Showing papers in "PLOS Biology in 2008"

Mapping the Structural Core of Human Cerebral Cortex

Abstract: Structurally segregated and functionally specialized regions of the human cerebral cortex are interconnected by a dense network of cortico-cortical axonal pathways. By using diffusion spectrum imaging, we noninvasively mapped these pathways within and across cortical hemispheres in individual human participants. An analysis of the resulting large-scale structural brain networks reveals a structural core within posterior medial and parietal cerebral cortex, as well as several distinct temporal and frontal modules. Brain regions within the structural core share high degree, strength, and betweenness centrality, and they constitute connector hubs that link all major structural modules. The structural core contains brain regions that form the posterior components of the human default network. Looking both within and outside of core regions, we observed a substantial correspondence between structural connectivity and resting-state functional connectivity measured in the same participants. The spatial and topological centrality of the core within cortex suggests an important role in functional integration.

Senescence-Associated Secretory Phenotypes Reveal Cell-Nonautonomous Functions of Oncogenic RAS and the p53 Tumor Suppressor

Abstract: Cellular senescence suppresses cancer by arresting cell proliferation, essentially permanently, in response to oncogenic stimuli, including genotoxic stress. We modified the use of antibody arrays to provide a quantitative assessment of factors secreted by senescent cells. We show that human cells induced to senesce by genotoxic stress secrete myriad factors associated with inflammation and malignancy. This senescence-associated secretory phenotype (SASP) developed slowly over several days and only after DNA damage of sufficient magnitude to induce senescence. Remarkably similar SASPs developed in normal fibroblasts, normal epithelial cells, and epithelial tumor cells after genotoxic stress in culture, and in epithelial tumor cells in vivo after treatment of prostate cancer patients with DNA-damaging chemotherapy. In cultured premalignant epithelial cells, SASPs induced an epithelial-mesenchyme transition and invasiveness, hallmarks of malignancy, by a paracrine mechanism that depended largely on the SASP factors interleukin (IL)-6 and IL-8. Strikingly, two manipulations markedly amplified, and accelerated development of, the SASPs: oncogenic RAS expression, which causes genotoxic stress and senescence in normal cells, and functional loss of the p53 tumor suppressor protein. Both loss of p53 and gain of oncogenic RAS also exacerbated the promalignant paracrine activities of the SASPs. Our findings define a central feature of genotoxic stress-induced senescence. Moreover, they suggest a cell-nonautonomous mechanism by which p53 can restrain, and oncogenic RAS can promote, the development of age-related cancer by altering the tissue microenvironment.

The Pervasive Effects of an Antibiotic on the Human Gut Microbiota, as Revealed by Deep 16S rRNA Sequencing

Abstract: The human intestinal microbiota is essential to the health of the host and plays a role in nutrition, development, metabolism, pathogen resistance, and regulation of immune responses. Antibiotics may disrupt these coevolved interactions, leading to acute or chronic disease in some individuals. Our understanding of antibiotic-associated disturbance of the microbiota has been limited by the poor sensitivity, inadequate resolution, and significant cost of current research methods. The use of pyrosequencing technology to generate large numbers of 16S rDNA sequence tags circumvents these limitations and has been shown to reveal previously unexplored aspects of the “rare biosphere.” We investigated the distal gut bacterial communities of three healthy humans before and after treatment with ciprofloxacin, obtaining more than 7,000 full-length rRNA sequences and over 900,000 pyrosequencing reads from two hypervariable regions of the rRNA gene. A companion paper in PLoS Genetics (see Huse et al., doi: 10.1371/journal.pgen.1000255) shows that the taxonomic information obtained with these methods is concordant. Pyrosequencing of the V6 and V3 variable regions identified 3,300–5,700 taxa that collectively accounted for over 99% of the variable region sequence tags that could be obtained from these samples. Ciprofloxacin treatment influenced the abundance of about a third of the bacterial taxa in the gut, decreasing the taxonomic richness, diversity, and evenness of the community. However, the magnitude of this effect varied among individuals, and some taxa showed interindividual variation in the response to ciprofloxacin. While differences of community composition between individuals were the largest source of variability between samples, we found that two unrelated individuals shared a surprising degree of community similarity. In all three individuals, the taxonomic composition of the community closely resembled its pretreatment state by 4 weeks after the end of treatment, but several taxa failed to recover within 6 months. These pervasive effects of ciprofloxacin on community composition contrast with the reports by participants of normal intestinal function and with prior assumptions of only modest effects of ciprofloxacin on the intestinal microbiota. These observations support the hypothesis of functional redundancy in the human gut microbiota. The rapid return to the pretreatment community composition is indicative of factors promoting community resilience, the nature of which deserves future investigation.

The bacterial symbiont Wolbachia induces resistance to RNA viral infections in Drosophila melanogaster.

Abstract: Wolbachia are vertically transmitted, obligatory intracellular bacteria that infect a great number of species of arthropods and nematodes. In insects, they are mainly known for disrupting the reproductive biology of their hosts in order to increase their transmission through the female germline. In Drosophila melanogaster, however, a strong and consistent effect of Wolbachia infection has not been found. Here we report that a bacterial infection renders D. melanogaster more resistant to Drosophila C virus, reducing the load of viruses in infected flies. We identify these resistance-inducing bacteria as Wolbachia. Furthermore, we show that Wolbachia also increases resistance of Drosophila to two other RNA virus infections (Nora virus and Flock House virus) but not to a DNA virus infection (Insect Iridescent Virus 6). These results identify a new major factor regulating D. melanogaster resistance to infection by RNA viruses and contribute to the idea that the response of a host to a particular pathogen also depends on its interactions with other microorganisms. This is also, to our knowledge, the first report of a strong beneficial effect of Wolbachia infection in D. melanogaster. The induced resistance to natural viral pathogens may explain Wolbachia prevalence in natural populations and represents a novel Wolbachia–host interaction.

Towards an Integrated Framework for Assessing the Vulnerability of Species to Climate Change

Abstract: [Extract] Global climate change threatens global biodiversity, ecosystem function, and human well-being, with thousands of publications demonstrating impacts across a wide diversity of taxonomic groups, ecosystems, economics, and social structure. A review by Hughes [1] identified many of the ways that organisms may be affected by and/or respond to climate change. Since then, there has been a dramatic increase in the number of case studies attesting to ecological impacts [2], prompting several recent reviews on the subject (e.g., [3–6]). Several global meta-analyses confirm the pervasiveness of the global climate change "fingerprint" across continents, ecosystems, processes, and species [7–9]. Some studies have predicted increasingly severe future impacts with potentially high extinction rates in natural systems around the world [10,11]. Responding to this threat will require a concerted, multi-disciplinary, multi-scale, multi-taxon research effort that improves our predictive capacity to identify and prioritise vulnerable species in order to inform governments of the seriousness of the threat and to facilitate conservation adaptation and management [12,13]. If we are to minimise global biodiversity loss, we need significant decreases in global emissions to be combined with environmental management that is guided by sensible prioritisation of relative vulnerability. That is, we need to determine which species, habitats, and ecosystems will be most vulnerable, exactly what aspects of their ecological and evolutionary biology determine their vulnerability, and what we can do about managing this vulnerability and minimising the realised impacts. There is an emerging literature on specific traits that promote vulnerability under climate change (e.g., thermal tolerance [14]) as well as a broad literature on the traits that influence species' vulnerability generally (e.g., review by [15]). Less is known about the various mechanisms for either ecological or evolutionary adaptation to climate change, although it is increasingly recognised as a vital component of assessing vulnerability [16,17].

Mapping the Genetic Architecture of Gene Expression in Human Liver

Abstract: Genetic variants that are associated with common human diseases do not lead directly to disease, but instead act on intermediate, molecular phenotypes that in turn induce changes in higher-order disease traits. Therefore, identifying the molecular phenotypes that vary in response to changes in DNA and that also associate with changes in disease traits has the potential to provide the functional information required to not only identify and validate the susceptibility genes that are directly affected by changes in DNA, but also to understand the molecular networks in which such genes operate and how changes in these networks lead to changes in disease traits. Toward that end, we profiled more than 39,000 transcripts and we genotyped 782,476 unique single nucleotide polymorphisms (SNPs) in more than 400 human liver samples to characterize the genetic architecture of gene expression in the human liver, a metabolically active tissue that is important in a number of common human diseases, including obesity, diabetes, and atherosclerosis. This genome-wide association study of gene expression resulted in the detection of more than 6,000 associations between SNP genotypes and liver gene expression traits, where many of the corresponding genes identified have already been implicated in a number of human diseases. The utility of these data for elucidating the causes of common human diseases is demonstrated by integrating them with genotypic and expression data from other human and mouse populations. This provides much-needed functional support for the candidate susceptibility genes being identified at a growing number of genetic loci that have been identified as key drivers of disease from genome-wide association studies of disease. By using an integrative genomics approach, we highlight how the gene RPS26 and not ERBB3 is supported by our data as the most likely susceptibility gene for a novel type 1 diabetes locus recently identified in a large-scale, genome-wide association study. We also identify SORT1 and CELSR2 as candidate susceptibility genes for a locus recently associated with coronary artery disease and plasma low-density lipoprotein cholesterol levels in the process.

SARS-Coronavirus Replication Is Supported by a Reticulovesicular Network of Modified Endoplasmic Reticulum

Abstract: Positive-strand RNA viruses, a large group including human pathogens such as SARS-coronavirus (SARS-CoV), replicate in the cytoplasm of infected host cells. Their replication complexes are commonly associated with modified host cell membranes. Membrane structures supporting viral RNA synthesis range from distinct spherular membrane invaginations to more elaborate webs of packed membranes and vesicles. Generally, their ultrastructure, morphogenesis, and exact role in viral replication remain to be defined. Poorly characterized double-membrane vesicles (DMVs) were previously implicated in SARS-CoV RNA synthesis. We have now applied electron tomography of cryofixed infected cells for the three-dimensional imaging of coronavirus-induced membrane alterations at high resolution. Our analysis defines a unique reticulovesicular network of modified endoplasmic reticulum that integrates convoluted membranes, numerous interconnected DMVs (diameter 200-300 nm), and "vesicle packets" apparently arising from DMV merger. The convoluted membranes were most abundantly immunolabeled for viral replicase subunits. However, double-stranded RNA, presumably revealing the site of viral RNA synthesis, mainly localized to the DMV interior. Since we could not discern a connection between DMV interior and cytosol, our analysis raises several questions about the mechanism of DMV formation and the actual site of SARS-CoV RNA synthesis. Our data document the extensive virus-induced reorganization of host cell membranes into a network that is used to organize viral replication and possibly hide replicating RNA from antiviral defense mechanisms. Together with biochemical studies of the viral enzyme complex, our ultrastructural description of this "replication network" will aid to further dissect the early stages of the coronavirus life cycle and its virus-host interactions.

Promotion of Reprogramming to Ground State Pluripotency by Signal Inhibition

Abstract: Induced pluripotent stem (iPS) cells are generated from somatic cells by genetic manipulation. Reprogramming entails multiple transgene integrations and occurs apparently stochastically in rare cells over many days. Tissue stem cells may be subject to less-stringent epigenetic restrictions than other cells and might therefore be more amenable to deprogramming. We report that brain-derived neural stem (NS) cells acquire undifferentiated morphology rapidly and at high frequency after a single round of transduction with reprogramming factors. However, critical attributes of true pluripotency—including stable expression of endogenous Oct4 and Nanog, epigenetic erasure of X chromosome silencing in female cells, and ability to colonise chimaeras—were not attained. We therefore applied molecularly defined conditions for the derivation and propagation of authentic pluripotent stem cells from embryos. We combined dual inhibition (2i) of mitogen-activated protein kinase signalling and glycogen synthase kinase-3 (GSK3) with the self-renewal cytokine leukaemia inhibitory factor (LIF). The 2i/LIF condition induced stable up-regulation of Oct4 and Nanog, reactivation of the X chromosome, transgene silencing, and competence for somatic and germline chimaerism. Using 2i /LIF, NS cell reprogramming required only 1–2 integrations of each transgene. Furthermore, transduction with Sox2 and c-Myc is dispensable, and Oct4 and Klf4 are sufficient to convert NS cells into chimaera-forming iPS cells. These findings demonstrate that somatic cell state influences requirements for reprogramming and delineate two phases in the process. The ability to capture pre-pluripotent cells that can advance to ground state pluripotency simply and with high efficiency opens a door to molecular dissection of this remarkable phenomenon.

Control of jasmonate biosynthesis and senescence by miR319 targets.

Abstract: Considerable progress has been made in identifying the targets of plant microRNAs, many of which regulate the stability or translation of mRNAs that encode transcription factors involved in development In most cases, it is unknown, however, which immediate transcriptional targets mediate downstream effects of the microRNA-regulated transcription factors We identified a new process controlled by the miR319-regulated clade of TCP (TEOSINTE BRANCHED/CYCLOIDEA/PCF) transcription factor genes In contrast to other miRNA targets, several of which modulate hormone responses, TCPs control biosynthesis of the hormone jasmonic acid Furthermore, we demonstrate a previously unrecognized effect of TCPs on leaf senescence, a process in which jasmonic acid has been proposed to be a critical regulator We propose that miR319-controlled TCP transcription factors coordinate two sequential processes in leaf development: leaf growth, which they negatively regulate, and leaf senescence, which they positively regulate

A Novel CpG Island Set Identifies Tissue-Specific Methylation at Developmental Gene Loci

Abstract: CpG islands (CGIs) are dense clusters of CpG sequences that punctuate the CpG-deficient human genome and associate with many gene promoters. As CGIs also differ from bulk chromosomal DNA by their frequent lack of cytosine methylation, we devised a CGI enrichment method based on nonmethylated CpG affinity chromatography. The resulting library was sequenced to define a novel human blood CGI set that includes many that are not detected by current algorithms. Approximately half of CGIs were associated with annotated gene transcription start sites, the remainder being intra- or intergenic. Using an array representing over 17,000 CGIs, we established that 6%–8% of CGIs are methylated in genomic DNA of human blood, brain, muscle, and spleen. Inter- and intragenic CGIs are preferentially susceptible to methylation. CGIs showing tissue-specific methylation were overrepresented at numerous genetic loci that are essential for development, including HOX and PAX family members. The findings enable a comprehensive analysis of the roles played by CGI methylation in normal and diseased human tissues.

WikiPathways: Pathway Editing for the People

Abstract: The exponential growth of diverse types of biological data presents the research community with an unprecedented challenge and opportunity. The challenge is to stay afloat in the flood of biological data, keeping it as accessible, up-to-date, and integrated as possible. The opportunity is to cultivate new models of data curation and exchange that take advantage of direct participation by a greater portion of the community. This combination of challenge and opportunity is especially relevant to the task of collecting biological pathway information. Pathways are critical to understanding the functions of individual genes and proteins in terms of systems and processes that contribute to normal physiology and to disease. Each biological pathway must be hewn from a mass of biological information distributed across multiple publications and databases. The particular challenge of pathway curation is amplified, because pathways are often presented as static images that are not amenable to computation, integration, or data exchange. Furthermore, pathway experts are distributed throughout the world, and most have limited time to learn about complex databases that need their expertise. This challenge can be met by taking the opportunity to develop a new community-based model for pathway curation. One way to engage the community is with a wiki model, as exemplified by Wikipedia [1]. We see the potential for a wiki-based pathway curation resource, coupled with an embedded graphical pathway editing tool, to meet the growing challenge presented by the influx of biological data and to provide an innovative example of content curation by the biology community (Figure 1). Figure 1 Two Models for Managing Biological Data

Diverse RNA-Binding Proteins Interact with Functionally Related Sets of RNAs, Suggesting an Extensive Regulatory System

Abstract: RNA-binding proteins (RBPs) have roles in the regulation of many post-transcriptional steps in gene expression, but relatively few RBPs have been systematically studied. We searched for the RNA targets of 40 proteins in the yeast Saccharomyces cerevisiae: a selective sample of the approximately 600 annotated and predicted RBPs, as well as several proteins not annotated as RBPs. At least 33 of these 40 proteins, including three of the four proteins that were not previously known or predicted to be RBPs, were reproducibly associated with specific sets of a few to several hundred RNAs. Remarkably, many of the RBPs we studied bound mRNAs whose protein products share identifiable functional or cytotopic features. We identified specific sequences or predicted structures significantly enriched in target mRNAs of 16 RBPs. These potential RNA-recognition elements were diverse in sequence, structure, and location: some were found predominantly in 39-untranslated regions, others in 59-untranslated regions, some in coding sequences, and many in two or more of these features. Although this study only examined a small fraction of the universe of yeast RBPs, 70% of the mRNA transcriptome had significant associations with at least one of these RBPs, and on average, each distinct yeast mRNA interacted with three of the RBPs, suggesting the potential for a rich, multidimensional network of regulation. These results strongly suggest that combinatorial binding of RBPs to specific recognition elements in mRNAs is a pervasive mechanism for multi-dimensional regulation of their post-transcriptional fate.

Spinal Cord Injury Reveals Multilineage Differentiation of Ependymal Cells

Abstract: Spinal cord injury often results in permanent functional impairment. Neural stem cells present in the adult spinal cord can be expanded in vitro and improve recovery when transplanted to the injured spinal cord, demonstrating the presence of cells that can promote regeneration but that normally fail to do so efficiently. Using genetic fate mapping, we show that close to all in vitro neural stem cell potential in the adult spinal cord resides within the population of ependymal cells lining the central canal. These cells are recruited by spinal cord injury and produce not only scar-forming glial cells, but also, to a lesser degree, oligodendrocytes. Modulating the fate of ependymal progeny after spinal cord injury may offer an alternative to cell transplantation for cell replacement therapies in spinal cord injury.

Sparse Representation of Sounds in the Unanesthetized Auditory Cortex

Abstract: How do neuronal populations in the auditory cortex represent acoustic stimuli? Although sound-evoked neural responses in the anesthetized auditory cortex are mainly transient, recent experiments in the unanesthetized preparation have emphasized subpopulations with other response properties. To quantify the relative contributions of these different subpopulations in the awake preparation, we have estimated the representation of sounds across the neuronal population using a representative ensemble of stimuli. We used cell-attached recording with a glass electrode, a method for which single-unit isolation does not depend on neuronal activity, to quantify the fraction of neurons engaged by acoustic stimuli (tones, frequency modulated sweeps, white-noise bursts, and natural stimuli) in the primary auditory cortex of awake head-fixed rats. We find that the population response is sparse, with stimuli typically eliciting high firing rates (>20 spikes/second) in less than 5% of neurons at any instant. Some neurons had very low spontaneous firing rates (<0.01 spikes/second). At the other extreme, some neurons had driven rates in excess of 50 spikes/second. Interestingly, the overall population response was well described by a lognormal distribution, rather than the exponential distribution that is often reported. Our results represent, to our knowledge, the first quantitative evidence for sparse representations of sounds in the unanesthetized auditory cortex. Our results are compatible with a model in which most neurons are silent much of the time, and in which representations are composed of small dynamic subsets of highly active neurons.

Going, going, gone : is animal migration disappearing

Abstract: Animal migration surely ranks as one of nature's most visible and widespread phenomena. Every minute of every day, somewhere, some place, animals are on the move. The migrants span the animal kingdom, from whales and warblers to dragonflies and salamanders. But is migration an endangered phenomenon? Around the world, many of the most spectacular migrations have either disappeared due to human activities or are in steep decline. Those of us living in eastern North America can no longer experience the flocks of millions of passenger pigeons that temporarily obscured the sun as they migrated to and from their breeding grounds. Nor can residents of the Great Plains climb to the top of a hill and gaze down up hundreds of thousands of bison trekking across the prairies, as was possible less than two centuries ago. Even the less iconic migrations show signs of trouble. Birdwatchers in North America and Europe, for example, complain that fewer songbirds are returning each spring from their winter quarters in Latin America and Africa, respectively. Indeed, a recent continent-wide analysis of European breeding birds concluded that long-distance migrants (i.e., those species that breed in Europe but winter in sub-Saharan Africa) have suffered sustained and often severe population declines, more so than related nonmigratory species [1]. In central Asia, the number of saiga, a peculiar migratory antelope of the dry steppe grasslands and semi-desert, has dropped by over 95% in the past two decades, from over one million to fewer than 50,000 [2]. The causes of all these declines vary depending on the species and the locale, but in general, the threats to migrants fall into four nonexclusive categories: habitat destruction, the creation of obstacles and barriers such as dams and fences, overexploitation, and climate change. Most of the migrants are in little immediate danger of extinction; rather, they are becoming less and less common. Thus, birdwatchers can still see all of the species of migratory songbirds they seek each spring; they simply have to work harder to do so. Bison still roam national parks and private ranches in the American West, but today's herds number in the hundreds or low thousands, rather than the hundreds of thousands or millions. And there are still lots of salmon to catch off the coast of Norway or British Columbia—just not as many as there used to be. The question thus arises: Given the panoply of environmental problems we now face, is the fading glory of migration really a significant issue? We would argue that it is. Protecting the abundance of migrants is the key to protecting the ecological importance of migration. As the number of migrants declines, so too do many of the most important ecological properties and services associated with them. Consider the case of salmon in the Pacific Northwest. The seven species of salmon and seagoing trout in this region share a similar life history strategy: as young fish (smolt), they leave their natal rivers and head to the sea where, aided by the productivity of the ocean, they increase tremendously in size and weight. After a year or two at sea, they return to their natal rivers to spawn, whereupon they die. By migrating upstream, spawning, and dying, they transfer nutrients from the ocean to the rivers. A portion of the nutrients is delivered in the form of feces, sperm, and eggs from the living fish; much more comes from the decaying carcasses of the adults. Phosphorus and nitrogen from salmon carcasses enhance the growth of phytoplankton and zooplankton in the rivers, which provide food for smaller fish, including young salmon. Thus, salmon fry are literally sustained by their parents. Prior to European settlement, 160–226 million kilograms of salmon migrated each year up the rivers of Washington, Idaho, Oregon, and California. Today, after decades of dam construction, overfishing, water withdrawals for irrigation, logging, and streamside grazing by livestock, salmon populations have plummeted. The total biomass of spawning salmon in the Pacific Northwest is now estimated to be only 12–14 million kilograms. Gresh et al. [3] have calculated that the rivers of the Northwest receive just 6%–7% of the marine-derived nitrogen and phosphorus they once received from the abundant salmon population. How this shortfall may be affecting the ecology of the region's rivers or adjacent farmlands is largely unknown. We can imagine an analogous situation developing with respect to migratory birds. Each spring, more than 30,000 tons of migratory songbirds migrate from their wintering grounds in Latin America and the Caribbean to their breeding grounds in the United States and Canada. (This biomass value is derived by combining breeding population totals from the North American Landbird Conservation Plan with species-specific weights from various sources.) If we assume these birds consume 10%–35% of their body weight per day in insects (roughly matching the requirements of a 100-gram bird and a 10-gram bird, respectively), then they are eating anywhere from 3,000–10,500 tons of insects per day. (During the breeding season, when the birds are feeding offspring, these figures would be much higher.) Several studies have shown that birds reduce insect populations in temperate forests, thus raising the question of whether ongoing declines in migratory birds pose a threat to the health of our forests and farmlands. Similarly, one wonders how the ecology of the Serengeti would change if its migratory population of wildebeest (exceeding 1 million individuals) were to collapse, given the major role these animals surely play in terms of consuming herbaceous vegetation and redistributing nutrients via their urine and dung (Figure 1). Figure 1 Migratory Wildebeest in the Serengeti

High functional diversity in Mycobacterium tuberculosis driven by genetic drift and human demography.

Abstract: Mycobacterium tuberculosis infects one third of the human world population and kills someone every 15 seconds. For more than a century, scientists and clinicians have been distinguishing between the human- and animal-adapted members of the M. tuberculosis complex (MTBC). However, all human-adapted strains of MTBC have traditionally been considered to be essentially identical. We surveyed sequence diversity within a global collection of strains belonging to MTBC using seven megabase pairs of DNA sequence data. We show that the members of MTBC affecting humans are more genetically diverse than generally assumed, and that this diversity can be linked to human demographic and migratory events. We further demonstrate that these organisms are under extremely reduced purifying selection and that, as a result of increased genetic drift, much of this genetic diversity is likely to have functional consequences. Our findings suggest that the current increases in human population, urbanization, and global travel, combined with the population genetic characteristics of M. tuberculosis described here, could contribute to the emergence and spread of drug-resistant tuberculosis.

Shifting Baselines, Local Impacts, and Global Change on Coral Reefs

Abstract: The striking health of remote coral reefs provides clear evidence that protection from local overfishing and pollution can help mitigate the impacts of global warming.

Is Sleep Essential

Abstract: No current hypothesis can explain why animals need to sleep. Yet, sleep is universal, tightly regulated, and cannot be deprived without deleterious consequences. This suggests that searching for a core function of sleep, particularly at the cellular level, is still a worthwhile exercise.

Global Reorganization of Replication Domains During Embryonic Stem Cell Differentiation

Abstract: DNA replication in mammals is regulated via the coordinate firing of clusters of replicons that duplicate megabase-sized chromosome segments at specific times during S-phase. Cytogenetic studies show that these “replicon clusters” coalesce as subchromosomal units that persist through multiple cell generations, but the molecular boundaries of such units have remained elusive. Moreover, the extent to which changes in replication timing occur during differentiation and their relationship to transcription changes has not been rigorously investigated. We have constructed high-resolution replication-timing profiles in mouse embryonic stem cells (mESCs) before and after differentiation to neural precursor cells. We demonstrate that chromosomes can be segmented into multimegabase domains of coordinate replication, which we call “replication domains,” separated by transition regions whose replication kinetics are consistent with large originless segments. The molecular boundaries of replication domains are remarkably well conserved between distantly related ESC lines and induced pluripotent stem cells. Unexpectedly, ESC differentiation was accompanied by the consolidation of smaller differentially replicating domains into larger coordinately replicated units whose replication time was more aligned to isochore GC content and the density of LINE-1 transposable elements, but not gene density. Replication-timing changes were coordinated with transcription changes for weak promoters more than strong promoters, and were accompanied by rearrangements in subnuclear position. We conclude that replication profiles are cell-type specific, and changes in these profiles reveal chromosome segments that undergo large changes in organization during differentiation. Moreover, smaller replication domains and a higher density of timing transition regions that interrupt isochore replication timing define a novel characteristic of the pluripotent state.

Identifying neural drivers with functional MRI: an electrophysiological validation.

Abstract: Whether functional magnetic resonance imaging (fMRI) allows the identification of neural drivers remains an open question of particular importance to refine physiological and neuropsychological models of the brain, and/or to understand neurophysiopathology. Here, in a rat model of absence epilepsy showing spontaneous spike-and-wave discharges originating from the first somatosensory cortex (S1BF), we performed simultaneous electroencephalographic (EEG) and fMRI measurements, and subsequent intracerebral EEG (iEEG) recordings in regions strongly activated in fMRI (S1BF, thalamus, and striatum). fMRI connectivity was determined from fMRI time series directly and from hidden state variables using a measure of Granger causality and Dynamic Causal Modelling that relates synaptic activity to fMRI. fMRI connectivity was compared to directed functional coupling estimated from iEEG using asymmetry in generalised synchronisation metrics. The neural driver of spike-and-wave discharges was estimated in S1BF from iEEG, and from fMRI only when hemodynamic effects were explicitly removed. Functional connectivity analysis applied directly on fMRI signals failed because hemodynamics varied between regions, rendering temporal precedence irrelevant. This paper provides the first experimental substantiation of the theoretical possibility to improve interregional coupling estimation from hidden neural states of fMRI. As such, it has important implications for future studies on brain connectivity using functional neuroimaging.

Transcription factors bind thousands of active and inactive regions in the Drosophila blastoderm.

Abstract: Identifying the genomic regions bound by sequence-specific regulatory factors is central both to deciphering the complex DNA cis-regulatory code that controls transcription in metazoans and to determining the range of genes that shape animal morphogenesis. We used whole-genome tiling arrays to map sequences bound in Drosophila melanogaster embryos by the six maternal and gap transcription factors that initiate anterior–posterior patterning. We find that these sequence-specific DNA binding proteins bind with quantitatively different specificities to highly overlapping sets of several thousand genomic regions in blastoderm embryos. Specific high- and moderate-affinity in vitro recognition sequences for each factor are enriched in bound regions. This enrichment, however, is not sufficient to explain the pattern of binding in vivo and varies in a context-dependent manner, demonstrating that higher-order rules must govern targeting of transcription factors. The more highly bound regions include all of the over 40 well-characterized enhancers known to respond to these factors as well as several hundred putative new cis-regulatory modules clustered near developmental regulators and other genes with patterned expression at this stage of embryogenesis. The new targets include most of the microRNAs (miRNAs) transcribed in the blastoderm, as well as all major zygotically transcribed dorsal–ventral patterning genes, whose expression we show to be quantitatively modulated by anterior–posterior factors. In addition to these highly bound regions, there are several thousand regions that are reproducibly bound at lower levels. However, these poorly bound regions are, collectively, far more distant from genes transcribed in the blastoderm than highly bound regions; are preferentially found in protein-coding sequences; and are less conserved than highly bound regions. Together these observations suggest that many of these poorly bound regions are not involved in early-embryonic transcriptional regulation, and a significant proportion may be nonfunctional. Surprisingly, for five of the six factors, their recognition sites are not unambiguously more constrained evolutionarily than the immediate flanking DNA, even in more highly bound and presumably functional regions, indicating that comparative DNA sequence analysis is limited in its ability to identify functional transcription factor targets.

Fish invasions in the world's river systems: when natural processes are blurred by human activities.

Abstract: Because species invasions are a principal driver of the human-induced biodiversity crisis, the identification of the major determinants of global invasions is a prerequisite for adopting sound conservation policies. Three major hypotheses, which are not necessarily mutually exclusive, have been proposed to explain the establishment of non-native species: the “human activity” hypothesis, which argues that human activities facilitate the establishment of non-native species by disturbing natural landscapes and by increasing propagule pressure; the “biotic resistance” hypothesis, predicting that species-rich communities will readily impede the establishment of non-native species; and the “biotic acceptance” hypothesis, predicting that environmentally suitable habitats for native species are also suitable for non-native species. We tested these hypotheses and report here a global map of fish invasions (i.e., the number of non-native fish species established per river basin) using an original worldwide dataset of freshwater fish occurrences, environmental variables, and human activity indicators for 1,055 river basins covering more than 80% of Earth's surface. First, we identified six major invasion hotspots where non-native species represent more than a quarter of the total number of species. According to the World Conservation Union, these areas are also characterised by the highest proportion of threatened fish species. Second, we show that the human activity indicators account for most of the global variation in non-native species richness, which is highly consistent with the “human activity” hypothesis. In contrast, our results do not provide support for either the “biotic acceptance” or the “biotic resistance” hypothesis. We show that the biogeography of fish invasions matches the geography of human impact at the global scale, which means that natural processes are blurred by human activities in driving fish invasions in the world's river systems. In view of our findings, we fear massive invasions in developing countries with a growing economy as already experienced in developed countries. Anticipating such potential biodiversity threats should therefore be a priority.

Riding the Wave: Reconciling the Roles of Disease and Climate Change in Amphibian Declines

Abstract: We review the evidence for the role of climate change in triggering disease outbreaks of chytridiomycosis, an emerging infectious disease of amphibians. Both climatic anomalies and disease-related extirpations are recent phenomena, and effects of both are especially noticeable at high elevations in tropical areas, making it difficult to determine whether they are operating separately or synergistically. We compiled reports of amphibian declines from Lower Central America and Andean South America to create maps and statistical models to test our hypothesis of spatiotemporal spread of the pathogen Batrachochytrium dendrobatidis (Bd), and to update the elevational patterns of decline in frogs belonging to the genus Atelopus. We evaluated claims of climate change influencing the spread of Bd by including error into estimates of the relationship between air temperature and last year observed. Available data support the hypothesis of multiple introductions of this invasive pathogen into South America and subsequent spread along the primary Andean cordilleras. Additional analyses found no evidence to support the hypothesis that climate change has been driving outbreaks of amphibian chytridiomycosis, as has been posited in the climate-linked epidemic hypothesis. Future studies should increase retrospective surveys of museum specimens from throughout the Andes and should study the landscape genetics of Bd to map fine-scale patterns of geographic spread to identify transmission routes and processes.

A specificity map for the PDZ domain family.

Abstract: PDZ domains are protein–protein interaction modules that recognize specific C-terminal sequences to assemble protein complexes in multicellular organisms. By scanning billions of random peptides, we accurately map binding specificity for approximately half of the over 330 PDZ domains in the human and Caenorhabditis elegans proteomes. The domains recognize features of the last seven ligand positions, and we find 16 distinct specificity classes conserved from worm to human, significantly extending the canonical two-class system based on position −2. Thus, most PDZ domains are not promiscuous, but rather are fine-tuned for specific interactions. Specificity profiling of 91 point mutants of a model PDZ domain reveals that the binding site is highly robust, as all mutants were able to recognize C-terminal peptides. However, many mutations altered specificity for ligand positions both close and far from the mutated position, suggesting that binding specificity can evolve rapidly under mutational pressure. Our specificity map enables the prediction and prioritization of natural protein interactions, which can be used to guide PDZ domain cell biology experiments. Using this approach, we predicted and validated several viral ligands for the PDZ domains of the SCRIB polarity protein. These findings indicate that many viruses produce PDZ ligands that disrupt host protein complexes for their own benefit, and that highly pathogenic strains target PDZ domains involved in cell polarity and growth.

Strong Purifying Selection in Transmission of Mammalian Mitochondrial DNA

Abstract: There is an intense debate concerning whether selection or demographics has been most important in shaping the sequence variation observed in modern human mitochondrial DNA (mtDNA). Purifying selection is thought to be important in shaping mtDNA sequence evolution, but the strength of this selection has been debated, mainly due to the threshold effect of pathogenic mtDNA mutations and an observed excess of new mtDNA mutations in human population data. We experimentally addressed this issue by studying the maternal transmission of random mtDNA mutations in mtDNA mutator mice expressing a proofreading-deficient mitochondrial DNA polymerase. We report a rapid and strong elimination of nonsynonymous changes in protein-coding genes; the hallmark of purifying selection. There are striking similarities between the mutational patterns in our experimental mouse system and human mtDNA polymorphisms. These data show strong purifying selection against mutations within mtDNA protein-coding genes. To our knowledge, our study presents the first direct experimental observations of the fate of random mtDNA mutations in the mammalian germ line and demonstrates the importance of purifying selection in shaping mitochondrial sequence diversity.

Mirror-Induced Behavior in the Magpie (Pica pica): Evidence of Self-Recognition

Abstract: Comparative studies suggest that at least some bird species have evolved mental skills similar to those found in humans and apes. This is indicated by feats such as tool use, episodic-like memory, and the ability to use one's own experience in predicting the behavior of conspecifics. It is, however, not yet clear whether these skills are accompanied by an understanding of the self. In apes, self-directed behavior in response to a mirror has been taken as evidence of self-recognition. We investigated mirror-induced behavior in the magpie, a songbird species from the crow family. As in apes, some individuals behaved in front of the mirror as if they were testing behavioral contingencies. When provided with a mark, magpies showed spontaneous mark-directed behavior. Our findings provide the first evidence of mirror self-recognition in a non-mammalian species. They suggest that essential components of human self-recognition have evolved independently in different vertebrate classes with a separate evolutionary history.

Dynamic Remodeling of Individual Nucleosomes Across a Eukaryotic Genome in Response to Transcriptional Perturbation

Abstract: The eukaryotic genome is packaged as chromatin with nucleosomes comprising its basic structural unit, but the detailed structure of chromatin and its dynamic remodeling in terms of individual nucleosome positions has not been completely defined experimentally for any genome. We used ultra-high–throughput sequencing to map the remodeling of individual nucleosomes throughout the yeast genome before and after a physiological perturbation that causes genome-wide transcriptional changes. Nearly 80% of the genome is covered by positioned nucleosomes occurring in a limited number of stereotypical patterns in relation to transcribed regions and transcription factor binding sites. Chromatin remodeling in response to physiological perturbation was typically associated with the eviction, appearance, or repositioning of one or two nucleosomes in the promoter, rather than broader region-wide changes. Dynamic nucleosome remodeling tends to increase the accessibility of binding sites for transcription factors that mediate transcriptional changes. However, specific nucleosomal rearrangements were also evident at promoters even when there was no apparent transcriptional change, indicating that there is no simple, globally applicable relationship between chromatin remodeling and transcriptional activity. Our study provides a detailed, high-resolution, dynamic map of single-nucleosome remodeling across the yeast genome and its relation to global transcriptional changes.

Invasive Mutualists Erode Native Pollination Webs

Abstract: Plant–animal mutualisms are characterized by weak or asymmetric mutual dependences between interacting species, a feature that could increase community stability. If invasive species integrate into mutualistic webs, they may alter web structure, with consequences for species persistence. However, the effect of alien mutualists on the architecture of plant–pollinator webs remains largely unexplored. We analyzed the extent of mutual dependency between interacting species, as a measure of mutualism strength, and the connectivity of 10 paired plant–pollinator webs, eight from forests of the southern Andes and two from oceanic islands, with different incidences of alien species. Highly invaded webs exhibited weaker mutualism than less-invaded webs. This potential increase in network stability was the result of a disproportionate increase in the importance and participation of alien species in the most asymmetric interactions. The integration of alien mutualists did not alter overall network connectivity, but links were transferred from generalist native species to super-generalist alien species during invasion. Therefore, connectivity among native species declined in highly invaded webs. These modifications in the structure of pollination webs, due to dominance of alien mutualists, can leave many native species subject to novel ecological and evolutionary dynamics.

Gene-environment interaction in yeast gene expression

Abstract: The effects of genetic variants on phenotypic traits often depend on environmental and physiological conditions, but such gene-environment interactions are poorly understood. Recently developed approaches that treat transcript abundances of thousands of genes as quantitative traits offer the opportunity to broadly characterize the architecture of gene-environment interactions. We examined the genetic and molecular basis of variation in gene expression between two yeast strains (BY and RM) grown in two different conditions (glucose and ethanol as carbon sources). We observed that most transcripts vary by strain and condition, with 2,996, 3,448, and 2,037 transcripts showing significant strain, condition, and strain-condition interaction effects, respectively. We expression profiled over 100 segregants derived from a cross between BY and RM in both growth conditions, and identified 1,555 linkages for 1,382 transcripts that show significant gene-environment interaction. At the locus level, local linkages, which usually correspond to polymorphisms in cis-regulatory elements, tend to be more stable across conditions, such that they are more likely to show the same effect or the same direction of effect across conditions. Distant linkages, which usually correspond to polymorphisms influencing trans-acting factors, are more condition-dependent, and often show effects in different directions in the two conditions. We characterized a locus that influences expression of many growth-related transcripts, and showed that the majority of the variation is explained by polymorphism in the gene IRA2. The RM allele of IRA2 appears to inhibit Ras/PKA signaling more strongly than the BY allele, and has undergone a change in selective pressure. Our results provide a broad overview of the genetic architecture of gene-environment interactions, as well as a detailed molecular example, and lead to key insights into how the effects of different classes of regulatory variants are modulated by the environment. These observations will guide the design of studies aimed at understanding the genetic basis of complex traits.

Density-dependent cladogenesis in birds.

Abstract: A characteristic signature of adaptive radiation is a slowing of the rate of speciation toward the present. On the basis of molecular phylogenies, studies of single clades have frequently found evidence for a slowdown in diversification rate and have interpreted this as evidence for density dependent speciation. However, we demonstrated via simulation that large clades are expected to show stronger slowdowns than small clades, even if the probability of speciation and extinction remains constant through time. This is a consequence of exponential growth: clades, which, by chance, diversify at above the average rate early in their history, will tend to be large. They will also tend to regress back to the average diversification rate later on, and therefore show a slowdown. We conducted a meta-analysis of the distribution of speciation events through time, focusing on sequence-based phylogenies for 45 clades of birds. Thirteen of the 23 clades (57%) that include more than 20 species show significant slowdowns. The high frequency of slowdowns observed in large clades is even more extreme than expected under a purely stochastic constant-rate model, but is consistent with the adaptive radiation model. Taken together, our data strongly support a model of density-dependent speciation in birds, whereby speciation slows as ecological opportunities and geographical space place limits on clade growth.