RESUMO
SAMHD1 (sterile α motif and histidine (H) aspartate (D) domain-containing protein 1) is known for its antiviral activity of hydrolysing deoxynucleotides required for virus replication. Daddacha et al. identify a hydrolase-independent, moonlighting function of SAMHD1 that facilitates homologous recombination of DNA double-strand breaks (DSBs) by promoting recruitment of C-terminal binding protein interacting protein (CTIP), a DNA-end resection factor, to damaged DNA. These findings could benefit anticancer treatment.
Assuntos
Reparo do DNA/genética , DNA/genética , Proteína 1 com Domínio SAM e Domínio HD/genética , Oxirredutases do Álcool/genética , Quebras de DNA de Cadeia Dupla , Proteínas de Ligação a DNA/genética , Recombinação Homóloga/genética , Humanos , Proteínas Nucleares/genética , Ligação Proteica/genéticaRESUMO
Eukaryotic chromosomes reach their stable rod-shaped appearance in mitosis in a reaction dependent on the evolutionarily conserved condensin complex. Little is known about how and where condensin associates with chromosomes. Here, we analyze condensin binding to budding yeast chromosomes using high-resolution oligonucleotide tiling arrays. Condensin-binding sites coincide with those of the loading factor Scc2/4 of the related cohesin complex. The sites map to tRNA and other genes bound by the RNA polymerase III transcription factor TFIIIC, and ribosomal protein and SNR genes. An ectopic B-box element, recognized by TFIIIC, constitutes a minimal condensin-binding site, and TFIIIC and the Scc2/4 complex promote functional condensin association with chromosomes. A similar pattern of condensin binding is conserved along fission yeast chromosomes. This reveals that TFIIIC-binding sites, including tRNA genes, constitute a hitherto unknown chromosomal feature with important implications for chromosome architecture during both interphase and mitosis.
Assuntos
Adenosina Trifosfatases/fisiologia , Cromossomos Fúngicos/genética , Proteínas de Ligação a DNA/fisiologia , Mitose , Complexos Multiproteicos/fisiologia , Saccharomycetales/metabolismo , Fatores de Transcrição TFIII/fisiologia , Transcrição Gênica , Sítios de Ligação , Ciclo Celular , Proteínas de Ciclo Celular , Montagem e Desmontagem da Cromatina , Imunoprecipitação da Cromatina , Proteínas Cromossômicas não Histona , Proteínas Fúngicas/fisiologia , RNA Polimerase III/metabolismo , RNA de Transferência/genética , Saccharomycetales/citologia , Saccharomycetales/genética , CoesinasRESUMO
Sister chromatid cohesion is mediated by cohesin, but the process of cohesion establishment during S-phase is still enigmatic. In mammalian cells, cohesin binding to chromatin is dynamic in G1, but becomes stabilized during S-phase. Whether the regulation of cohesin stability is integral to the process of cohesion establishment is unknown. Here, we provide evidence that fission yeast cohesin also displays dynamic behavior. Cohesin association with G1 chromosomes requires continued activity of the cohesin loader Mis4/Ssl3, suggesting that repeated loading cycles maintain cohesin binding. Cohesin instability in G1 depends on wpl1, the fission yeast ortholog of mammalian Wapl, suggestive of a conserved mechanism that controls cohesin stability on chromosomes. wpl1 is nonessential, indicating that a change in wpl1-dependent cohesin dynamics is dispensable for cohesion establishment. Instead, we find that cohesin stability increases at the time of S-phase in a reaction that can be uncoupled from DNA replication. Hence, cohesin stabilization might be a pre-requisite for cohesion establishment rather than its consequence.
Assuntos
Proteínas de Ciclo Celular/metabolismo , Ciclo Celular/genética , Proteínas Cromossômicas não Histona/metabolismo , Cromossomos Fúngicos/metabolismo , Proteínas Nucleares/metabolismo , Schizosaccharomyces/genética , Schizosaccharomyces/metabolismo , Proteínas de Ciclo Celular/genética , Cromatina/metabolismo , Proteínas Cromossômicas não Histona/genética , Fase G1/genética , Fase G2/genética , Proteínas Nucleares/genética , Ligação Proteica , Fase S/genética , Schizosaccharomyces/citologia , Troca de Cromátide Irmã/genética , CoesinasRESUMO
Mice double deficient in LAMP-1 and -2 were generated. The embryos died between embryonic days 14.5 and 16.5. An accumulation of autophagic vacuoles was detected in many tissues including endothelial cells and Schwann cells. Fibroblast cell lines derived from the double-deficient embryos accumulated autophagic vacuoles and the autophagy protein LC3II after amino acid starvation. Lysosomal vesicles were larger and more peripherally distributed and showed a lower specific density in Percoll gradients in double deficient when compared with control cells. Lysosomal enzyme activities, cathepsin D processing and mannose-6-phosphate receptor expression levels were not affected by the deficiency of both LAMPs. Surprisingly, LAMP-1 and -2 deficiencies did not affect long-lived protein degradation rates, including proteolysis due to chaperone-mediated autophagy. The LAMP-1/2 double-deficient cells and, to a lesser extent, LAMP-2 single-deficient cells showed an accumulation of unesterified cholesterol in endo/lysosomal, rab7, and NPC1 positive compartments as well as reduced amounts of lipid droplets. The cholesterol accumulation in LAMP-1/2 double-deficient cells could be rescued by overexpression of murine LAMP-2a, but not by LAMP-1, highlighting the more prominent role of LAMP-2. Taken together these findings indicate partially overlapping functions for LAMP-1 and -2 in lysosome biogenesis, autophagy, and cholesterol homeostasis.