RESUMEN
Mammalian Sirtuin proteins (SIRTs) are homologs of yeast Sir2, and characterized as class III histone deacetylases of NAD(+) dependence. Unlike their lower counterparts that are directly involved in the extending of lifespan, mammalian SIRTs mainly function in metabolism and cellular homeostasis, among them, SIRT7 is the least understood. SIRT7 is localized in the nucleus and rich in nucleoli associated with RNA polymerase I, and correlated with cell proliferation. In contrast, SIRT7 has recently been demonstrated to specifically deacetylate H3K18ac in the chromatin, and in most cases represses proliferation. Although MicroRNA as miR-125b has been reported to down-regulate SIRT7 by binding to its 3'UTR, however, how SIRT7 gene is regulated remains unclear. Here, we identified the transcription initiation site of human SIRT7 gene at the upstream 23rd A nucleotide respective to the translational codon, and the SIRT7 is a TATA-less and initiator-less gene. The sequences in the upstream region between -256 and -129 bp are identical with important functions in the three species detected. A C/EBPα responding element is found that binds both C/EBPα and C/EBPß in vitro. We showed TSA induced SIRT7 gene transcription and only the HDAC3, but not its catalytic domain depleted mutant, interacted with C/EBPα to occupy the C/EBPα element and repressed SIRT7 gene in the hepatocellular carcinoma cells. To our knowledge, this is the first report on the regulation mechanism of SIRT7 gene, in which, HDAC3 collaborated with C/EBPα to occupy its responding element in the upstream region of SIRT7 gene and repressed its expression in human cells.
Asunto(s)
Proteínas Potenciadoras de Unión a CCAAT/genética , Carcinoma Hepatocelular/genética , Histona Desacetilasas/genética , Neoplasias Hepáticas/genética , Sirtuinas/genética , Regiones no Traducidas 3' , Animales , Proteína beta Potenciadora de Unión a CCAAT/genética , Proteínas Potenciadoras de Unión a CCAAT/biosíntesis , Carcinoma Hepatocelular/patología , Línea Celular Tumoral , Cromatina/genética , Histona Desacetilasas/biosíntesis , Humanos , Neoplasias Hepáticas/patología , Regiones Promotoras Genéticas , Sirtuinas/biosíntesisRESUMEN
Growth associated protein 43 (Gap43) is a neuron-specific phosphoprotein, which plays critical role in axon growth and synapses functions during neurogenesis. Here we identified two transcription start sites (TSSs) of the mouse Gap43 gene designated as a proximal site at +1, and a distal TSS at -414. RT-qPCR data reveal that the transcripts from +1 increase 10-fold on day-1 post-all-trans retinoic acid (RA) treatment, reached a peak value at day-4 and gradually reduced. By contrast, the distal TSS directs a late, remarkably sharp increase of the transcripts from the day-5 on. An intense signal of Gap43 at the neurites and neural network is determined by the efficient transcription of the distal promoter as shown in Northern blot and RT-qPCR assay. In addition, the targeting of p300 in combination with a differential enrichment of Brm to Brg1 change at the distal promoter region of the gene is induced under RA treatment. The over hundreds of GA rich stretches and the GAGAG elements located between the two TSSs may take parts in the differential transcription of the two TSSs of the Gap43. Our findings provide the first evidence on the identification and differential transcription of the two TSSs of the mouse Gap43 gene, and the preferential distribution of their protein products in the specific stages of RA induced P19 differentiation. These data suggest the efficient transcription of the distal promoter of Gap43 is an important mark for the transition of P19 cells from the progenitor stage into neuronal differentiation.