RESUMO
The bcl-X gene plays a critical role in apoptosis. Six different isoforms generated by tissue-specific promoter usage and alternative splicing were described. Some of them exert opposite effects on cell death. In mammary epithelial cells glucocorticoids induce bcl-X expression and increase the ratio bcl-X(L) (antiapoptotic)/bcl-X(S) (apoptotic) by activating P4 promoter, which contains two hormone response elements. Here we show that, on mouse thymocytes and T lymphocyte derivative S49 cells, glucocorticoids inhibited transcription from P4 and decreased the ratio bcl-X(L)/bcl-X(S) favoring apoptosis. Upon hormonal treatment, glucocorticoid receptor (GR), steroid receptor coactivator-1, and RNA polymerase II were transiently recruited to P4 promoter, whereas STAT5B was also recruited but remained bound. Concomitant with the release of GR, silencing mediator for retinoic acid receptor and thyroid hormone receptor and histone deacetylase 3 were recruited, histone H3 was deacetylated, and RNA polymerase II left the promoter. Inhibition of STAT5 activity reverted glucocorticoid repression to activation of transcription and was accompanied by stable recruitment of GR and RNA polymerase II to P4.
Assuntos
Dexametasona/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Linfócitos/metabolismo , Regiões Promotoras Genéticas/genética , Fator de Transcrição STAT5/metabolismo , Timo/metabolismo , Proteína bcl-X/metabolismo , Acetilação , Animais , Apoptose , Células COS , Chlorocebus aethiops , Imunoprecipitação da Cromatina , Ensaio de Desvio de Mobilidade Eletroforética , Histona Acetiltransferases/metabolismo , Histonas/metabolismo , Antagonistas de Hormônios/farmacologia , Linfócitos/citologia , Masculino , Camundongos , Mifepristona/farmacologia , Coativador 1 de Receptor Nuclear , Plasmídeos/metabolismo , RNA Polimerase II/metabolismo , RNA Mensageiro/metabolismo , Receptores de Glucocorticoides/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Timo/citologia , Fatores de Transcrição/metabolismo , Transcrição Gênica , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/genéticaRESUMO
Bcl-X exists in at least five different isoforms with complex effects on programmed cell death. Glucocorticoids and progestins control bcl-X expression and influence the ratio between bcl-X(L) (antiapoptotic isoform) and bcl-X(S) (proapoptotic isoform) in different tissues. The 5'-UTR region of the mouse bcl-X gene contains at least five different promoters, which exhibit a tissue-specific pattern of promoter usage. Several mRNAs with different 5'-leading exons can be generated upon promoter activation. Here we explore the potential of the various bcl-X gene promoters to be regulated by glucocorticoids or progestins. We found that the region located immediately upstream of promoter 4 (P4) contains two hormone response element (HRE)-like sequences at positions -3040 (HRE I) and -3001 (HRE II) relative to the translation initiation codon. These HRE-like sequences confer hormone responsiveness to a core promoter and bind glucocorticoid or progesterone receptors in vitro. Point mutations of both HREs that prevent steroid receptor binding also eliminate hormonal inducibility. In cells treated with glucocorticoids, the hormone receptor is recruited to the P4 region containing the HREs. Analysis of the products of the endogenous bcl-X in epithelial mammary cells showed that only transcripts originating from P4 increased upon hormone treatment. This observation correlates with the induction of the bcl-X(L) mRNA, suggesting that P4 is one of the bcl-X promoters responsible for the generation of this antiapoptotic isoform.