microRNA Expression and Its Association With Disability and Brain Atrophy in Multiple Sclerosis Patients Treated With Glatiramer Acetate.

Background: MicroRNAs are small non-coding RNA that regulate gene expression at a post-transcriptional level affecting several cellular processes including inflammation, neurodegeneration and remyelination. Different patterns of miRNAs expression have been demonstrated in multiple sclerosis compared to controls, as well as in different courses of the disease. For these reason they have been postulated as promising biomarkers candidates in multiple sclerosis. Objective: To correlate serum microRNAs profile expression with disability, cognitive functioning and brain volume in patients with remitting-relapsing multiple sclerosis. Methods: Cross-sectional study in relapsing-remitting multiple sclerosis patients treated with glatiramer acetate. Disability was measured with Expanded Disability Status Scale (EDSS) and cognitive function was studied with Symbol Digit Modalities Test (SDMT). Brain volume was analyzed with automatic software NeuroQuant® . Results: We found an association between miR.146a.5p (rs:0.434, p=0.03) and miR.9.5p (rs:0.516, p=0.028) with EDSS; and miR-146a.5p (rs:-0.476, p=0.016) and miR-126.3p (rs:-0.528, p=0.007) with SDMT. Regarding to the brain volume, miR.9.5p correlated with thalamus (rs:-0.545, p=0.036); miR.200c.3p with pallidum (rs:-0.68, p=0.002) and cerebellum (rs:-0.472, p=0.048); miR-138.5p with amygdala (rs:0.73, p=0.016) and pallidum (rs:0.64, p=0.048); and miR-223.3p with caudate (rs:0.46, p=0.04). Conclusions: These data support the hypothesis of microRNA as potential biomarkers in this disease. More studies are needed to validate these results and to better understand the role of microRNAs in the pathogenesis, monitoring and therapeutic response of multiple sclerosis.post-print1410 K

SGNP: An Essential Stress Granule/Nucleolar Protein Potentially Involved in 5.8s rRNA Processing/Transport

Background: Stress Granules (SG) are sites of accumulation of stalled initiation complexes that are induced following a variety of cellular insults. In a genetic screen for factors involved in protecting human myoblasts from acute oxidative stress, we identified a gene encoding a protein we designate SGNP (Stress Granule and Nucleolar Protein). Methodology/Principal Findings: A gene-trap insertional mutagenesis screen produced one insertion that conferred resistance to sodium arsenite. RT-PCR/39 RACE was used to identify the endogenous gene expressed as a GFP-fusion transcript. SGNP is localized in both the cytoplasm and nucleolus and defines a non-nucleolar compartment containing 5.8S rRNA, a component of the 60S ribosomal subunit. Under oxidative stress, SGNP nucleolar localization decreases and it rapidly co-localizes with stress granules. The decrease in nucleolar SGNP following oxidative stress was accompanied by a large increase in nucleolar 5.8S rRNA. Knockdown of SGNP with shRNA increased global mRNA translation but induced growth arrest and cell death. Conclusions: These results suggest that SGNP is an essential gene that may be involved in ribosomal biogenesis and translational control in response to oxidative stress

Hypoxia Negatively Regulates Antimetastatic PEDF in Melanoma Cells by a Hypoxia Inducible Factor-Independent, Autophagy Dependent Mechanism

Pigment epithelium-derived factor (PEDF), a member of the serine protease inhibitor (SERPIN) superfamily, displays a potent antiangiogenic and antimetastatic activity in a broad range of tumor types. Melanocytes and low aggressive melanoma cells secrete high levels of PEDF, while its expression is lost in highly aggressive melanomas. PEDF efficiently abrogates a number of functional properties critical for the acquisition of metastatic ability by melanoma cells, such as neovascularization, proliferation, migration, invasiveness and extravasation. In this study, we identify hypoxia as a relevant negative regulator of PEDF in melanocytes and low aggressive melanoma cells. PEDF was regulated at the protein level. Importantly, although downregulation of PEDF was induced by inhibition of 2-oxoglutarate-dependent dioxygenases, it was independent of the hypoxia inducible factor (HIF), a key mediator of the adaptation to hypoxia. Decreased PEDF protein was not mediated by inhibition of translation through untranslated regions (UTRs) in melanoma cells. Degradation by metalloproteinases, implicated on PEDF degradation in retinal pigment epithelial cells, or by the proteasome, was also excluded as regulatory mechanism in melanoma cells. Instead, we found that degradation by autophagy was critical for PEDF downregulation under hypoxia in human melanoma cells. Our findings show that hypoxic conditions encountered during primary melanoma growth downregulate antiangiogenic and antimetastasic PEDF by a posttranslational mechanism involving degradation by autophagy and could therefore contribute to the acquisition of highly metastatic potential characteristic of aggressive melanoma cells

The RNA binding protein HuR differentially regulates unique subsets of mRNAs in estrogen receptor negative and estrogen receptor positive breast cancer

<p>Abstract</p> <p>Background</p> <p>The discordance between steady-state levels of mRNAs and protein has been attributed to posttranscriptional control mechanisms affecting mRNA stability and translation. Traditional methods of genome wide microarray analysis, profiling steady-state levels of mRNA, may miss important mRNA targets owing to significant posttranscriptional gene regulation by RNA binding proteins (RBPs).</p> <p>Methods</p> <p>The ribonomic approach, utilizing RNA immunoprecipitation hybridized to microarray (RIP-Chip), provides global identification of putative endogenous mRNA targets of different RBPs. HuR is an RBP that binds to the AU-rich elements (ARE) of labile mRNAs, such as proto-oncogenes, facilitating their translation into protein. HuR has been shown to play a role in cancer progression and elevated levels of cytoplasmic HuR directly correlate with increased invasiveness and poor prognosis for many cancers, including those of the breast. HuR has been described to control genes in several of the acquired capabilities of cancer and has been hypothesized to be a tumor-maintenance gene, allowing for cancers to proliferate once they are established.</p> <p>Results</p> <p>We used HuR RIP-Chip as a comprehensive and systematic method to survey breast cancer target genes in both MCF-7 (estrogen receptor positive, ER+) and MDA-MB-231 (estrogen receptor negative, ER-) breast cancer cell lines. We identified unique subsets of HuR-associated mRNAs found individually or in both cell types. Two novel HuR targets, <it>CD9 </it>and <it>CALM2 </it>mRNAs, were identified and validated by quantitative RT-PCR and biotin pull-down analysis.</p> <p>Conclusion</p> <p>This is the first report of a side-by-side genome-wide comparison of HuR-associated targets in wild type ER+ and ER- breast cancer. We found distinct, differentially expressed subsets of cancer related genes in ER+ and ER- breast cancer cell lines, and noted that the differential regulation of two cancer-related genes by HuR was contingent upon the cellular environment.</p

Post-transcriptional control during chronic inflammation and cancer: a focus on AU-rich elements

A considerable number of genes that code for AU-rich mRNAs including cytokines, growth factors, transcriptional factors, and certain receptors are involved in both chronic inflammation and cancer. Overexpression of these genes is affected by aberrations or by prolonged activation of several signaling pathways. AU-rich elements (ARE) are important cis-acting short sequences in the 3′UTR that mediate recognition of an array of RNA-binding proteins and affect mRNA stability and translation. This review addresses the cellular and molecular mechanisms that are common between inflammation and cancer and that also govern ARE-mediated post-transcriptional control. The first part examines the role of the ARE-genes in inflammation and cancer and sequence characteristics of AU-rich elements. The second part addresses the common signaling pathways in inflammation and cancer that regulate the ARE-mediated pathways and how their deregulations affect ARE-gene regulation and disease outcome

Role of the RNA-Binding Protein Nrd1 in Stress Granule Formation and Its Implication in the Stress Response in Fission Yeast

We have previously identified the RNA recognition motif (RRM)-type RNA-binding protein Nrd1 as an important regulator of the posttranscriptional expression of myosin in fission yeast. Pmk1 MAPK-dependent phosphorylation negatively regulates the RNA-binding activity of Nrd1. Here, we report the role of Nrd1 in stress-induced RNA granules. Nrd1 can localize to poly(A)-binding protein (Pabp)-positive RNA granules in response to various stress stimuli, including heat shock, arsenite treatment, and oxidative stress. Interestingly, compared with the unphosphorylatable Nrd1, Nrd1DD (phosphorylation-mimic version of Nrd1) translocates more quickly from the cytoplasm to the stress granules in response to various stimuli; this suggests that the phosphorylation of Nrd1 by MAPK enhances its localization to stress-induced cytoplasmic granules. Nrd1 binds to Cpc2 (fission yeast RACK) in a phosphorylation-dependent manner and deletion of Cpc2 affects the formation of Nrd1-positive granules upon arsenite treatment. Moreover, the depletion of Nrd1 leads to a delay in Pabp-positive RNA granule formation, and overexpression of Nrd1 results in an increased size and number of Pabp-positive granules. Interestingly, Nrd1 deletion induced resistance to sustained stresses and enhanced sensitivity to transient stresses. In conclusion, our results indicate that Nrd1 plays a role in stress-induced granule formation, which affects stress resistance in fission yeast

Screening for Microsatellite Instability Identifies Frequent 3′-Untranslated Region Mutation of the RB1-Inducible Coiled-Coil 1 Gene in Colon Tumors

BACKGROUND: Coding region microsatellite instability (MSI) results in loss of gene products and promotion of microsatellite-unstable (MSI-H) carcinogenesis. Recent studies have indicated that MSI within 3'-untranslated regions (3'UTRs) may post-transcriptionally dysregulate gene products. Within this context, we conducted a broad mutational survey of 42 short 3'UTR microsatellites (MSs) in 45 MSI-H colorectal tumors and their corresponding normal colonic mucosae. METHODOLOGY/PRINCIPAL FINDINGS: In order to estimate the overall susceptibility of MSs to MSI in MSI-H tumors, the observed MSI frequency of each MS was correlated with its length, interspecies sequence conservation level, and distance from some genetic elements (i.e., stop codon, polyA signal, and microRNA binding sites). All MSs were stable in normal colonic mucosae. The MSI frequency at each MS in MSI-H tumors was independent of sequence conservation level and distance from other genetic elements. In contrast, MS length correlated significantly with MSI frequency in MSI-H tumors (r=0.86, p=7.2x10(-13)). 3'UTR MSs demonstrated MSI frequencies in MSI-H tumors higher than the 99% upper limit predicted by MS length for RB1-inducible coiled-coil 1(RB1CC1, mutation frequency 68.4%), NUAK family SNF1-like kinase 1(NUAK1, 31.0%), and Rtf1, Paf1/RNA polymerase II complex component, homolog (RTF1, 25.0%). An in silico prediction of RNA structure alterations was conducted for these MSI events to gauge their likelihood of affecting post-transcriptional regulation. RB1CC1 mutant was predicted to lose a microRNA-accessible loop structure at a putative binding site for the tumor-suppressive microRNA, miR-138. In contrast, the predicted 3'UTR structural change was minimal for NUAK1- and RTF1 mutants. Notably, real-time quantitative RT-PCR analysis revealed significant RB1CC1 mRNA overexpression vs. normal colonic mucosae in MSI-H cancers manifesting RB1CC1 3'UTR MSI (9.0-fold; p = 3.6x10(-4)). CONCLUSIONS: This mutational survey of well-characterized short 3'UTR MSs confirms that MSI incidence in MSI-H colorectal tumors correlates with MS length, but not with sequence conservation level or distance from other genetic elements. This study also identifies RB1CC1 as a novel target of frequent mutation and aberrant upregulation in MSI-H colorectal tumors. The predicted loss of a microRNA-accessible structure in mutant RB1CC1 RNA fits the hypothesis that 3'UTR MSI involves in aberrant RB1CC1 posttranscriptional upregulation. Further direct assessments are indicated to investigate this possibility.Bogdan C. Paun, Yulan Cheng, Barbara A. Leggett, Joanne Young, Stephen J. Meltzer, Yuriko Mor

Post-transcriptional gene regulation: From genome-wide studies to principles

Post-transcriptional regulation of gene expression plays important roles in diverse cellular processes such as development, metabolism and cancer progression. Whereas many classical studies explored the mechanistics and physiological impact on specific mRNA substrates, the recent development of genome-wide analysis tools enables the study of post-transcriptional gene regulation on a global scale. Importantly, these studies revealed distinct programs of RNA regulation, suggesting a complex and versatile post-transcriptional regulatory network. This network is controlled by specific RNA-binding proteins and/or non-coding RNAs, which bind to specific sequence or structural elements in the RNAs and thereby regulate subsets of mRNAs that partly encode functionally related proteins. It will be a future challenge to link the spectra of targets for RNA-binding proteins to post-transcriptional regulatory programs and to reveal its physiological implications

iCLIP Predicts the Dual Splicing Effects of TIA-RNA Interactions

Transcriptome-wide analysis of protein-RNA interactions predicts the dual splicing effects of TIA proteins, showing that their local enhancing function is associated with diverse distal splicing silencing effects