RESUMO
AU-rich elements (AREs, usually containing repeated copies of AUUUA), when present in the 3'-untranslated regions (UTRs) of many mammalian mRNAs, confer instability on their host RNA molecules. The viral small nuclear RNA (snRNA) Herpesvirus saimiri U RNA 1 (HSUR 1) also contains an AUUUA-rich sequence. Here, we report that this ARE induces rapid degradation of HSUR 1 itself and of other snRNAs including HSUR 2 and cellular U1. Mutational analyses of the viral ARE establish that sequence requirements for mRNA and snRNA decay are the same, suggesting a similar mechanism. Moreover, the in vivo degradation activity of mutant AREs correlates with their in vitro binding to the HuR protein, implicated previously as a component of the mRNA degradation machinery. Our results suggest that ARE-mediated instability can be uncoupled from both ongoing translation and deadenylation of the target RNA.
Assuntos
Antígenos de Superfície , Herpesvirus Saimiriíneo 2/química , RNA Mensageiro/metabolismo , RNA Nuclear Pequeno/metabolismo , RNA Viral/metabolismo , Sequência de Bases , Proteínas ELAV , Proteína Semelhante a ELAV 1 , Regulação da Expressão Gênica/genética , Genes Reporter , Globinas/genética , Herpesvirus Saimiriíneo 2/genética , Dados de Sequência Molecular , Mutação , RNA Mensageiro/genética , RNA Nuclear Pequeno/genética , RNA Viral/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Sequências Repetitivas de Ácido Nucleico , Ribonucleases/metabolismo , Transcrição Gênica , TransfecçãoRESUMO
Expression of many proto-oncogenes, cytokines and lymphokines is regulated by targeting their messenger RNAs for rapid degradation. Essential signals for this control are AU-rich elements (AREs) in the 3' untranslated region (UTR) of these messages. The ARE is loosely defined as the five-nucleotide sequence AUUUA embedded in a uracil-rich region. A transacting factor, presumably a protein, binds the ARE and initiates recognition by the destabilization machinery. Numerous candidate ARE-binding proteins have been proposed. We show that a 32 kDa protein in HeLa nuclear extracts characterized previously has RNA-binding specificity that correlates with the activity of an ARE in directing mRNA decay. Purification and subsequent analyses demonstrate that this 32 kDa protein is identical to a recently identified member of the Elav-like gene family (ELG) called HuR. The in vitro binding selectivity of HuR is indicative of an ARE sequence's ability to destabilize a mRNA in vivo, suggesting a critical role for HuR in the regulation of mRNA degradation.
Assuntos
Antígenos de Superfície , Regulação da Expressão Gênica/fisiologia , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Células 3T3 , Sequência de Aminoácidos , Animais , Composição de Bases , Extratos Celulares , Reagentes de Ligações Cruzadas , Proteínas ELAV , Proteína Semelhante a ELAV 1 , Células HeLa , Humanos , Camundongos , Dados de Sequência Molecular , Peso Molecular , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/isolamento & purificação , Sequências Reguladoras de Ácido Nucleico/genética , Raios UltravioletaRESUMO
Herpesvirus saimiri (HVS) is one of several primate viruses that carry genes for small RNAs. The five H. saimiri-encoded U RNAs (HSURs) are the most abundant viral transcripts expressed in transformed marmoset T lymphocytes. They assemble with host proteins common to spliceosomal small nuclear ribonucleoproteins (snRNPs). HSURs 1, 2, and 5 exhibit sequences at their 5' ends identical to the AUUUA motif, which targets a number of protooncogene, cytokine, and lymphokine mRNAs for rapid degradation. We show that a 32-kDa protein previously demonstrated to bind to the 3' untranslated region of several unstable messages can be UV crosslinked specifically to HSUR 1, 2, and 5 transcripts in vitro, as well as to endogenous HSUR snRNPs. Our results suggest an unusual role for these viral snRNPs: HSURs may function to attenuate the rapid degradation of certain cellular mRNAs, thereby facilitating viral transformation of host T lymphocytes.