RESUMO
DNA replication results in the doubling of the genome prior to cell division. This process requires the assembly of 50 or more protein factors into a replication fork. Here, we review recent structural and biochemical insights that start to explain how specific proteins recognize DNA replication origins, load the replicative helicase on DNA, unwind DNA, synthesize new DNA strands, and reassemble chromatin. We focus on the minichromosome maintenance (MCM2-7) proteins, which form the core of the eukaryotic replication fork, as this complex undergoes major structural rearrangements in order to engage with DNA, regulate its DNA-unwinding activity, and maintain genome stability.
Assuntos
Replicação do DNA/fisiologia , Animais , Cromatina/metabolismo , DNA Helicases/metabolismo , Replicação do DNA/genética , Evolução Molecular , Instabilidade Genômica/genética , Humanos , Proteínas de Manutenção de Minicromossomo/genética , Proteínas de Manutenção de Minicromossomo/metabolismo , Origem de Replicação/fisiologiaRESUMO
RNA polymerase III (RNAPIII) synthesizes most small RNAs, the most prominent being tRNAs. Although the basic mechanism of RNAPIII transcription is well understood, recent evidence suggests that additional proteins play a role in RNAPIII transcription. Here, we discovered by a genome-wide approach that Nab2, a poly(A)-binding protein important for correct poly(A) tail length and nuclear mRNA export, is present at all RNAPIII transcribed genes. The occupancy of Nab2 at RNAPIII transcribed genes is dependent on transcription. Using a novel temperature-sensitive allele of NAB2, nab2-34, we show that Nab2 is required for the occupancy of RNAPIII and TFIIIB at target genes. Furthermore, Nab2 interacts with RNAPIII, TFIIIB, and RNAPIII transcripts. Importantly, impairment of Nab2 function causes an RNAPIII transcription defect in vivo and in vitro. Taken together, we establish Nab2, an important mRNA biogenesis factor, as a novel player required for RNAPIII transcription by stabilizing TFIIIB and RNAPIII at promoters.
Assuntos
Regulação Fúngica da Expressão Gênica , Proteínas de Transporte Nucleocitoplasmático/metabolismo , RNA Polimerase III/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Transporte Nucleocitoplasmático/genética , Regiões Promotoras Genéticas , Ligação Proteica , RNA Polimerase III/genética , Proteínas de Ligação a RNA/genética , Proteínas de Saccharomyces cerevisiae/genéticaRESUMO
Mutations in the l(3)mbt tumour suppressor result in overproliferation of Drosophila larval brains. Recently, the derepression of different gene classes in l(3)mbt mutants was shown to be causal for transformation. However, the molecular mechanisms of dL(3)mbt-mediated gene repression are not understood. Here, we identify LINT, the major dL(3)mbt complex of Drosophila. LINT has three core subunits-dL(3)mbt, dCoREST, and dLint-1-and is expressed in cell lines, embryos, and larval brain. Using genome-wide ChIP-Seq analysis, we show that dLint-1 binds close to the TSS of tumour-relevant target genes. Depletion of the LINT core subunits results in derepression of these genes. By contrast, histone deacetylase, histone methylase, and histone demethylase activities are not required to maintain repression. Our results support a direct role of LINT in the repression of brain tumour-relevant target genes by restricting promoter access.
Assuntos
Neoplasias Encefálicas/genética , Proteínas de Drosophila , Drosophila melanogaster/genética , Complexos Multiproteicos , Proteínas Repressoras , Animais , Animais Geneticamente Modificados , Sítios de Ligação , Linhagem Celular , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Regulação da Expressão Gênica , Genoma de Inseto , Células Germinativas/metabolismo , Histonas/genética , Histonas/metabolismo , Larva/genética , Larva/metabolismo , Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Mutação , Cromossomos Politênicos/genética , Interferência de RNA , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismoRESUMO
RNA polymerase III (RNAPIII) synthesizes diverse, small, non-coding RNAs with many important roles in the cellular metabolism. One of the open questions of RNAPIII transcription is whether and how additional factors are involved. Recently, Nab2 was identified as the first messenger ribonucleoprotein particle (mRNP) biogenesis factor with a function in RNAPIII transcription.