Additional file 4: of RIP-Chip analysis supports different roles for AGO2 and GW182 proteins in recruiting and processing microRNA targets

Overview of gene expression levels in IP and FT samples. Focus on the enriched genes in AGO2-IP and GW182-IP vs FT samples. The reported expression levels are computed as the average values of the three performed experimental replicates. (PDF 1423 kb)

A multiomics analysis of S100 protein family in breast cancer

The S100 gene family is the largest subfamily of calcium binding proteins of EF-hand type, expressed in tissue and cell-specific manner, acting both as intracellular regulators and extracellular mediators. There is a growing interest in the S100 proteins and their relationships with different cancers because of their involvement in a variety of biological events closely related to tumorigenesis and cancer progression. However, the collective role and the possible coordination of this group of proteins, as well as the functional implications of their expression in breast cancer (BC) is still poorly known. We previously reported a large-scale proteomic investigation performed on BC patients f…

Statistical Validation of a Comprehensive Gene/miRNA Expression Profile Dataset for miRNA:mRNA Interaction Analysis

Identification of a prognostic gene signature associated with MBP-1 expression in ErbB2-negative breast carcinomas

The ENO1 transcript, which encodes the glycolitic enzyme alpha-enolase, can be translated into a shorter nuclear protein called Myc-promoter Binding Protein-1 (MBP-1) by using an alternative translation start site. MBP-1 acts as a negative regulator of c-Myc, ErbB2 and Cox2 genes (1). Several evidences indicate that MBP-1 acts as a tumor suppressor in breast carcinoma and prostate cancer and its expression results in a reduced invasive ability (2). In our previous studies, we showed that MBP-1 is expressed and easily detectable in normal breast epithelial cells, but a loss of expression occurs in most primary invasive ductal carcinomas (IDC) of the breast. Furthermore, in these tumors MBP-1…

Transcriptomic Changes Following Partial Depletion of CENP-E in Normal Human Fibroblasts

The centromere is a fundamental chromosome structure in which the macro-molecular kinetochore assembles and is bound by spindle microtubules, allowing the segregation of sister chromatids during mitosis. Any alterations in kinetochore assembly or functioning or kinetochore–microtubule attachments jeopardize chromosome stability, leading to aneuploidy, a common feature of cancer cells. The spindle assembly checkpoint (SAC) supervises this process, ensuring a faithful segregation of chromosomes. CENP-E is both a protein of the kinetochore and a crucial component of the SAC required for kinetochore–microtubule capture and stable attachment, as well as congression of chromosomes to the metaphas…

Statistical validation of a comprehensive gene/miRNA expression profile dataset for miRNA:mRNA interctome analysis

Additional file 6: of RIP-Chip analysis supports different roles for AGO2 and GW182 proteins in recruiting and processing microRNA targets

Wilcoxon test p-values summary. Wilcoxon test p-values (log10) obtained by comparing the variable values associated with the enriched/underrepresented genes sets. Three different miRNA target prediction tools (Targetscan, PITA and miRanda) were used to compute the necessary binding sites (BS) matrices. The BS matrices used to compute the p-values in the last panel were obtained by considering BS predicted by at least two of the three prediction tools. In each panel, the variables computed with the three AGO2 IN profiles were used to distinguish enriched and underrepresented genes in AGO2-IP vs FT and the variables computed with the three GW182 IN profiles were used to distinguish enriched a…

Detecting significant features in modeling microRNA-target interactions

MicroRNAs (miRNAs) are small non-coding RNA molecules mediating the translational repression and degradation of target mRNAs in the cell. Mature miRNAs are used as a template by the RNA-induced silencing complex (RISC) to recognize the complementary mRNAs to be regulated. Up to 60% of human genes are putative targets of one or more miRNAs. Several prediction tools are available to suggest putative miRNA targets, however, only a small part of the interaction pairs has been validated by experimental approaches. The analysis of the expression profile of the RNA fraction immunoprecipitated (IP) with the RISC proteins is an established method to detect which genes are actually regulated by the R…

Additional file 7: of RIP-Chip analysis supports different roles for AGO2 and GW182 proteins in recruiting and processing microRNA targets

Summary of miRNA expression profiles shuffling effects. ROC analysis was performed to evaluate the performance of F6 and F4d variables, computed with simulated miRNA profiles, in distinguishing enriched/underrepresented genes in AGO2 or GW182-IP samples. Each panel reports the AUC values obtained with simulated variables. Each boxplot refers to AUC values obtained with a specific set of simulations, where the expression profile of a set of miRNAs was shuffled. The boxplot in the center was obtained by shuffling all miRNAs. The boxplots from the center to the right refer to simulations where all the miRNAs were shuffled with the exception of n top expressed miRNAs, n increasing in the right …

Relazione tra l’espressione del repressore trascrizionale MBP-1 e di miRNAs nel carcinoma mammario

MBP-1 (Myc-promoter-binding-protein-1) è una proteina di 37 kDa, prodotta dalla traduzione alternativa del mRNA del gene ENO1, codificante per l’enzima glicolitico α-enolasi. MBP-1 svolge il ruolo di repressore trascrizionale agendo direttamente sul promotore di c-MYC, ERBB2 e COX2, tutti geni coinvolti in diverse fasi della progressione tumorale (1-3). Nei carcinomi duttali infiltranti della mammella (IDC), l’espressione di MBP-1 è inversamente correlata alla comparsa di metastasi linfonodali e di recidive (4). In diverse linee cellulari tumorali l’overespressione di MBP-1 risulta nella riduzione della proliferazione cellulare e dell’invasività e in un aumento nella morte cellulare per apo…

Dal trascrittoma all’interattoma di miRNA: identificazione sperimentale e bioinformatica delle interazioni funzionali miRNA:mRNA

I miRNA, piccole molecole endogene di RNA non codificante, regolano l’espressione genica attraverso la degradazione dei messaggeri (mRNA) o l’inibizione della traduzione. I miRNA maturi interagiscono con le proteine del complesso RISC (RNA-induced silencing complex) tra cui le proteine Argonaute (Ago), capaci di legare direttamente i miRNA e di mediare la regolazione dell’espressione genica in seguito alla interazione del miRNA con il proprio mRNA target. Un singolo miRNA può legare diversi mRNA e ciascun mRNA può essere regolato da diversi miRNA. La maggior parte dei software di predizione oggi disponibili individuano i putativi target di singoli miRNA ignorando caratteristiche di tipo glo…

RIP-Chip analysis supports different roles for AGO2 and GW182 proteins in recruiting and processing microRNA targets.

Background MicroRNAs (miRNAs) are small non-coding RNA molecules mediating the translational repression and degradation of target mRNAs in the cell. Mature miRNAs are used as a template by the RNA-induced silencing complex (RISC) to recognize the complementary mRNAs to be regulated. To discern further RISC functions, we analyzed the activities of two RISC proteins, AGO2 and GW182, in the MCF-7 human breast cancer cell line. Methods We performed three RIP-Chip experiments using either anti-AGO2 or anti-GW182 antibodies and compiled a data set made up of the miRNA and mRNA expression profiles of three samples for each experiment. Specifically, we analyzed the input sample, the immunoprecipita…

Additional file 5: of RIP-Chip analysis supports different roles for AGO2 and GW182 proteins in recruiting and processing microRNA targets

Venn diagram of lists of enriched genes. The considered lists are: AGO2-IP (UP_AGO2 set), the list of enriched genes detected by Fan et al. [13] (UP_AGO2_Fan) and our list of enriched genes in GW182-IP sample (UP_GW182). The reported p-values refer to the closest intersection set of genes and are computed with one tail Fisher-test. (PDF 34 kb)

The miRNA profile associated with MBP-1 expression in breast cancer SKBR3 cells

Additional file 1: of RIP-Chip analysis supports different roles for AGO2 and GW182 proteins in recruiting and processing microRNA targets

Analysis of miRNA expression in AGO2 and GW182-IP samples. a) miRNA expression level in AGO2-IP samples (average value from the three performed experiments) vs the expression level in IN samples (average value from the three performed experiments). The Pearson correlation values reported on the top of the picture were computed by using all the expressed miRNA, and the top 100 or 50 expressed miRNAs. The colored points refer to miRNA that have been validated by RT-PCR data. Green points refer to hsa-miR-141-3p, hsa-miR-21-5p, hsa-let-7f-5p, hsa-miR-16-5p, hsa-miR-24-3p, hsa-miR-27a-3p, hsa-miR-23a-3p. The red point refers to hsa-miR-1260a. b) Comparison of IP/IN ratios obtained by RT-PCR dat…

Additional file 8: of RIP-Chip analysis supports different roles for AGO2 and GW182 proteins in recruiting and processing microRNA targets

Empirical Cumulative Distribution Function of 3â UTR and coding region length of IP-Enriched genes. Enriched genes in AGO (1â 4) and in GW182 protein family IP selected by considering log2 IP-Enrichment of transcript greater than 1. Data are downloaded from Landthaler et al. [14]. The Empirical Cumulative Distribution Function of the 3â UTR length (top) and coding region length (bottom) of genes enriched exclusively by AGO-IP (red line), GW182-IP (blue line) and both IPs (black line) are reported. The reported p-value is computed by performing a Wilcoxon test to compare the length distributions of genes enriched exclusively in AGO-IP and in GW182-IP. (PDF 145 kb)

Additional file 9: of RIP-Chip analysis supports different roles for AGO2 and GW182 proteins in recruiting and processing microRNA targets

Summary of miRNA expression profiles switch between experiment replicas. ROC analysis of F6&F4d SVM model trained with variables calculated with miRNA expression profiles from each of the three anti-AGO2 RIP experiments. SVM models were used to classify the top 1000 and the bottom 1000 genes with respect to the IP/FT mRNA expression ratio, computed for each of the three AGO2 RIP experiments. (PDF 653 kb)

Aneuploid IMR90 cells induced by depletion of pRB, DNMT1 and MAD2 show a common gene expression signature

Chromosome segregation defects lead to aneuploidy which is a major feature of solid tumors. How diploid cells face chromosome mis-segregation and how aneuploidy is tolerated in tumor cells are not completely defined yet. Thus, an important goal of cancer genetics is to identify gene networks that underlie aneuploidy and are involved in its tolerance. To this aim, we induced aneuploidy in IMR90 human primary cells by depleting pRB, DNMT1 and MAD2 and analyzed their gene expression profiles by microarray analysis. Bioinformatic analysis revealed a common gene expression profile of IMR90 cells that became aneuploid. Gene Set Enrichment Analysis (GSEA) also revealed gene-sets/pathways that are …

MULTIOMIC ANALYSIS OF TRANSCRIPTIONAL REPRESSOR MBP-1 FUNCTIONS IN GYNECOLOGICAL TUMORS

Additional file 3: of RIP-Chip analysis supports different roles for AGO2 and GW182 proteins in recruiting and processing microRNA targets

Summary of enriched and underrepresented genes. Summary of enriched and underrepresented genes in AGO2 and GW182-IP vs FT comparisons performed by SAMR (column 2–3). The enrichment results obtained with the REA algorithm are reported in columns 4–5. Columns 6 and 7 report the 3’UTR and Coding region (CR) lengths respectively. In columns 8–21 we report the number of binding sites predicted by Targetscan in the 3’UTR and the Coding region of seven highly expressed miRNAs. (XLS 294 kb)

Additional file 2: of RIP-Chip analysis supports different roles for AGO2 and GW182 proteins in recruiting and processing microRNA targets

Gene set enrichment analysis results with seven top expressed miRNA predicted targets sets. Predicted targets of miRNAs (column 1) were predicted with three different target prediction tools (column 2). The total number of predicted targets is indicated in column 3. Five lists of genes were analyzed. For each list of genes the number of genes in common with the predicted targets and the associated hypergeometric test pvalue are provided. The total number of genes considered in the analysis is 16,392. The five considered lists are: a list of genes enriched in AGO2 IP sample from [13]; lists of genes enriched in AGO2 IP vs IN and IP vs FT samples; lists of genes enriched in GW182 IP vs IN and…