RESUMEN
Activating transcription factor (ATF) 5 is a transcription factor belonging to the ATF/cAMP-response element-binding protein gene family. We previously reported that ATF5 mRNA expression increased in response to amino acid limitation. The ATF5 gene allows transcription of mRNAs with at least two alternative 5'-untranslated regions (5'-UTRs), 5'-UTRalpha and 5'-UTRbeta, derived from exon1alpha and exon1beta. 5'-UTRalpha contains highly conserved sequences, in which the upstream open reading frames (uORFs) uORF1 and uORF2 are found in many species. This study was designed to investigate the potential role of 5'-UTRs in translational control. These 5'-UTRs differentially determined translation efficiency from mRNA. The presence of 5'-UTRalpha or 5'-UTRbeta represses translation from the downstream ATF5 ORF. Moreover, 5'-UTRalpha-regulated translational repression is released by amino acid limitation or NaAsO(2) exposure. This release was not seen for 5'-UTRbeta. Mutation of uAUG2 in the uORF2 of 5'-UTRalpha restored the basal expression and abolished the positive regulation by amino acid limitation or arsenite exposure. We demonstrated that phosphorylation of eukaryotic initiation factor 2alpha was required for amino acid limitation-induced translational regulation of ATF5. Furthermore, arsenite exposure activated the exogenously expressed heme-regulated inhibitor kinase and induced the phosphorylation of eukaryotic initiation factor 2alpha in nonerythroid cells. These results suggest that translation of ATF5 is regulated by the alternative 5'-UTR region of its mRNA, and ATF5 may play a role in protecting cells from amino acid limitation or arsenite-induced oxidative stress.
Asunto(s)
Factores de Transcripción Activadores/genética , ARN Mensajero/genética , Regiones no Traducidas 5' , Aminoácidos/metabolismo , Animales , Arsenitos/farmacología , Secuencia de Bases , Células COS , Línea Celular , Chlorocebus aethiops , Cartilla de ADN/genética , Factor 2 Eucariótico de Iniciación/metabolismo , Células HeLa , Humanos , Ratones , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Sistemas de Lectura Abierta , Estrés Oxidativo , Fosforilación , Biosíntesis de Proteínas/efectos de los fármacos , Homología de Secuencia de Ácido NucleicoRESUMEN
ATF5 is a transcription factor in the cAMP response element (CRE)-binding protein/activating transcription factor (CREB/ATF) family. We studied the effect of amino acid limitation on ATF5 mRNA levels in a mammalian cell line. Northern-blot analysis demonstrated that limitation of a single amino acid, glutamine, methionine, or leucine, resulted in increased ATF5 mRNA levels in HeLaS3 cells. This resulted, at least in part, from increased half-life of the ATF5 mRNA transcript. Cycloheximide inhibited the increase in ATF5 mRNA expression induced by glutamine limitation, indicating that it was dependent on de novo protein synthesis. Moreover, rapamycin had no effect on basal ATF5 mRNA expression or on increased expression induced by glutamine limitation. These results indicate that amino acid limitation regulates ATF5 mRNA expression during post-transcription in a rapamycin-independent manner. The potential role for ATF5 in protecting cells from amino acid-limitation is of considerable interest.
