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Nucleotide composition of cellular internal ribosome entry sites defines dependence on NF45 and predicts a posttranscriptional mitotic regulon.

ABSTRACT: The vast majority of cellular mRNAs initiate their translations through a well-defined mechanism of ribosome recruitment that occurs at the 5'-terminal 7-methylguanosine cap with the help of several canonical protein factors. A subset of cellular and viral mRNAs contain regulatory motifs in their 5' untranslated regions (UTRs), termed internal ribosome entry sites (IRES), that sidestep this canonical mode of initiation. On cellular mRNAs, this mechanism requires IRES trans-acting protein factors (ITAFs) that facilitate ribosome recruitment downstream of the cap. While several ITAFs and their target mRNAs have been empirically identified, the in silico prediction of targets has proved difficult. Here, we report that a high AU content (>60%) of the IRES-containing 5' UTRs serves as an excellent predictor of dependence on NF45, a recently identified ITAF. Moreover, we provide evidence that cells deficient in NF45 ITAF activity exhibit reduced IRES-mediated translation of X-linked inhibitor of apoptosis protein (XIAP) and cellular inhibitor of apoptosis protein 1 (cIAP1) mRNAs that, in turn, leads to dysregulated expression of their respective targets, survivin and cyclin E. This specific defect in IRES translation explains in part the cytokinesis impairment and senescence-like phenotype observed in HeLa cells expressing NF45 RNA interference (RNAi). This study uncovers a novel role for NF45 in regulating ploidy and highlights the importance of IRES-mediated translation in cellular homeostasis.

SUBMITTER: Faye MD

PROVIDER: S-EPMC3554113 | biostudies-literature | 2013 Jan

REPOSITORIES: biostudies-literature

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Nucleotide composition of cellular internal ribosome entry sites defines dependence on NF45 and predicts a posttranscriptional mitotic regulon.

Faye Mame Daro MD Graber Tyson E TE Liu Peng P Thakor Nehal N Baird Stephen D SD Durie Danielle D Holcik Martin M

Molecular and cellular biology 20121105 2

The vast majority of cellular mRNAs initiate their translations through a well-defined mechanism of ribosome recruitment that occurs at the 5'-terminal 7-methylguanosine cap with the help of several canonical protein factors. A subset of cellular and viral mRNAs contain regulatory motifs in their 5' untranslated regions (UTRs), termed internal ribosome entry sites (IRES), that sidestep this canonical mode of initiation. On cellular mRNAs, this mechanism requires IRES trans-acting protein factors ...[more]

PMID: 23129811

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Project description:In recent years, hepatitis C virus (HCV)-related viruses were identified in several species, including dogs, horses, bats, and rodents. In addition, a novel virus of the genus Hepacivirus has been discovered in bovine samples and was termed bovine hepacivirus (BovHepV). Prediction of the BovHepV internal ribosome entry site (IRES) structure revealed strong similarities to the HCV IRES structure comprising domains II, IIIabcde, pseudoknot IIIf, and IV with the initiation codon AUG. Unlike HCV, only one microRNA-122 (miR-122) binding site could be identified in the BovHepV 5' nontranslated region. In this study, we analyzed the necessity of BovHepV IRES domains to initiate translation and investigated possible interactions between the IRES and core coding sequences by using a dual luciferase reporter assay. Our results suggest that such long-range interactions within the viral genome can affect IRES-driven translation. Moreover, the significance of a possible miR-122 binding to the BovHepV IRES was investigated. When analyzing translation in human Huh-7 cells with large amounts of endogenous miR-122, introduction of point mutations to the miR-122 binding site resulted in reduced translation efficiency. Similar results were observed in HeLa cells after substitution of miR-122. Nevertheless, the absence of pronounced effects in a bovine hepatocyte cell line expressing hardly any miR-122 as well suggests additional functions of this host factor in virus replication.IMPORTANCE Several members of the family Flaviviridae, including HCV, have adapted cap-independent translation strategies to overcome canonical eukaryotic translation pathways and use cis-acting RNA-elements, designated viral internal ribosome entry sites (IRES), to initiate translation. Although novel hepaciviruses have been identified in different animal species, only limited information is available on their biology on molecular level. Therefore, our aim was a fundamental analysis of BovHepV IRES functions. The findings which show that functional IRES elements are also crucial for BovHepV translation expand our knowledge on molecular mechanism of hepacivirus propagation. We also studied the possible effects of one major host factor implicated in HCV pathogenesis, miR-122. The results of mutational analyses suggested that miR-122 enhances virus translation mediated by BovHepV IRES.

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Nucleotide composition of cellular internal ribosome entry sites defines dependence on NF45 and predicts a posttranscriptional mitotic regulon.

Publications

Nucleotide composition of cellular internal ribosome entry sites defines dependence on NF45 and predicts a posttranscriptional mitotic regulon.

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