Identification of candidate regulatory sequences in mammalian 3' UTRs by statistical analysis of oligonucleotide distributions

3' untranslated regions (3' UTRs) contain binding sites for many regulatory elements, and in particular for microRNAs (miRNAs). The importance of miRNA-mediated post-transcriptional regulation has become increasingly clear in the last few years. We propose two complementary approaches to the statistical analysis of oligonucleotide frequencies in mammalian 3' UTRs aimed at the identification of candidate binding sites for regulatory elements. The first method is based on the identification of sets of genes characterized by evolutionarily conserved overrepresentation of an oligonucleotide. The second method is based on the identification of oligonucleotides showing statistically significant strand asymmetry in their distribution in 3' UTRs. Both methods are able to identify many previously known binding sites located in 3'UTRs, and in particular seed regions of known miRNAs. Many new candidates are proposed for experimental verification.Comment: Added two reference

Mammalian Cis-Acting RNA Sequence Elements

Cis-acting regulatory sequence elements are sequences contained in the 3′ and 5′ untranslated region, introns, or coding regions of precursor RNAs and mature mRNAs that are selectively recognized by a complementary set of one or more trans-acting factors to regulate posttranscriptional gene expression. This chapter focuses on mammalian cis-acting regulatory elements that had been recently discovered in different regions: pre-processed and mature. The chapter begins with an overview of two large networks of mRNAs that contain conserved AU-rich elements (AREs) or GU-rich elements (GREs), and their role in mammalian cell physiology. Other, less conserved, cis-acting elements and their functional role in different steps of RNA maturation and metabolism will be discussed. The molecular characteristics of pathological cis-acting sequences that rose from gene mutations or transcriptional aberrations are briefly outlined, with the proposed approach to restore normal gene expression. Concise models of the function of posttranscriptional regulatory networks within different cellular compartments conclude this chapter

Regulating the Many to Benefit the Few: Role of Weak Small RNA Targets

AbstractSmall regulatory RNAs are central players in the regulation of many cellular processes across all kingdoms of life. Experiments in mouse and human have shown that a typical small RNA may regulate the expression of many different genes, suggesting that small RNAs act as global regulators. It is noted though that most targets respond only weakly to the presence of the small RNA. At the same time, evidence in bacteria and animals suggest that the phenotypes associated with small RNA mutants are only due to a few of their targets. Here we assume that targets regulated by a small RNA to control function is in fact small, and propose that the role of the many other weak targets is to confer robustness to the regulation of these few principal targets. Through mathematical modeling we show that auxiliary targets may significantly buffer both number and kinetic fluctuations of the principal targets, with only minor slowdown in the kinetics of response. Analysis of genomic data suggests that auxiliary targets experience a nonspecific evolutionary pressure, playing a role at the system level. Our work is of importance for studies on small RNA functions, and impacts on the understanding of small RNA evolution

Multiple Elements RegulateMash1Expression in the Developing CNS

AbstractMash1, a transcription factor of the basic helix–loop–helix class, is expressed during embryogenesis in restricted regions of the nervous system. An essential role for Mash1 in neural development was demonstrated previously in mice carrying a targeted disruption of theMash1gene. Regulation of the precise temporal and spatial expression ofMash1is thus likely to be important for proper neural development. In this study, sequences that regulateMash1expression in the central nervous system were characterized by assaying the expression oflacZreporter genes in transgenic embryos. A 1158-bp enhancer localized ∼7 kb upstream of theMash1coding region was identified. Deletions within this enhancer region reveal the presence of both positive and negativecis-acting elements. Analysis of multiple sequences within the enhancer demonstrate that different elements preferentially function in different regions within theMash1-specific CNS expression domain. In addition, a role for sequences 3′ of theMash1coding region is revealed, providing evidence for posttranscriptional control ofMash1expression in multiple CNS domains

Dissecting the expression relationships between RNA-binding proteins and their cognate targets in eukaryotic post-transcriptional regulatory networks

RNA-binding proteins (RBPs) are pivotal in orchestrating several steps in the metabolism of RNA in eukaryotes thereby controlling an extensive network of RBP-RNA interactions. Here, we employed CLIP (cross-linking immunoprecipitation)-seq datasets for 60 human RBPs and RIP-ChIP (RNP immunoprecipitation-microarray) data for 69 yeast RBPs to construct a network of genome-wide RBP- target RNA interactions for each RBP. We show in humans that majority (~78%) of the RBPs are strongly associated with their target transcripts at transcript level while ~95% of the studied RBPs were also found to be strongly associated with expression levels of target transcripts when protein expression levels of RBPs were employed. At transcript level, RBP - RNA interaction data for the yeast genome, exhibited a strong association for 63% of the RBPs, confirming the association to be conserved across large phylogenetic distances. Analysis to uncover the features contributing to these associations revealed the number of target transcripts and length of the selected protein-coding transcript of an RBP at the transcript level while intensity of the CLIP signal, number of RNA-Binding domains, location of the binding site on the transcript, to be significant at the protein level. Our analysis will contribute to improved modelling and prediction of post-transcriptional network

Target RNAs strike back on MicroRNAs

MicroRNAs are extensively studied regulatory non-coding small RNAs that silence animal genes throughout most biological processes, typically doing so by binding to partially complementary sequences within target RNAs. A plethora of studies has described detailed mechanisms for microRNA biogenesis and function, as well as their temporal and spatial regulation during development. By inducing translational repression and/or degradation of their target RNAs, microRNAs can contribute to achieve highly specific cell-or tissue-specific gene expression, while their aberrant expression can lead to disease. Yet an unresolved aspect of microRNA biology is how such small RNA molecules are themselves cleared from the cell, especially under circumstances where fast microRNA turnover or specific degradation of individual microRNAs is required. In recent years, it was unexpectedly found that binding of specific target RNAs to microRNAs with extensive complementarity can reverse the outcome, triggering degradation of the bound microRNAs. This emerging pathway, named TDMD for Target RNA-Directed MicroRNA Degradation, leads to microRNA 3′-end tailing by the addition of A/U non-templated nucleotides, trimming or shortening from the 3′ end, and highly specific microRNA loss, providing a new layer of microRNA regulation. Originally described in flies and known to be triggered by viral RNAs, novel endogenous instances of TDMD have been uncovered and are now starting to be understood. Here, we review our current knowledge of this pathway and its potential role in the control and diversification of microRNA expression patterns.Fil: Fuchs Wightman, Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Giono, Luciana Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Fededa, Juan Pablo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: de la Mata, Manuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; Argentin

Identifications of conserved 7-mers in 3'-UTRs and microRNAs in Drosophila

<p>Abstract</p> <p>Background</p> <p>MicroRNAs (miRNAs) are a class of endogenous regulatory small RNAs which play an important role in posttranscriptional regulations by targeting mRNAs for cleavage or translational repression. The base-pairing between the 5'-end of miRNA and the target mRNA 3'-UTRs is essential for the miRNA:mRNA recognition. Recent studies show that many seed matches in 3'-UTRs, which are fully complementary to miRNA 5'-ends, are highly conserved. Based on these features, a two-stage strategy can be implemented to achieve the <it>de novo </it>identification of miRNAs by requiring the complete base-pairing between the 5'-end of miRNA candidates and the potential seed matches in 3'-UTRs.</p> <p>Results</p> <p>We presented a new method, which combined multiple pairwise conservation information, to identify the frequently-occurred and conserved 7-mers in 3'-UTRs. A pairwise conservation score (PCS) was introduced to describe the conservation of all 7-mers in 3'-UTRs between any two <it>Drosophila </it>species. Using PCSs computed from 6 pairs of flies, we developed a support vector machine (SVM) classifier ensemble, named Cons-SVM and identified 689 conserved 7-mers including 63 seed matches covering 32 out of 38 known miRNA families in the reference dataset. In the second stage, we searched for 90 nt conserved stem-loop regions containing the complementary sequences to the identified 7-mers and used the previously published miRNA prediction software to analyze these stem-loops. We predicted 47 miRNA candidates in the genome-wide screen.</p> <p>Conclusion</p> <p>Cons-SVM takes advantage of the independent evolutionary information from the 6 pairs of flies and shows high sensitivity in identifying seed matches in 3'-UTRs. Combining the multiple pairwise conservation information by the machine learning approach, we finally identified 47 miRNA candidates in <it>D. melanogaster</it>.</p

Integrated analysis of transcript-level regulation of metabolism reveals disease-relevant nodes of the human metabolic network

Metabolic diseases and comorbidities represent an ever-growing epidemic where multiple cell types impact tissue homeostasis. Here, the link between the metabolic and gene regulatory networks was studied through experimental and computational analysis. Integrating gene regulation data with a human metabolic network prompted the establishment of an open-sourced web portal, IDARE (Integrated Data Nodes of Regulation), for visualizing various gene-related data in context of metabolic pathways. Motivated by increasing availability of deep sequencing studies, we obtained ChIP-seq data from widely studied human umbilical vein endothelial cells. Interestingly, we found that association of metabolic genes with multiple transcription factors (TFs) enriched disease-associated genes. To demonstrate further extensions enabled by examining these networks together, constraint-based modeling was applied to data from human preadipocyte differentiation. In parallel, data on gene expression, genome-wide ChIP-seq profiles for peroxisome proliferator-activated receptor (PPAR) γ, CCAAT/enhancer binding protein (CEBP) α, liver X receptor (LXR) and H3K4me3 and microRNA target identification for miR-27a, miR-29a and miR-222 were collected. Disease-relevant key nodes, including mitochondrial glycerol-3-phosphate acyltransferase (GPAM), were exposed from metabolic pathways predicted to change activity by focusing on association with multiple regulators. In both cell types, our analysis reveals the convergence of microRNAs and TFs within the branched chain amino acid (BCAA) metabolic pathway, possibly providing an explanation for its downregulation in obese and diabetic condition