together, the influence of gut bacteria on the host soma and germline could potentially give rise to emergent phenotypes, which may be partially inherited by three distinguishable modes of transgenerational influence of gut bacteria: 1) "soma-to-soma" 2) "soma-to-germline" and 3) "soma-germline-soma". Here, we discuss these possibilities in light of evidence supporting bacterial-mediated modes of transgenerational inheritance.
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(2018) Microbiome. 6, 17. Abstract
Background: Most of our knowledge about the remarkable microbial diversity on Earth comes from sequencing the 16S rRNA gene. The use of next-generation sequencing methods has increased sample number and sequencing depth, but the read length of the most widely used sequencing platforms today is quite short, requiring the researcher to choose a subset of the gene to sequence (typically 16-33% of the total length). Thus, many bacteria may share the same amplified region, and the resolution of profiling is inherently limited. Platforms that offer ultra-long read lengths, whole genome shotgun sequencing approaches, and computational frameworks formerly suggested by us and by others all allow different ways to circumvent this problem yet suffer various shortcomings. There is a need for a simple and low-cost 16S rRNA gene-based profiling approach that harnesses the short read length to provide a much larger coverage of the gene to allow for high resolution, even in harsh conditions of low bacterial biomass and fragmented DNA. Results: This manuscript suggests Short MUltiple Regions Framework (SMURF), a method to combine sequencing results from different PCR-amplified regions to provide one coherent profiling. The de facto amplicon length is the total length of all amplified regions, thus providing much higher resolution compared to current techniques. Computationally, the method solves a convex optimization problem that allows extremely fast reconstruction and requires only moderate memory. We demonstrate the increase in resolution by in silico simulations and by profiling two mock mixtures and real-world biological samples. Reanalyzing a mock mixture from the Human Microbiome Project achieved about twofold improvement in resolution when combing two independent regions. Using a custom set of six primer pairs spanning about 1200 bp (80%) of the 16S rRNA gene, we were able to achieve ~ 100-fold improvement in resolution compared to a single region, over a mock mixture of common human gut bacterial isolates. Finally, the profiling of a Drosophila melanogaster microbiome using the set of six primer pairs provided a ~ 100-fold increase in resolution and thus enabling efficient downstream analysis. Conclusions: SMURF enables the identification of near full-length 16S rRNA gene sequences in microbial communities, having resolution superior compared to current techniques. It may be applied to standard sample preparation protocols with very little modifications. SMURF also paves the way to high-resolution profiling of low-biomass and fragmented DNA, e.g., in the case of formalin-fixed and paraffin-embedded samples, fossil-derived DNA, or DNA exposed to other degrading conditions. The approach is not restricted to combining amplicons of the 16S rRNA gene and may be applied to any set of amplicons, e.g., in multilocus sequence typing (MLST).
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(2017) Nature Communications. 8, 14826. Abstract
The capacity of cells and organisms to respond to challenging conditions in a repeatable manner is limited by a finite repertoire of pre-evolved adaptive responses. Beyond this capacity, cells can use exploratory dynamics to cope with a much broader array of conditions. However, the process of adaptation by exploratory dynamics within the lifetime of a cell is not well understood. Here we demonstrate the feasibility of exploratory adaptation in a high-dimensional network model of gene regulation. Exploration is initiated by failure to comply with a constraint and is implemented by random sampling of network configurations. It ceases if and when the network reaches a stable state satisfying the constraint. We find that successful convergence (adaptation) in high dimensions requires outgoing network hubs and is enhanced by their auto-regulation. The ability of these empirically validated features of gene regulatory networks to support exploratory adaptation without fine-tuning, makes it plausible for biological implementation.
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(2016) Nature Communications. 7, 11280. Abstract
Unlike vertically transmitted endosymbionts, which have broad effects on their host's germ line, the extracellular gut microbiota is transmitted horizontally and is not known to influence the germ line. Here we provide evidence supporting the influence of these gut bacteria on the germ line of Drosophila melanogaster. Removal of the gut bacteria represses oogenesis, expedites maternal-to-zygotic-transition in the offspring and unmasks hidden phenotypic variation in mutants. We further show that the main impact on oogenesis is linked to the lack of gut Acetobacter species, and we identify the Drosophila Aldehyde dehydrogenase (Aldh) gene as an apparent mediator of repressed oogenesis in Acetobacter-depleted flies. The finding of interactions between the gut microbiota and the germ line has implications for reproduction, developmental robustness and adaptation.
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(2015) Biology Direct. 10, 68. Abstract
BACKGROUND: During the lifetime of an organism, every individual encounters many combinations of diverse changes in the somatic genome, epigenome and microbiome. This gives rise to many novel combinations of internal failures which are unique to each individual. How any individual can tolerate this high load of new, individual-specific scenarios of failure is not clear. While stress-induced plasticity and hidden variation have been proposed as potential mechanisms of tolerance, the main conceptual problem remains unaddressed, namely: how largely non-beneficial random variation can be rapidly and safely organized into net benefits to every individual.PRESENTATION OF THE HYPOTHESIS: We propose an organizational principle which explains how every individual can alleviate a high load of novel stressful scenarios using many random variations in flexible and inherently less harmful traits. Random changes which happen to reduce stress, benefit the organism and decrease the drive for additional changes. This adaptation (termed 'Adaptive Improvisation') can be further enhanced, propagated, stabilized and memorized when beneficial changes reinforce themselves by auto-regulatory mechanisms. This principle implicates stress not only in driving diverse variations in cells tissues and organs, but also in organizing these variations into adaptive outcomes. Specific (but not exclusive) examples include stress reduction by rapid exchange of mobile genetic elements (or exosomes) in unicellular, and rapid changes in the symbiotic microorganisms of animals. In all cases, adaptive changes can be transmitted across generations, allowing rapid improvement and assimilation in a few generations.TESTING THE HYPOTHESIS: We provide testable predictions derived from the hypothesis.IMPLICATIONS OF THE HYPOTHESIS: The hypothesis raises a critical, but thus far overlooked adaptation problem and explains how random variation can self-organize to confer a wide range of individual-specific adaptations beyond the existing outcomes of natural selection. It portrays gene regulation as an inseparable synergy between natural selection and adaptation by improvisation. The latter provides a basis for Lamarckian adaptation that is not limited to a specific mechanism and readily accounts for the remarkable resistance of tumors to treatment.
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(2015) Biochimica et Biophysica Acta - Gene Regulatory Mechanisms. 1849, 4, p. 462-466 Abstract
Organisms have to be sufficiently robust to environmental and genetic perturbations, yet plastic enough to cope with stressful scenarios to which they are not fully adapted. How this apparent conflict between robustness and plasticity is resolved at the cellular and whole organism levels is not clear. Here we review and discuss evidence in flies suggesting that the environment can modulate the balance between robustness and plasticity. The outcomes of this modulation can vary from mild sensitizations that are hardly noticeable, to overt qualitative changes in phenotype. The effects could be at both the cellular and whole organism levels and can include cellular de-/trans-differentiation ('Cellular reprogramming') and gross disfigurements such as homeotic transformations (Tissue/whole organism reprogramming'). When the stress is mild enough, plastic changes in some processes may prevent drastic changes in more robust traits such as cell identity and tissue integrity. However, when the stress is sufficiently severe, this buffering may no longer be able to prevent such overt changes, and the resulting phenotypic variability could be subjected to selection and might assist survival at the population level. This article is part of a Special Issue entitled: Stress as a fundamental theme in cell plasticity.
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(2015) PLoS ONE. 10, 2, e0115100. Abstract
The current world-wide epidemic of diabetes has prompted attempts to generate new sources of insulin-producing cells for cell replacement therapy. An inherent challenge in many of these strategies is the lack of cell-surface markers permitting isolation and characterization of specific cell types from differentiating stem cell populations. Here we introduce an iterative proteomics procedure allowing tag-free isolation of cell types based on their function. Our method detects and associates specific cell-surface markers with particular cell functionality by coupling cell capture on antibody arrays with immunofluorescent labeling. Using this approach in an iterative manner, we discovered marker combinations capable of enriching for discrete pancreatic cell subtypes from human islets of Langerhans: insulin-producing beta cells (CD9high/CD56+), glucagon-producing alpha cells (CD9-/CD56+) and trypsin-producing acinar cells (CD9-/CD56-). This strategy may assist future beta cell research and the development of diagnostic tools for diabetes. It can also be applied more generally for function-based purification of desired cell types from other limited and heterogeneous biological samples.
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(2014) Journal Of Physiology-London. 592, 11, p. 2343-2355 Abstract
Transgenerational persistence of parental responses to environmental stimuli has been reported in various organisms, but the underlying mechanisms remain underexplored. In one of these reported examples, we have shown that exposure of fly larvae to G418 antibiotic leads to non-Mendelian inheritance of ectopic induction of certain developmental genes. Here we investigate if this inheritance involves changes in mRNA composition within the early, maternal-stage offspring embryos of exposed flies. Exposure to G418 in F1 modified the maternal RNA levels of many genes in their early (F2) embryos. This includes reduction of maternal Polycomb group genes which persisted in the following generation of embryos (F3). To investigate the functional meaning of this reduction, we compared genetically normal embryos of Polycomb mutant females to normal embryos of normal females. Analysis with two different alleles of Polycomb, Pc1 and Pc3, revealed that maternal reduction in Polycomb gene dosage has a positive influence on the inheritance of induced expression. Together, this shows that exposure to G418 stress reduces the maternal levels of Polycomb in the offspring embryos and this reduction contributes to the inheritance of induced expression.
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(2014) Frontiers in Genetics. 5, JUN, 168. Abstract
The microbiome is known to have a profound effect on the development, physiology and health of its host. Whether and how it also contributes to evolutionary diversification of the host is, however, unclear. Here we hypothesize that disruption of the microbiome by new stressful environments interferes with host-microbe co-adaptation, contributes to host destabilization, and can drive irreversible changes in the host prior to its genetic adaptation. This hypothesis is based on three presumptions: (1) the microbiome consists of heritable partners which contribute to the stability (canalization) of host development and physiology in frequently encountered environments, (2) upon encountering a stressful new environment, the microbiome adapts much faster than the host, and (3) this differential response disrupts cooperation, contributes to host destabilization and promotes reciprocal changes in the host and its microbiome. This dynamic imbalance relaxes as the host and its microbiome establish a new equilibrium state in which they are adapted to one another and to the altered environment. Over long time in this new environment, the changes in the microbiome contribute to the canalization of the altered state. This scenario supports stability of the adapted patterns, while promoting variability which may be beneficial in new stressful conditions, thus allowing the organism to balance stability and flexibility based on contextual demand. Additionally, interaction between heritable microbial and epigenetic/physiological changes can promote new outcomes which persist over a wide range of timescales. A sufficiently persistent stress can further induce irreversible changes in the microbiome which may permanently alter the organism prior to genetic changes in the host. Epigenetic and microbial changes therefore provide a potential infrastructure for causal links between immediate responses to new environments and longer-term establishment of evolutionary adaptations.
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(2014) Frontiers in Genetics. 5, FEB, 27. Abstract
Commensal gut bacteria in many species including flies are integral part of their host, and are known to influence its development and homeostasis within generation. Here we report an unexpected impact of host-microbe interactions, which mediates multi-generational, non-Mendelian inheritance of a stress-induced phenotype. We have previously shown that exposure of fly larvae to G418 antibiotic induces transgenerationally heritable phenotypes, including a delay in larval development, gene induction in the gut and morphological changes. We now show that G418 selectively depletes commensal Acetobacter species and that this depletion explains the heritable delay, but not the inheritance of the other phenotypes. Notably, the inheritance of the delay was mediated by a surprising trans-generational effect. Specifically, bacterial removal from F1 embryos did not induce significant delay in F1 larvae, but nonetheless led to a considerable delay in F2. This effect maintains a delay induced by bacterial-independent G418 toxicity to the host. In line with these findings, reintroduction of isolated Acetobacter species prevented the inheritance of the delay. We further show that this prevention is partly mediated by vitamin B2 (Riboflavin) produced by these bacteria; exogenous Riboflavin led to partial prevention and inhibition of Riboflavin synthesis compromised the ability of the bacteria to prevent the inheritance. These results identify host-microbe interactions as a hitherto unrecognized factor capable of mediating non-Mendelian inheritance of a stress-induced phenotype.
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(2013) Nucleic Acids Research. 41, 22, p. e205 Abstract
The emergence of massively parallel sequencing technology has revolutionized microbial profiling, allowing the unprecedented comparison of microbial diversity across time and space in a wide range of host-associated and environmental ecosystems. Although the high-throughput nature of such methods enables the detection of low-frequency bacteria, these advances come at the cost of sequencing read length, limiting the phylogenetic resolution possible by current methods. Here, we present a generic approach for integrating short reads from large genomic regions, thus enabling phylogenetic resolution far exceeding current methods. The approach is based on a mapping to a statistical model that is later solved as a constrained optimization problem. We demonstrate the utility of this method by analyzing human saliva and Drosophila samples, using Illumina single-end sequencing of a 750 bp amplicon of the 16S rRNA gene. Phylogenetic resolution is significantly extended while reducing the number of falsely detected bacteria, as compared with standard single-region Roche 454 Pyrosequencing. Our approach can be seamlessly applied to simultaneous sequencing of multiple genes providing a higher resolution view of the composition and activity of complex microbial communities.
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(2013) Stem Cells. 31, 10, p. 2266-2272 Abstract
microRNAs of the miR-290-295 family are selectively expressed at high levels in mouse embryonic stem cells (mESCs) and have established roles in regulating self-renewal. However, the potential influence of these microRNAs on cell fate acquisition during differentiation has been overlooked. Here, we show that miR-290-295 regulate the propensity of mESCs to acquire specific fates. We generated a new miR-290-295-null mESC model, which exhibits increased propensity to generate ectoderm, at the expense of endoderm and mesoderm lineages. We further found that in wild-type cells, miR-290-295 repress Pax6 and ectoderm differentiation; accordingly, Pax6 knockdown partially rescues the mESCs differentiation impairment that is caused by loss of miR-290-295. Thus, in addition to regulating self-renewal, the large reservoir of miR-290-295 in undifferentiated mESCs fine-tunes the expression of master transcriptional factors, such as Pax6, thereby regulating the equilibrium of fate acquisition by mESC descendants. Stem Cells 2013;31:2266-2272
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(2012) Proceedings of the National Academy of Sciences of the United States of America. 109, 39, p. 15811-15816 Abstract
Neurodegenerative diseases constitute a class of illnesses marked by pathological protein aggregation in the brains of affected individuals. Although these disorders are invariably characterized by the degeneration of highly specific subpopulations of neurons, protein aggregation occurs in all cells, which indicates that toxicity arises only in particular cell biological contexts. Aggregation-associated disorders are unified by a common cell biological feature: the deposition of the culprit proteins in inclusion bodies. The precise function of these inclusions remains unclear. The starting point for uncovering the origins of disease pathology must therefore be a thorough understanding of the general cell biological function of inclusions and their potential role inmodulating the consequences of aggregation. Here, we show that in human cells certain aggregate inclusions are active compartments. We find that toxic aggregates localize to one of these compartments, the juxtanuclear quality control compartment (JUNQ), and interfere with its quality control function. The accumulation of SOD1G93A aggregates sequesters Hsp70, preventing the delivery of misfolded proteins to the proteasome. Preventing the accumulation of SOD1G93A in the JUNQ by enhancing its sequestration in an insoluble inclusion reduces the harmful effects of aggregation on cell viability.
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(2012) Rna-A Publication Of The Rna Society. 18, 9, p. 1612-1623 Abstract
While microRNAs have been shown to copurify with polysomes, their relative fraction in the translation pool (polysome occupancy) has not yet been measured. Here, we introduce a high-throughput method for quantifying polysome occupancies of hundreds of microRNAs and use it to investigate factors affecting these occupancies. Analysis in human embryonic stem cells (hESCs) and foreskin fibroblasts (hFFs) revealed microRNA-specific preferences for low, medium, or high polysome occupancy. Bioinformatics and functional analysis based on overexpression of endogenous and chimeric microRNAs showed that the polysome occupancy of microRNAs is specified by its mature sequence and depends on the choice of seed. Nuclease treatment further suggested that the differential occupancy of the microRNAs reflects interactions with their mRNA targets. Indeed, analysis of microNRdmRNA duplexes showed that pairs involving high occupancy microRNAs exhibit significantly higher binding energy compared to pairs with low occupancy microRNAs. Since mRNAs reside primarily in polysomes, strong interactions lead to high association of microRNAs with polysomes and vice versa for weak interactions. Comparison between hESCs and hFFs data revealed that hESCs tend to express lower occupancy microRNAs, suggesting that cell type-dependent translational features may be affected by expression of a particular set of microRNAs. Published by Cold Spring Harbor Laboratory Press.
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(2012) Molecular & Cellular Proteomics. 11, 9, p. 586-595 Abstract
Heterogeneity, shortage of material, and lack of progenitor-specific cell surface markers are major obstacles to elucidating the mechanisms underlying developmental processes. Here we report a proteomics platform that alleviates these difficulties and demonstrate its effectiveness in fractionating heterogeneous cultures of early endoderm derived from human embryonic stem cells. The approach, designated differential cell-capture antibody array, is based on highly parallel, comparative screening of live cell populations using hundreds of antibodies directed against cell-surface antigens. We used this platform to fractionate the hitherto unresolved early endoderm compartment of CXCR4+ cells and identify several endoderm (CD61+ and CD63+) and non-endoderm (CD271+, CD49F+, CD44+ and B2M+) sub-populations. We provide evidence that one of these sub-populations, CD61+, is directly derived from CXCR4+ cells, displays characteristic kinetics of emergence, and exhibits a distinct gene expression profile. The results demonstrate the potential of the cell-capture antibody array as a powerful proteomics tool for detailed dissection of heterogeneous cellular systems.
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(2012) Stem Cells. 30, 6, p. 1097-1108 Abstract
While experimentally induced arrest of human embryonic stem cells (hESCs) in G1 has been shown to stimulate differentiation, it remains unclear whether the unperturbed G1 phase in hESCs is causally related to differentiation. Here, we use centrifugal elutriation to isolate and investigate differentiation propensities of hESCs in different phases of their cell cycle. We found that isolated G1 cells exhibit higher differentiation propensity compared with S and G2 cells, and they differentiate at low cell densities even under self-renewing conditions. This differentiation of G1 cells was partially prevented in dense cultures of these cells and completely abrogated in coculture with S and G2 cells. However, coculturing without cell-to-cell contact did not rescue the differentiation of G1 cells. Finally, we show that the subset of G1 hESCs with reduced phosphorylation of retinoblastoma has the highest propensity to differentiate and that the differentiation is preceded by cell cycle arrest. These results provide direct evidence for increased propensity of hESCs to differentiate in G1 and suggest a role for neighboring cells in preventing differentiation of hESCs as they pass through a differentiation sensitive, G1 phase.
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(2012) Nature Biotechnology. 30, 6, p. 531-+ Abstract
To identify early populations of committed progenitors derived from human embryonic stem cells (hESCs), we screened self-renewing, BMP4-treated and retinoic acid-treated cultures with >400 antibodies recognizing cell-surface antigens. Sorting of >30 subpopulations followed by transcriptional analysis of developmental genes identified four distinct candidate progenitor groups. Subsets detected in self-renewing cultures, including CXCR4(+) cells, expressed primitive endoderm genes. Expression of Cxcr4 in primitive endoderm was confirmed in visceral endoderm of mouse embryos. BMP4-induced progenitors exhibited gene signatures of mesoderm, trophoblast and vascular endothelium, suggesting correspondence to gastrulation-stage primitive streak, chorion and allantois precursors, respectively. Functional studies in vitro and in vivo confirmed that ROR2(+) cells produce mesoderm progeny, APA(+) cells generate syncytiotrophoblasts and CD87(+) cells give rise to vasculature. The same progenitor classes emerged during the differentiation of human induced pluripotent stem cells (hiPSCs). These markers and progenitors provide tools for purifying human tissue-regenerating progenitors and for studying the commitment of pluripotent stem cells to lineage progenitors.
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(2012) Cell Reports. 1, 5, p. 528-542 Abstract
Developing organisms have evolved a wide range of mechanisms for coping with recurrent environmental challenges. How they cope with rare or unforeseen challenges is, however, unclear as are the implications to their unchallenged offspring. Here, we investigate these questions by confronting the development of the fly, . D. . melanogaster, with artificial tissue distributions of toxic stress that are not expected to occur during fly development. We show that under a wide range of toxic scenarios, this challenge can lead to modified development that may coincide with increased tolerance to an otherwise lethal condition. Part of this response was mediated by suppression of Polycomb group genes, which in turn leads to derepression of developmental regulators and their expression in new domains. Importantly, some of the developmental alterations were epigenetically inherited by subsequent generations of unchallenged offspring. These results show that the environment can induce alternative patterns of development that are stable across multiple generations.
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(2011) Molecular Systems Biology. 7, 529. Abstract[All authors]
Transcriptional responses to extracellular stimuli involve tuning the rates of transcript production and degradation. Here, we show that the time-dependent profiles of these rates can be inferred from simultaneous measurements of precursor mRNA (pre-mRNA) and mature mRNA profiles. Transcriptome-wide measurements demonstrate that genes with similar mRNA profiles often exhibit marked differences in the amplitude and onset of their production rate. The latter is characterized by a large dynamic range, with a group of genes exhibiting an unexpectedly strong transient production overshoot, thereby accelerating their induction and, when combined with time-dependent degradation, shaping transient responses with precise timing and amplitude.