Anna Maria D’Erchia

BALB/c and C57BL/6 Mice Differ in Polyreactive IgA Abundance, which Impacts the Generation of Antigen-Specific IgA and Microbiota Diversity

The interrelationship between IgAs and microbiota diversity is still unclear. Here we show that BALB/c mice had higher abundance and diversity of IgAs than C57BL/6 mice and that this correlated with increased microbiota diversity. We show that polyreactive IgAs mediated the entrance of non-invasive bacteria to Peyers patches, independently of CX3CR1(+) phagocytes. This allowed the induction of bacteria-specific IgA and the establishment of a positive feedback loop of IgA production. Cohousing of mice or fecal transplantation had little or no influence on IgA production and had only partial impact on microbiota composition. Germ-free BALB/c, but not C57BL/6, mice already had polyreactive IgAs that influenced microbiota diversity and selection after colonization. Together, these data suggest that genetic predisposition to produce polyreactive IgAs has a strong impact on the generation of antigen-specific IgAs and the selection and maintenance of microbiota diversity.

Identification and functional characterization of two new transcriptional variants of the human p63 gene

p63 belongs to a family of transcription factors, which, while demonstrating striking conservation of functional domains, regulate distinct biological functions. Its principal role is in the regulation of epithelial commitment, differentiation and maintenance programs, during embryogenesis and in adult tissues. The p63 gene has a complex transcriptional pattern, producing two subclasses of N-terminal isoforms (TA and ΔN) which are alternatively spliced at the C-terminus. Here, we report the identification of two new C-terminus p63 variants, we named p63 δ and ε, that increase from 6 to 10 the number of the p63 isoforms. Expression analysis of all p63 variants demonstrates a tissue/cell-type-specific nature of p63 alternative transcript expression, probably related to their different cellular functions. We demonstrate that the new p63 variants as ΔN isoforms are active as transcription factors as they have nuclear localization and can modulate the expression of p63 target genes. Moreover, we report that, like ΔNp63α, ΔNp63δ and ε sustain cellular proliferation and that their expression decreases during keratinocyte differentiation, suggesting their involvement in this process. Taken together, our results demonstrate the existence of novel p63 proteins whose expression should be considered in future studies on the roles of p63 in the regulation of cellular functions.

Profiling RNA editing in human tissues: towards the inosinome Atlas.

Adenine to Inosine RNA editing is a widespread co- and post-transcriptional mechanism mediated by ADAR enzymes acting on double stranded RNA. It has a plethora of biological effects, appears to be particularly pervasive in humans with respect to other mammals, and is implicated in a number of diverse human pathologies. Here we present the first human inosinome atlas comprising 3,041,422 A-to-I events identified in six tissues from three healthy individuals. Matched directional total-RNA-Seq and whole genome sequence datasets were generated and analysed within a dedicated computational framework, also capable of detecting hyper-edited reads. Inosinome profiles are tissue specific and edited gene sets consistently show enrichment of genes involved in neurological disorders and cancer. Overall frequency of editing also varies, but is strongly correlated with ADAR expression levels. The inosinome database is available at: http://srv00.ibbe.cnr.it/editing/.

ASPicDB: a database of annotated transcript and protein variants generated by alternative splicing

Alternative splicing is emerging as a major mechanism for the expansion of the transcriptome and proteome diversity, particularly in human and other vertebrates. However, the proportion of alternative transcripts and proteins actually endowed with functional activity is currently highly debated. We present here a new release of ASPicDB which now provides a unique annotation resource of human protein variants generated by alternative splicing. A total of 256 939 protein variants from 17 191 multi-exon genes have been extensively annotated through state of the art machine learning tools providing information of the protein type (globular and transmembrane), localization, presence of PFAM domains, signal peptides, GPI-anchor propeptides, transmembrane and coiled-coil segments. Furthermore, full-length variants can be now specifically selected based on the annotation of CAGE-tags and polyA signal and/or polyA sites, marking transcription initiation and termination sites, respectively. The retrieval can be carried out at gene, transcript, exon, protein or splice site level allowing the selection of data sets fulfilling one or more features settled by the user. The retrieval interface also enables the selection of protein variants showing specific differences in the annotated features. ASPicDB is available at http://www.caspur.it/ASPicDB/.

SpliceAid-F: a database of human splicing factors and their RNA-binding sites

A comprehensive knowledge of all the factors involved in splicing, both proteins and RNAs, and of their interaction network is crucial for reaching a better understanding of this process and its functions. A large part of relevant information is buried in the literature or collected in various different databases. By hand-curated screenings of literature and databases, we retrieved experimentally validated data on 71 human RNA-binding splicing regulatory proteins and organized them into a database called ‘SpliceAid-F’ (http://www.caspur.it/SpliceAidF/). For each splicing factor (SF), the database reports its functional domains, its protein and chemical interactors and its expression data. Furthermore, we collected experimentally validated RNA–SF interactions, including relevant information on the RNA-binding sites, such as the genes where these sites lie, their genomic coordinates, the splicing effects, the experimental procedures used, as well as the corresponding bibliographic references. We also collected information from experiments showing no RNA–SF binding, at least in the assayed conditions. In total, SpliceAid-F contains 4227 interactions, 2590 RNA-binding sites and 1141 ‘no-binding’ sites, including information on cellular contexts and conditions where binding was tested. The data collected in SpliceAid-F can provide significant information to explain an observed splicing pattern as well as the effect of mutations in functional regulatory elements.

Intestinal microbiota sustains inflammation and autoimmunity induced by hypomorphic RAG defects

Rigoni et al. report that hypomorphic Rag2R229Q mutation is associated with altered microbiota composition and defects in the gut–blood barrier and suggest that intestinal microbes may play a critical role in the distinctive immune dysregulation of Omenn syndrome.

Uncovering RNA Editing Sites in Long Non-Coding RNAs.

RNA editing is an important co/post-transcriptional molecular process able to modify RNAs by nucleotide insertions/deletions or substitutions. In human, the most common RNA editing event involves the deamination of adenosine (A) into inosine (I) through the adenosine deaminase acting on RNA proteins. Although A-to-I editing can occur in both coding and non-coding RNAs, recent findings, based on RNA-seq experiments, have clearly demonstrated that a large fraction of RNA editing events alter non-coding RNAs sequences including untranslated regions of mRNAs, introns, long non-coding RNAs (lncRNAs), and low molecular weight RNAs (tRNA, miRNAs, and others). An accurate detection of A-to-I events occurring in non-coding RNAs is of utmost importance to clarify yet unknown functional roles of RNA editing in the context of gene expression regulation and maintenance of cell homeostasis. In the last few years, massive transcriptome sequencing has been employed to identify putative RNA editing changes at genome scale. Despite several efforts, the computational prediction of A-to-I sites in complete eukaryotic genomes is yet a challenging task. We have recently developed a software package, called REDItools, in order to simplify the detection of RNA editing events from deep sequencing data. In the present work, we show the potential of our tools in recovering A-to-I candidates from RNA-Seq experiments as well as guidelines to improve the RNA editing detection in non-coding RNAs, with specific attention to the lncRNAs.

Comparative Genomics of Listeria Sensu Lato: Genus-Wide Differences in Evolutionary Dynamics and the Progressive Gain of Complex, Potentially Pathogenicity-Related Traits through Lateral Gene Transfer

Historically, genome-wide and molecular characterization of the genus Listeria has concentrated on the important human pathogen Listeria monocytogenes and a small number of closely related species, together termed Listeria sensu strictu. More recently, a number of genome sequences for more basal, and nonpathogenic, members of the Listeria genus have become available, facilitating a wider perspective on the evolution of pathogenicity and genome level evolutionary dynamics within the entire genus (termed Listeria sensu lato). Here, we have sequenced the genomes of additional Listeria fleischmannii and Listeria newyorkensis isolates and explored the dynamics of genome evolution in Listeria sensu lato. Our analyses suggest that acquisition of genetic material through gene duplication and divergence as well as through lateral gene transfer (mostly from outside Listeria) is widespread throughout the genus. Novel genetic material is apparently subject to rapid turnover. Multiple lines of evidence point to significant differences in evolutionary dynamics between the most basal Listeria subclade and all other congeners, including both sensu strictu and other sensu lato isolates. Strikingly, these differences are likely attributable to stochastic, population-level processes and contribute to observed variation in genome size across the genus. Notably, our analyses indicate that the common ancestor of Listeria sensu lato lacked flagella, which were acquired by lateral gene transfer by a common ancestor of Listeria grayi and Listeria sensu strictu, whereas a recently functionally characterized pathogenicity island, responsible for the capacity to produce cobalamin and utilize ethanolamine/propane-2-diol, was acquired in an ancestor of Listeria sensu strictu.

T Follicular Helper Cells Promote a Beneficial Gut Ecosystem for Host Metabolic Homeostasis by sensing Microbiota-Derived Extracellular ATP.

Summary The ATP-gated ionotropic P2X7 receptor regulates T follicular helper (Tfh) cell abundance in the Peyer’s patches (PPs) of the small intestine; deletion of P2rx7, encoding for P2X7, in Tfh cells results in enhanced IgA secretion and binding to commensal bacteria. Here, we show that Tfh cell activity is important for generating a diverse bacterial community in the gut and that sensing of microbiota-derived extracellular ATP via P2X7 promotes the generation of a proficient gut ecosystem for metabolic homeostasis. The results of this study indicate that Tfh cells play a role in host-microbiota mutualism beyond protecting the intestinal mucosa by induction of affinity-matured IgA and suggest that extracellular ATP constitutes an inter-kingdom signaling molecule important for selecting a beneficial microbial community for the host via P2X7-mediated regulation of B cell help.

Methods for screening tumors for p53 status and therapeutic exploitation.

Mutations in the p53 oncosuppressor gene occur in most human cancers and regulation of the protein is defective in a variety of others. Novel strategies are emerging for the treatment of tumors that have p53 mutations. In this context, the analysis of p53 status is useful in diagnosis and prognosis, and could serve to evaluate the effectiveness of a cancer treatment. In this review, we report an overview of major methods for screening tumors for p53 status and the major strategies suggested for restoring p53 function.