Search

Single-cell technologies have transformed our understanding of human tissues. Yet, studies typically capture only a limited number of donors and disagree on cell type definitions. Integrating many single-cell datasets can address these limitations of individual studies and capture the variability present in the population. Here we present the integrated Human Lung Cell Atlas (HLCA), combining 49 datasets of the human respiratory system into a single atlas spanning over 2.4 million cells from 486 individuals. The HLCA presents a consensus cell type re-annotation with matching marker genes, including annotations of rare and previously undescribed cell types. Leveraging the number and diversity of individuals in the HLCA, we identify gene modules that are associated with demographic covariates such as age, sex and body mass index, as well as gene modules changing expression along the proximal-to-distal axis of the bronchial tree. Mapping new data to the HLCA enables rapid data annotation and interpretation. Using the HLCA as a reference for the study of disease, we identify shared cell states across multiple lung diseases, including SPP1 + profibrotic monocyte-derived macrophages in COVID-19, pulmonary fibrosis and lung carcinoma. Overall, the HLCA serves as an example for the development and use of large-scale, cross-dataset organ atlases within the Human Cell Atlas., (© 2023. The Author(s).)

The genus Yersinia includes a large variety of nonpathogenic and life-threatening pathogenic bacteria, which cause a broad spectrum of diseases in humans and animals, such as plague, enteritis, Far East scarlet-like fever (FESLF), and enteric redmouth disease. Like most clinically relevant microorganisms, Yersinia spp. are currently subjected to intense multi-omics investigations whose numbers have increased extensively in recent years, generating massive amounts of data useful for diagnostic and therapeutic developments. The lack of a simple and centralized way to exploit these data led us to design Yersiniomics, a web-based platform allowing straightforward analysis of Yersinia omics data. Yersiniomics contains a curated multi-omics database at its core, gathering 200 genomic, 317 transcriptomic, and 62 proteomic data sets for Yersinia species. It integrates genomic, transcriptomic, and proteomic browsers, a genome viewer, and a heatmap viewer to navigate within genomes and experimental conditions. For streamlined access to structural and functional properties, it directly links each gene to GenBank, the Kyoto Encyclopedia of Genes and Genomes (KEGG), UniProt, InterPro, IntAct, and the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) and each experiment to Gene Expression Omnibus (GEO), the European Nucleotide Archive (ENA), or the Proteomics Identifications Database (PRIDE). Yersiniomics provides a powerful tool for microbiologists to assist with investigations ranging from specific gene studies to systems biology studies. IMPORTANCE The expanding genus Yersinia is composed of multiple nonpathogenic species and a few pathogenic species, including the deadly etiologic agent of plague, Yersinia pestis. In 2 decades, the number of genomic, transcriptomic, and proteomic studies on Yersinia grew massively, delivering a wealth of data. We developed Yersiniomics, an interactive web-based platform, to centralize and analyze omics data sets on Yersinia species. The platform allows user-friendly navigation between genomic data, expression data, and experimental conditions. Yersiniomics will be a valuable tool to microbiologists.

Listeria monocytogenes is a foodborne intracellular bacterial pathogen leading to human listeriosis. Despite a high mortality rate and increasing antibiotic resistance no clinically approved vaccine against Listeria is available. Attenuated Listeria strains offer protection and are tested as antitumor vaccine vectors, but would benefit from a better knowledge on immunodominant vector antigens. To identify novel antigens, we screen for Listeria peptides presented on the surface of infected human cell lines by mass spectrometry-based immunopeptidomics. In between more than 15,000 human self-peptides, we detect 68 Listeria immunopeptides from 42 different bacterial proteins, including several known antigens. Peptides presented on different cell lines are often derived from the same bacterial surface proteins, classifying these antigens as potential vaccine candidates. Encoding these highly presented antigens in lipid nanoparticle mRNA vaccine formulations results in specific CD8 +  T-cell responses and induces protection in vaccination challenge experiments in mice. Our results can serve as a starting point for the development of a clinical mRNA vaccine against Listeria and aid to improve attenuated Listeria vaccines and vectors, demonstrating the power of immunopeptidomics for next-generation bacterial vaccine development., (© 2022. The Author(s).)

Summary: To face up to the exponential growth of heterogeneous datasets of various organisms, we developed a user-friendly platform for building multi-omics websites, which is named Bacnet. This platform helps bioinformaticians to construct four key web interfaces: (i) an interactive genome viewer; (ii) an expression and protein atlas; (iii) an interface for analysis of co-expression network; (iv) an interface for exploring homolog presence. We believe our platform will help the bioinformaticians to construct personalized user interfaces dedicated to biologists studying non-reference organisms., Availability and Implementation: https://github.com/becavin-lab/bacnet; Java; Eclipse RAP; Eclipse RCP., (© The Author(s) 2020. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Angiotensin-converting enzyme 2 (ACE2) and accessory proteases (TMPRSS2 and CTSL) are needed for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) cellular entry, and their expression may shed light on viral tropism and impact across the body. We assessed the cell-type-specific expression of ACE2, TMPRSS2 and CTSL across 107 single-cell RNA-sequencing studies from different tissues. ACE2, TMPRSS2 and CTSL are coexpressed in specific subsets of respiratory epithelial cells in the nasal passages, airways and alveoli, and in cells from other organs associated with coronavirus disease 2019 (COVID-19) transmission or pathology. We performed a meta-analysis of 31 lung single-cell RNA-sequencing studies with 1,320,896 cells from 377 nasal, airway and lung parenchyma samples from 228 individuals. This revealed cell-type-specific associations of age, sex and smoking with expression levels of ACE2, TMPRSS2 and CTSL. Expression of entry factors increased with age and in males, including in airway secretory cells and alveolar type 2 cells. Expression programs shared by ACE2 + TMPRSS2 + cells in nasal, lung and gut tissues included genes that may mediate viral entry, key immune functions and epithelial-macrophage cross-talk, such as genes involved in the interleukin-6, interleukin-1, tumor necrosis factor and complement pathways. Cell-type-specific expression patterns may contribute to the pathogenesis of COVID-19, and our work highlights putative molecular pathways for therapeutic intervention.

We investigated SARS-CoV-2 potential tropism by surveying expression of viral entry-associated genes in single-cell RNA-sequencing data from multiple tissues from healthy human donors. We co-detected these transcripts in specific respiratory, corneal and intestinal epithelial cells, potentially explaining the high efficiency of SARS-CoV-2 transmission. These genes are co-expressed in nasal epithelial cells with genes involved in innate immunity, highlighting the cells' potential role in initial viral infection, spread and clearance. The study offers a useful resource for further lines of inquiry with valuable clinical samples from COVID-19 patients and we provide our data in a comprehensive, open and user-friendly fashion at www.covid19cellatlas.org.

The intestinal microbiota modulates host physiology and gene expression via mechanisms that are not fully understood. Here we examine whether host epitranscriptomic marks are affected by the gut microbiota. We use methylated RNA-immunoprecipitation and sequencing (MeRIP-seq) to identify N6-methyladenosine (m 6 A) modifications in mRNA of mice carrying conventional, modified, or no microbiota. We find that variations in the gut microbiota correlate with m 6 A modifications in the cecum, and to a lesser extent in the liver, affecting pathways related to metabolism, inflammation and antimicrobial responses. We analyze expression levels of several known writer and eraser enzymes, and find that the methyltransferase Mettl16 is downregulated in absence of a microbiota, and one of its target mRNAs, encoding S-adenosylmethionine synthase Mat2a, is less methylated. We furthermore show that Akkermansia muciniphila and Lactobacillus plantarum affect specific m 6 A modifications in mono-associated mice. Our results highlight epitranscriptomic modifications as an additional level of interaction between commensal bacteria and their host.

RNA-binding proteins (RBPs) perform key cellular activities by controlling the function of bound RNAs. The widely held assumption that RBPs are strictly intracellular has been challenged by the discovery of secreted RBPs. However, extracellular RBPs have been described in eukaryotes, while secreted bacterial RBPs have not been reported. Here, we show that the bacterial pathogen Listeria monocytogenes secretes a small RBP that we named Zea. We show that Zea binds a subset of L. monocytogenes RNAs, causing their accumulation in the extracellular medium. Furthermore, during L. monocytogenes infection, Zea binds RIG-I, the non-self-RNA innate immunity sensor, potentiating interferon-β production. Mouse infection studies reveal that Zea affects L. monocytogenes virulence. Together, our results unveil that bacterial RNAs can be present extracellularly in association with RBPs, acting as "social RNAs" to trigger a host response during infection., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Small regulatory RNAs (sRNAs) are widely found in bacteria and play key roles in many important physiological and adaptation processes. Studying their evolution and screening for events of coevolution with other genomic features is a powerful way to better understand their origin and assess a common functional or adaptive relationship between them. However, evolution and coevolution of sRNAs with coding genes have been sparsely investigated in bacterial pathogens., Results: We designed a robust and generic phylogenomics approach that detects correlated evolution between sRNAs and protein-coding genes using their observed and inferred patterns of presence-absence in a set of annotated genomes. We applied this approach on 79 complete genomes of the Listeria genus and identified fifty-two accessory sRNAs, of which most were present in the Listeria common ancestor and lost during Listeria evolution. We detected significant coevolution between 23 sRNA and 52 coding genes and inferred the Listeria sRNA-coding genes coevolution network. We characterized a main hub of 12 sRNAs that coevolved with genes encoding cell wall proteins and virulence factors. Among them, an sRNA specific to L. monocytogenes species, rli133, coevolved with genes involved either in pathogenicity or in interaction with host cells, possibly acting as a direct negative post-transcriptional regulation., Conclusions: Our approach allowed the identification of candidate sRNAs potentially involved in pathogenicity and host interaction, consistent with recent findings on known pathogenicity actors. We highlight four sRNAs coevolving with seven internalin genes, some of which being important virulence factors in Listeria.

As for many model organisms, the amount of Listeria omics data produced has recently increased exponentially. There are now >80 published complete Listeria genomes, around 350 different transcriptomic data sets, and 25 proteomic data sets available. The analysis of these data sets through a systems biology approach and the generation of tools for biologists to browse these various data are a challenge for bioinformaticians. We have developed a web-based platform, named Listeriomics, that integrates different tools for omics data analyses, i.e., (i) an interactive genome viewer to display gene expression arrays, tiling arrays, and sequencing data sets along with proteomics and genomics data sets; (ii) an expression and protein atlas that connects every gene, small RNA, antisense RNA, or protein with the most relevant omics data; (iii) a specific tool for exploring protein conservation through the Listeria phylogenomic tree; and (iv) a coexpression network tool for the discovery of potential new regulations. Our platform integrates all the complete Listeria species genomes, transcriptomes, and proteomes published to date. This website allows navigation among all these data sets with enriched metadata in a user-friendly format and can be used as a central database for systems biology analysis. IMPORTANCE In the last decades, Listeria has become a key model organism for the study of host-pathogen interactions, noncoding RNA regulation, and bacterial adaptation to stress. To study these mechanisms, several genomics, transcriptomics, and proteomics data sets have been produced. We have developed Listeriomics, an interactive web platform to browse and correlate these heterogeneous sources of information. Our website will allow listeriologists and microbiologists to decipher key regulation mechanism by using a systems biology approach.

To adapt to changing environments, bacteria have evolved numerous pathways that activate stress response genes. In Gram-positive bacteria, the stressosome, a cytoplasmic complex, relays external cues and activates the sigma B regulon. The stressosome is structurally well-characterized in Bacillus, but how it senses stress remains elusive. Here, we report a genome-wide N-terminomic approach in Listeria that strikingly led to the discovery of 19 internal translation initiation sites and 6 miniproteins, among which one, Prli42, is conserved in Firmicutes. Prli42 is membrane-anchored and interacts with orthologues of Bacillus stressosome components. We reconstituted the Listeria stressosome in vitro and visualized its supramolecular structure by electron microscopy. Analysis of a series of Prli42 mutants demonstrated that Prli42 is important for sigma B activation, bacterial growth following oxidative stress and for survival in macrophages. Taken together, our N-terminonic approach unveiled Prli42 as a long-sought link between stress and the stressosome.

Background: Plasmodium falciparum malaria in India is characterized by high rates of severe disease, with multiple organ dysfunction (MOD)-mainly associated with acute renal failure (ARF)-and increased mortality. The objective of this study is to identify cytokine signatures differentiating severe malaria patients with MOD, cerebral malaria (CM), and cerebral malaria with MOD (CM-MOD) in India. We have previously shown that two cytokines clusters differentiated CM from mild malaria in Maharashtra. Hence, we also aimed to determine if these cytokines could discriminate malaria subphenotypes in Odisha., Methods: P. falciparum malaria patients from the SCB Medical College Cuttack in the Odisha state in India were enrolled along with three sets of controls: healthy individuals, patients with sepsis and encephalitis (n = 222). We determined plasma concentrations of pro- and anti-inflammatory cytokines and chemokines for all individuals using a multiplex assay. We then used an ensemble of statistical analytical methods to ascertain whether particular sets of cytokines/chemokines were predictors of severity or signatures of a disease category., Results: Of the 26 cytokines/chemokines tested, 19 increased significantly during malaria and clearly distinguished malaria patients from controls, as well as sepsis and encephalitis patients. High amounts of IL-17, IP-10, and IL-10 predicted MOD, decreased IL-17 and MIP-1α segregated CM-MOD from MOD, and increased IL-12p40 differentiated CM from CM-MOD. Most severe malaria patients with ARF exhibited high levels of IL-17., Conclusion: We report distinct differences in cytokine production correlating with malarial disease severity in Odisha and Maharashtra populations in India. We show that CM, CM-MOD and MOD are clearly distinct malaria-associated pathologies. High amounts of IL-17, IP-10, and IL-10 were predictors of MOD; decreased IL-17 and MIP-1α separated CM-MOD from MOD; and increased IL-12p40 differentiated CM from CM-MOD. Data also suggest that the IL-17 pathway may contribute to malaria pathogenesis via different regulatory mechanisms and may represent an interesting target to mitigate the pathological processes in malaria-associated ARF.

Background: Mechanisms of acquired protection to malaria in asymptomatic Plasmodium falciparum carriers are only partially understood. Among them, the role plays by the self-reactive antibodies has not been clarified yet. In this study, the relationship between repertoires of circulating self-reactive and parasite-specific immunoglobulin G (IgG), their correlation with cytokine levels, and their association with protection against malaria was investigated in asymptomatic Plasmodium falciparum-infected Gabonese children., Methods: The diversity of P. falciparum-specific antibody repertoire was analysed using a protein micro-array immunoassay, the total auto-antibody repertoire by quantitative immunoblotting and circulating cytokine levels were measured by ELISA in endemic controls (EC) and P. falciparum-infected children from Gabon with asymptomatic (AM) or mild malaria (MM). The association of self- and parasite-specific antibody repertoires with circulating cytokines was evaluated using single linkage hierarchical clustering, Kruskal-Wallis tests and Spearman's rank correlation., Results: Children with AM exhibited an IgG response to merozoite surface protein 3 (MSP3) but not to MSP1-19, although their levels of total P. falciparum-specific IgG were similar to those in the MM group. Moreover, the asymptomatic children had increased levels of autoantibodies recognising brain antigens. In addition, a correlation between IL-10 levels and parasite load was found in AM and MM children. These two groups also exhibited significant correlations between plasma levels of IL-10 and IFN-γ with age and with total plasma IgG levels. IL-10 and IFN-γ levels were also associated with auto-antibody responses in AM., Conclusions: Altogether, these results indicate that a self-reactive polyclonal response associated with increased IgG to MSP3 and high plasma levels of IL-10 and IFN-γ may contribute to protective immune mechanisms triggered in asymptomatic P. falciparum infection in Gabonese children.

For nearly 3 decades, listeriologists and immunologists have used mainly three strains of the same serovar (1/2a) to analyze the virulence of the bacterial pathogen Listeria monocytogenes. The genomes of two of these strains, EGD-e and 10403S, were released in 2001 and 2008, respectively. Here we report the genome sequence of the third reference strain, EGD, and extensive genomic and phenotypic comparisons of the three strains. Strikingly, EGD-e is genetically highly distinct from EGD (29,016 single nucleotide polymorphisms [SNPs]) and 10403S (30,296 SNPs), and is more related to serovar 1/2c than 1/2a strains. We also found that while EGD and 10403S strains are genetically very close (317 SNPs), EGD has a point mutation in the transcriptional regulator PrfA (PrfA*), leading to constitutive expression of several major virulence genes. We generated an EGD-e PrfA* mutant and showed that EGD behaves like this strain in vitro, with slower growth in broth and higher invasiveness in human cells than those of EGD-e and 10403S. In contrast, bacterial counts in blood, liver, and spleen during infection in mice revealed that EGD and 10403S are less virulent than EGD-e, which is itself less virulent than EGD-e PrfA*. Thus, constitutive expression of PrfA-regulated virulence genes does not appear to provide a significant advantage to the EGD strain during infection in vivo, highlighting the fact that in vitro invasion assays are not sufficient for evaluating the pathogenic potential of L. monocytogenes strains. Together, our results pave the way for deciphering unexplained differences or discrepancies in experiments using different L. monocytogenes strains. IMPORTANCE Over the past 3 decades, Listeria has become a model organism for host-pathogen interactions, leading to critical discoveries in a broad range of fields, including bacterial gene regulation, cell biology, and bacterial pathophysiology. Scientists studying Listeria use primarily three pathogenic strains: EGD, EGD-e, and 10403S. Despite many studies on EGD, it is the only one of the three strains whose genome has not been sequenced. Here we report the sequence of its genome and a series of important genomic and phenotypic differences between the three strains, in particular, a critical mutation in EGD's PrfA, the main regulator of Listeria virulence. Our results show that the three strains display differences which may play an important role in the virulence differences observed between the strains. Our findings will be of critical relevance to listeriologists and immunologists who have used or may use Listeria as a tool to study the pathophysiology of listeriosis and immune responses.

Unlabelled: The intestinal tract is the largest reservoir of microbes in the human body. The intestinal microbiota is thought to be able to modulate alterations of the gut induced by enteropathogens, thereby maintaining homeostasis. Listeria monocytogenes is the agent of listeriosis, an infection transmitted to humans upon ingestion of contaminated food. Crossing of the intestinal barrier is a critical step of the infection before dissemination into deeper organs. Here, we investigated the role of the intestinal microbiota in the regulation of host protein-coding genes and microRNA (miRNA or miR) expression during Listeria infection. We first established the intestinal miRNA signatures corresponding to the 10 most highly expressed miRNAs in the murine ileum of conventional and germfree mice, noninfected and infected with Listeria. Next, we identified 6 miRNAs whose expression decreased upon Listeria infection in conventional mice. Strikingly, five of these miRNA expression variations (in miR-143, miR-148a, miR-200b, miR-200c, and miR-378) were dependent on the presence of the microbiota. In addition, as is already known, protein-coding genes were highly affected by infection in both conventional and germfree mice. By crossing bioinformatically the predicted targets of the miRNAs to our whole-genome transcriptomic data, we revealed an miRNA-mRNA network that suggested miRNA-mediated global regulation during intestinal infection. Other recent studies have revealed an miRNA response to either bacterial pathogens or commensal bacteria. In contrast, our work provides an unprecedented insight into the impact of the intestinal microbiota on host transcriptional reprogramming during infection by a human pathogen., Importance: While the crucial role of miRNAs in regulating the host response to bacterial infection is increasingly recognized, the involvement of the intestinal microbiota in the regulation of miRNA expression has not been explored in detail. Here, we investigated the impact of the intestinal microbiota on the regulation of protein-coding genes and miRNA expression in a host infected by L. monocytogenes, a food-borne pathogen. We show that the microbiota interferes with the microRNA response upon oral Listeria infection and identify several protein-coding target genes whose expression correlates inversely with that of the miRNA. Further investigations of the regulatory networks involving miR-143, miR-148a, miR-200b, miR-200c, and miR-378 will provide new insights into the impact of the intestinal microbiota on the host upon bacterial infection.

Riboswitches are ligand-binding elements located in 5' untranslated regions of messenger RNAs, which regulate expression of downstream genes. In Listeria monocytogenes, a vitamin B12-binding (B12) riboswitch was identified, not upstream of a gene but downstream, and antisense to the adjacent gene, pocR, suggesting it might regulate pocR in a nonclassical manner. In Salmonella enterica, PocR is a transcription factor that is activated by 1,2-propanediol, and subsequently activates expression of the pdu genes. The pdu genes mediate propanediol catabolism and are implicated in pathogenesis. As enzymes involved in propanediol catabolism require B12 as a cofactor, we hypothesized that the Listeria B12 riboswitch might be involved in pocR regulation. Here we demonstrate that the B12 riboswitch is transcribed as part of a noncoding antisense RNA, herein named AspocR. In the presence of B12, the riboswitch induces transcriptional termination, causing aspocR to be transcribed as a short transcript. In contrast, in the absence of B12, aspocR is transcribed as a long antisense RNA, which inhibits pocR expression. Regulation by AspocR ensures that pocR, and consequently the pdu genes, are maximally expressed only when both propanediol and B12 are present. Strikingly, AspocR can inhibit pocR expression in trans, suggesting it acts through a direct interaction with pocR mRNA. Together, this study demonstrates how pocR and the pdu genes can be regulated by B12 in bacteria and extends the classical definition of riboswitches from elements governing solely the expression of mRNAs to a wider role in controlling transcription of noncoding RNAs.

Background: Influenza infection causes respiratory disease that can lead to death. The complex interplay between virus-encoded and host-specific pathogenicity regulators - and the relative contributions of each toward viral pathogenicity - is not well-understood., Results: By analyzing a collection of lung samples from mice infected by A/Vietnam/1203/2004 (H5N1; VN1203), we characterized a signature of transcripts and proteins associated with the kinetics of the host response. Using a new geometrical representation method and two criteria, we show that inoculation concentrations and four specific mutations in VN1203 mainly impact the magnitude and velocity of the host response kinetics, rather than specific sets of up- and down- regulated genes. We observed analogous kinetic effects using lung samples from mice infected with A/California/04/2009 (H1N1), and we show that these effects correlate with morbidity and viral titer., Conclusions: We have demonstrated the importance of the kinetics of the host response to H5N1 pathogenesis and its relationship with clinical disease severity and virus replication. These kinetic properties imply that time-matched comparisons of 'omics profiles to viral infections give limited views to differentiate host-responses. Moreover, these results demonstrate that a fast activation of the host-response at the earliest time points post-infection is critical for protective mechanisms against fast replicating viruses.

Listeria monocytogenes is a foodborne pathogen that crosses the intestinal barrier and disseminates within the host. Here, we report a unique comprehensive analysis of the impact of two Lactobacillus species, Lactobacillus paracasei CNCM I-3689 and Lactobacillus casei BL23, on L. monocytogenes and orally acquired listeriosis in a gnotobiotic humanized mouse model. We first assessed the effect of treatment with each Lactobacillus on L. monocytogenes counts in host tissues and showed that each decreases L. monocytogenes systemic dissemination in orally inoculated mice. A whole genome intestinal transcriptomic analysis revealed that each Lactobacillus changes expression of a specific subset of genes during infection, with IFN-stimulated genes (ISGs) being the most affected by both lactobacilli. We also examined microRNA (miR) expression and showed that three miRs (miR-192, miR-200b, and miR-215) are repressed during L. monocytogenes infection. Treatment with each Lactobacillus increased miR-192 expression, whereas only L. casei association increased miR-200b and miR-215 expression. Finally, we showed that treatment with each Lactobacillus significantly reshaped the L. monocytogenes transcriptome and up-regulated transcription of L. monocytogenes genes encoding enzymes allowing utilization of intestinal carbon and nitrogen sources in particular genes involved in propanediol and ethanolamine catabolism and cobalamin biosynthesis. Altogether, these data reveal that the modulation of L. monocytogenes infection by treatment with lactobacilli correlates with a decrease in host gene expression, in particular ISGs, miR regulation, and a dramatic reshaping of L. monocytogenes transcriptome.

Listeria monocytogenes is a human, food-borne pathogen. Genomic comparisons between L. monocytogenes and Listeria innocua, a closely related non-pathogenic species, were pivotal in the identification of protein-coding genes essential for virulence. However, no comprehensive comparison has focused on the non-coding genome. We used strand-specific cDNA sequencing to produce genome-wide transcription start site maps for both organisms, and developed a publicly available integrative browser to visualize and analyze both transcriptomes in different growth conditions and genetic backgrounds. Our data revealed conservation across most transcripts, but significant divergence between the species in a subset of non-coding RNAs. In L. monocytogenes, we identified 113 small RNAs (33 novel) and 70 antisense RNAs (53 novel), significantly increasing the repertoire of ncRNAs in this species. Remarkably, we identified a class of long antisense transcripts (lasRNAs) that overlap one gene while also serving as the 5' UTR of the adjacent divergent gene. Experimental evidence suggests that lasRNAs transcription inhibits expression of one operon while activating the expression of another. Such a lasRNA/operon structure, that we named 'excludon', might represent a novel form of regulation in bacteria.

Allostery is a key concept of molecular biology which refers to the control of an enzyme activity by an effector molecule binding the enzyme at another site rather than the active site (allos = other in Greek). We revisit here allostery in the context of chromatin and argue that allosteric principles underlie and explain the functional architecture required for spacetime coordination of gene expression at all scales from DNA to the whole chromosome. We further suggest that this functional architecture is provided by the chromatin fiber itself. The structural, mechanical and topological features of the chromatin fiber endow chromosomes with a tunable signal transduction from specific (or nonspecific) effectors to specific (or nonspecific) active sites. Mechanical constraints can travel along the fiber all the better since the fiber is more compact and regular, which speaks in favor of the actual existence of the (so-called 30 nm) chromatin fiber. Chromatin fiber allostery reconciles both the physical and biochemical approaches of chromatin. We illustrate this view with two supporting specific examples. Moreover, from a methodological point of view, we suggest that the notion of chromatin fiber allostery is particularly relevant for systemic approaches. Finally we discuss the evolutionary power of allostery in the context of chromatin and its relation to modularity.

Motivation: Multidimensional scaling (MDS) is a well-known multivariate statistical analysis method used for dimensionality reduction and visualization of similarities and dissimilarities in multidimensional data. The advantage of MDS with respect to singular value decomposition (SVD) based methods such as principal component analysis is its superior fidelity in representing the distance between different instances specially for high-dimensional geometric objects. Here, we investigate the importance of the choice of initial conditions for MDS, and show that SVD is the best choice to initiate MDS. Furthermore, we demonstrate that the use of the first principal components of SVD to initiate the MDS algorithm is more efficient than an iteration through all the principal components. Adding stochasticity to the molecular dynamics simulations typically used for MDS of large datasets, contrary to previous suggestions, likewise does not increase accuracy. Finally, we introduce a k nearest neighbor method to analyze the local structure of the geometric objects and use it to control the quality of the dimensionality reduction., Results: We demonstrate here the, to our knowledge, most efficient and accurate initialization strategy for MDS algorithms, reducing considerably computational load. SVD-based initialization renders MDS methodology much more useful in the analysis of high-dimensional data such as functional genomics datasets.

The small nuclear 7SK RNA negatively controls transcription by inactivating positive transcription elongation factor b (P-TEFb) and is an integral component of Tat-dependent and independent HIV-1 transcription initiation complexes. 7SK RNA has recently been shown to also directly control HMGA1 transcription activity. HMGA1 is a master regulator of gene expression and its deregulation is associated with virtually any type of human cancer. The degree of HMGA1 over-expression thereby correlates with tumor malignancy and metastatic potential. 7SK snRNA directly interacts through its loop2 (7SK L2) with the first A/T-hook DNA binding motif of HMGA1. We have developed several 7SK L2 RNA chimera with the Epstein Barr Virus expressed RNA 2 (EBER2) to target HMGA1 function in transcription regulation. The efficiency of interfering with HMGA1 transcription activity by the chimeric 7SK L2-EBER2 fusions by large exceeds the efficiency of 7SK wild-type RNA due to the stronger EBER2 promoter activity. Furthermore, the 7SK L2-EBER2 chimera do not interfere with P-TEFb controlled transcription elongation or the formation of 7SK sn/hnRNPs. The comparison of the effects of wild-type 7SK RNA on cellular transcriptome dynamics with those induced by the two 7SK L2 mutants as well as the changes in gene expression following inhibition of HMGA1 allow the identification and characterization of HMGA1-dependent and independent effects of 7SK snRNA. We furthermore also present evidence for P-TEFb and HMGA1-independent 7SK RNA L2 regulatory activity.

'Omics' data have increased very rapidly in quantity and resolution, and are increasingly recognized as very valuable experimental observations in the systematic study of biological phenomena. The increase in availability, complexity and nonexpert interest in such data requires the urgent development of accurate and efficient dimensionality reduction and visualization techniques. To illustrate this need for new approaches we extensively discuss current methodology in terms of the limitations encountered. We then illustrate a recent example of how combinations of existing techniques can be used to overcome some of the present limitations, and discuss possible future directions for research in this important field of study.

During eukaryotic transcription, RNA-polymerase activity generates torsional stress in DNA, having a negative impact on the elongation process. Using our previous studies of chromatin fiber structure and conformational transitions, we suggest that this torsional stress can be alleviated, thanks to a tradeoff between the fiber twist and nucleosome conformational transitions into an activated state named "reversome". Our model enlightens the origin of polymerase pauses, and leads to the counterintuitive conclusion that chromatin-organized compaction might facilitate polymerase progression. Indeed, in a compact and well-structured chromatin loop, steric hindrance between nucleosomes enforces sequential transitions, thus ensuring that the polymerase always meets a permissive nucleosomal state., (2010 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

Background: The main processes in the pathogenesis of cerebral malaria caused by Plasmodium falciparum involved sequestration of parasitized red blood cells and immunopathological responses. Among immune factors, IgG autoantibodies to brain antigens are increased in P. falciparum infected patients and correlate with disease severity in African children. Nevertheless, their role in the pathophysiology of cerebral malaria (CM) is not fully defined. We extended our analysis to an Indian population with genetic backgrounds and endemic and environmental status different from Africa to determine if these autoantibodies could be either a biomarker or a risk factor of developing CM., Methods/principal Findings: We investigated the significance of these self-reactive antibodies in clinically well-defined groups of P. falciparum infected patients manifesting mild malaria (MM), severe non-cerebral malaria (SM), or cerebral malaria (CM) and in control subjects from Gondia, a malaria epidemic site in central India using quantitative immunoprinting and multivariate statistical analyses. A two-fold complete-linkage hierarchical clustering allows classifying the different patient groups and to distinguish the CM from the others on the basis of their profile of IgG reactivity to brain proteins defined by PANAMA Blot. We identified beta tubulin III (TBB3) as a novel discriminant brain antigen in the prevalence of CM. In addition, circulating IgG from CM patients highly react with recombinant TBB3. Overall, correspondence analyses based on singular value decomposition show a strong correlation between IgG anti-TBB3 and elevated concentration of cluster-II cytokine (IFNgamma, IL1beta, TNFalpha, TGFbeta) previously demonstrated to be a predictor of CM in the same population., Conclusions/significance: Collectively, these findings validate the relationship between antibody response to brain induced by P. falciparum infection and plasma cytokine patterns with clinical outcome of malaria. They also provide significant insight into the immune mechanisms associated to CM by the identification of TBB3 as a new disease-specific marker and potential therapeutic target.