IDENTIFICATION OF SMARCAL1 AS A COMPONENT OF THE DNA DAMAGE RESPONSE

鉴定 SMARCAL1 作为 DNA 损伤反应的一个组成部分

• 批准号: 8169194
• 负责人: Hironori Funabiki
• 金额: $ 0.58万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-03-01 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8169194
• 关键词: ATP phosphohydrolase; Binding; Binding Proteins; Caffeine; Cell physiology; Cells; Computer Retrieval of Information on Scientific Projects Database; DNA; DNA Binding; DNA Damage; DNA Double Strand Break; Disease; Dysplasia; Frequencies; Functional disorder; Funding; Generations; Grant; Growth; Human; Institution; Kidney; Mass Spectrum Analysis; Mutation; Phosphotransferases; Protein Family; Proteins; Publishing; Recruitment Activity; Research; Research Personnel; Resources; Single-Stranded DNA; Source; T-Cell Immunodeficiency; United States National Institutes of Health; Work; Xenopus; egg; homologous recombination; novel; recombinational repair; response

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. SMARCAL1 (also known as HARP) is a SWI/SNF family protein with an ATPase activity stimulated by DNA containing both single-stranded and double-stranded regions. Mutations in SMARCAL1 are associated with the disease Schimke immuno-osseous dysplasia, a multisystem autosomal recessive disorder characterized by T cell immunodeficiency, growth inhibition, and renal dysfunction. The cellular function of SMARCAL1, however, is unknown. Here, using Xenopus egg extracts and mass spectrometry, we identify SMARCAL1 as a protein recruited to double-stranded DNA breaks. SMARCAL1 binds to double-stranded breaks and stalled replication forks in both egg extract and human cells, specifically colocalizing with the single-stranded DNA binding factor RPA. In addition, SMARCAL1 interacts physically with RPA independently of DNA. SMARCAL1 is phosphorylated in a caffeine-sensitive manner in response to double-stranded breaks and stalled replication forks. It has been suggested that stalled forks can be stabilized by a mechanism involving caffeine-sensitive kinases, or they collapse and subsequently recruit Rad51 to promote homologous recombination repair. We show that depletion of SMARCAL1 from U2OS cells leads to increased frequency of RAD51 foci upon generation of stalled replication forks, indicating that fork breakdown is more prevalent in the absence of SMARCAL1. We propose that SMARCAL1 is a novel DNA damage-binding protein involved in replication fork stabilization. This work ha been published: (305. L. Postow, E.M. Woo, B.T. Chait, H. Funabiki "Identification of SMARCAL1 as a component of the DNA damage response" J Biol Chem. 284(2009) 35951-61 )

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目和 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 SMARCAL1（也称为 HARP）是一种 SWI/SNF 家族蛋白，其 ATP 酶活性受包含单链和双链区域的 DNA 刺激。 SMARCAL1 突变与 Schimke 免疫性骨发育不良疾病相关，这是一种多系统常染色体隐性遗传疾病，其特征为 T 细胞免疫缺陷、生长抑制和肾功能障碍。然而，SMARCAL1 的细胞功能尚不清楚。在这里，使用非洲爪蟾卵提取物和质谱分析，我们将 SMARCAL1 鉴定为一种招募到双链 DNA 断裂的蛋白质。 SMARCAL1 与鸡蛋提取物和人类细胞中的双链断裂和停滞的复制叉结合，特别是与单链 DNA 结合因子 RPA 共定位。此外，SMARCAL1 与 RPA 的物理相互作用不依赖于 DNA。 SMARCAL1 以咖啡因敏感的方式磷酸化，以响应双链断裂和复制叉停滞。有人建议，停滞的分叉可以通过涉及咖啡因敏感激酶的机制来稳定，或者它们崩溃并随后招募 Rad51 来促进同源重组修复。我们发现，U2OS 细胞中 SMARCAL1 的耗尽会导致在产生停滞的复制叉时 RAD51 焦点的频率增加，这表明在缺乏 SMARCAL1 的情况下，复制叉断裂更为普遍。我们认为 SMARCAL1 是一种参与复制叉稳定的新型 DNA 损伤结合蛋白。 该工作已发表：（305. L. Postow, E.M. Woo, B.T. Chait, H. Funabiki “Identification of SMARCAL1 as a component of the DNA Damage response” J Biol Chem. 284(2009) 35951-61）