RESUMO
Apoptosis, an intrinsic cellular pathway that eliminates unwanted cells from multicellular organisms, represents an important mechanism for protection against viral infections. When cells infected by viruses get recognized by immune cells, apoptosis is triggered in the infected cells. Among the many regulators of apoptosis involved in this process, a family of proteins homologous to oncogene Bcl-2 plays a central role. Their concerted activities converge to permeabilization of mitochondrial membranes and activation of apoptotic pathways in the presence of diverse apoptotic signals, including virus infection. In the genomes of many viruses, genes encoding for homologues of antiapoptotic proteins of Bcl-2 family can be found. These proteins, collectively referred to as vBcl-2 proteins, inhibit apoptosis in infected cells at the different stages of virus life cycle to enable the virus to complete its replication and to spread.
Assuntos
Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Virais/genética , Animais , Apoptose/genética , Humanos , Viroses/genética , Replicação Viral/genética , Vírus/genéticaRESUMO
The mitochondrial ADP/ATP carrier in Saccharomyces cerevisiae is encoded by three independent genes. AAC1, AAC2, and AAC3. In this work, we analysed the 5' upstream region of AAC1 by sequencing and by mapping the transcription initiation site of the gene. By monitoring the level of AAC1 mRNA and the beta-galactosidase activity of AAC1-lacZ fusion constructs, we showed that expression of AAC1 is subjected to regulation by oxygen. In contrast to the other two AAC genes, the effect of oxygen on AAC1 is not mediated by heme and heme-dependent transcription factors. The AAC1 expression was reduced eightfold in anaerobically grown cells compared to expression in cells grown aerobically, but it was not affected by the nature of carbon source used for growth. The data presented show that AAC1 expression, while constitutive under all aerobic conditions tested, is repressed during anaerobiosis in a heme-independent manner.
Assuntos
Regulação Fúngica da Expressão Gênica , Mitocôndrias/enzimologia , Translocases Mitocondriais de ADP e ATP/genética , Oxigênio/farmacologia , Saccharomyces cerevisiae/genética , Transcrição Gênica , Aerobiose , Anaerobiose , Sequência de Bases , Repressão Enzimática , Genes Fúngicos , Genes Reporter , Heme/farmacologia , Translocases Mitocondriais de ADP e ATP/biossíntese , Dados de Sequência Molecular , Mutação , RNA Fúngico/análise , RNA Fúngico/genética , RNA Mensageiro/análise , RNA Mensageiro/genética , Proteínas Recombinantes , Sequências Reguladoras de Ácido Nucleico , Saccharomyces cerevisiae/enzimologia , Análise de Sequência de DNA , Fatores de Transcrição/metabolismoAssuntos
Fator de Ligação a CCAAT , Carbono-Oxigênio Liases , Proteínas de Transporte/genética , DNA Liase (Sítios Apurínicos ou Apirimidínicos) , Regulação Fúngica da Expressão Gênica , Saccharomyces cerevisiae/genética , Difosfato de Adenosina/metabolismo , Trifosfato de Adenosina/metabolismo , Aerobiose , Ácido Aminolevulínico/metabolismo , Proteínas de Ligação a DNA/genética , Escherichia coli/genética , Proteínas Fúngicas/genética , Óperon Lac , Proteínas de Membrana/genética , Mitocôndrias/metabolismo , Proteínas Nucleares/genética , Recombinação Genética , Proteínas Repressoras/genética , Proteínas de Saccharomyces cerevisiae , Fatores de Transcrição/genética , Transcrição Gênica , beta-Galactosidase/metabolismoRESUMO
Expression of the Saccharomyces cerevisiae AAC2 gene encoding the major mitochondrial ADP/ATP carrier was examined. The intracellular level of the carrier protein, as well as the level of the AAC2-gene-specific mRNA, is influenced by the presence or absence of oxygen or of heme, and it is subject to carbon-source control. In addition, the expression of AAC2 gene requires the products of the HAP2 and HAP3 genes, but not that of the HAP1 gene. The 5'-flanking region of the gene was isolated, sequenced and fused to the lacZ reporter gene in order to study the effect of carbon sources and of specific deletion mutations on expression of the gene in yeast transformants. The expression of the reporter gene reveals that the AAC2 gene possesses a strong inducible promoter. The promoter analysis, combined with expression studies in the wild-type as well as in various mutant strains, identified an upstream activation site (UAS) contained within a sequence between -393 bp and -268 bp, and several major initiation sites of AAC2 mRNA between -105bp and -95 bp. Deletion analysis also shows that the TATA boxes located 45 bp and 104 bp upstream of the 5'-ends of AAC2 mRNA are not essential for the transcription. The UAS of the AAC2 gene is required for activation by HAP2 and heme and for release from glucose repressin. A restriction fragment containing the UAS conferred oxygen and carbon source regulation when placed upstream of another yeast gene encoding ADP/ATP carrier (AAC3), deleted of its regulatory sequences. The UAS of the AAC2 gene contains at least two distinct motifs for DNA-binding transcriptional activators, including one which is identical with the core HAP2/3/4 binding motif, and a second one with the ABF1 consensus binding sequence. Our results indicate that these sequences mediate the effects of the respective transactivator on the oxygen- and carbon-source-dependent transcription of the AAC2 gene.
Assuntos
Fator de Ligação a CCAAT , Proteínas Fúngicas/farmacologia , Regulação Fúngica da Expressão Gênica/efeitos dos fármacos , Heme/farmacologia , Translocases Mitocondriais de ADP e ATP/genética , Oxigênio/farmacologia , Saccharomyces cerevisiae/genética , Fatores de Transcrição/farmacologia , Sequência de Bases , Sequência Consenso , Proteínas Fúngicas/genética , Genes Fúngicos , Mitocôndrias/enzimologia , Translocases Mitocondriais de ADP e ATP/biossíntese , Dados de Sequência Molecular , Regiões Promotoras Genéticas/genética , RNA Mensageiro/biossíntese , Saccharomyces cerevisiae/metabolismo , Fatores de Transcrição/genética , Transcrição GênicaRESUMO
Saccharomyces cerevisiae strains expressing a single type of ADP/ATP carrier (AAC) protein were prepared from a mutant in which all AAC genes were disrupted, by transformation with plasmids containing a chosen AAC gene. As demonstrated by measurements of [14C]ADP specific binding and transport, all three translocator proteins, AAC1, AAC2 and AAC3 when present in the mitochondrial membrane, exhibited similar translocation properties. The disruption of some AAC genes, however, resulted in phenotypes indicating that the function of these proteins in whole cells can be quite different. Specifically, we found that the disruption of AAC1 gene, but not AAC2 and AAC3, resulted in a change in colony phenotype.
