Bilateral NSF/BIO-BBSRC- Remodelling Replication Roadblocks: Regulatory Systems that Integrate DNA Replication, Recombination and Protein Modification

双边 NSF/BIO-BBSRC-重塑复制障碍：整合 DNA 复制、重组和蛋白质修饰的监管系统

• 批准号: 1642283
• 负责人: Julie Maupin-Furlow
• 金额: $ 40.23万
• 依托单位: University of Florida
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-08-15 至 2020-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1642283&HistoricalAwards=false
• 关键词: Bilateral; NSF; BIO; BBSRC; Remodelling

In the absence of replication, most DNA lesions (damaged areas of DNA) are tolerated remarkably well by living cells. However, all organisms must replicate (copy) their genome in order to divide, and replication forks are prone to stalling at lesions. Attempting to replicate a DNA lesion will turn a small problem affecting one DNA strand to potentially damaging both DNA strands. Cells can disassemble and rebuild replication forks, which is accomplished by a type of protein degradation. Such remodeling is necessary before the stalled fork can be restarted. In this collaborative project investigators from the US (University of Florida and Duke University) and the UK (Nottingham University) will investigate the regulatory networks that connect the processes of DNA replication, recombination and protein degradation in Archaea. The project provides interdisciplinary training opportunities in quantitative experimental methods. The PIs will also participate in outreach activities to the general public to promote science education.Replication is prone to stalling at DNA lesions but to avoid reinitiating at origins, stalled forks are rebuilt by ubiquitin-mediated protein degradation. Such remodeling is necessary before the stalled fork can be restarted by homologous recombination. The PIs have uncovered a regulatory network centered on the CBS domain protein RcrA in Haloferax volcanii that connects the processes of DNA replication, recombination and ubiquitin-like modification. CBS domain proteins are used as sensors of intracellular metabolites suggesting a regulatory function for RcrA. To unravel this network, a systems biology approach will be used that incorporates transcriptomics, proteomics and two-hybrid analysis. Cells will be challenged by agents that block replication, loss-of-function mutations, and drugs that inhibit key regulatory enzymes. The results will be used to inform computational models, which will be refined using data from iterative rounds of transcriptomics and proteomics.This collaborative US/UK project is supported by the US National Science Foundation and the UK Biotechnology and Biological Sciences Research Council.

在没有复制的情况下，活细胞对大多数 DNA 损伤（DNA 受损区域）的耐受性非常好。然而，所有生物体都必须复制（复制）其基因组才能分裂，而复制叉很容易在病变处停滞。尝试复制 DNA 损伤会将影响一条 DNA 链的小问题变成可能损害两条 DNA 链的问题。细胞可以分解和重建复制叉，这是通过一种蛋白质降解来完成的。在停滞的分叉重新启动之前，这种重塑是必要的。在这个合作项目中，来自美国（佛罗里达大学和杜克大学）和英国（诺丁汉大学）的研究人员将研究古生菌中连接 DNA 复制、重组和蛋白质降解过程的调控网络。该项目提供定量实验方法的跨学科培训机会。 PI 还将参与面向公众的推广活动，以促进科学教育。复制很容易在 DNA 损伤处停滞，但为了避免在起点重新启动，停滞的分叉通过泛素介导的蛋白质降解来重建。 在通过同源重组重新启动停滞的分叉之前，这种重塑是必要的。 PI 发现了一个以 Haloferax volcanii 中 CBS 结构域蛋白 RcrA 为中心的调控网络，该网络连接 DNA 复制、重组和类泛素修饰过程。 CBS 结构域蛋白被用作细胞内代谢物的传感器，表明 RcrA 具有调节功能。为了解开这个网络，将使用一种系统生物学方法，结合转录组学、蛋白质组学和双杂交分析。细胞将受到阻碍复制、功能丧失突变和抑制关键调节酶的药物的挑战。结果将用于为计算模型提供信息，该模型将使用转录组学和蛋白质组学迭代的数据进行完善。这个美国/英国合作项目得到了美国国家科学基金会和英国生物技术和生物科学研究委员会的支持。

项目成果

Gene Expression of Haloferax volcanii on Intermediate and Abundant Sources of Fixed Nitrogen

中等充足固定氮源下火山盐鳅基因表达

• DOI: 10.3390/ijms20194784
• 发表时间: 2019-10
• 期刊: International Journal of Molecular Sciences
• 影响因子: 5.6
• 作者: Hwang, Sungmin;Chavarria, Nikita;Hackley, Rylee;Schmid, Amy;Maupin
• 通讯作者: Maupin

Molecular Factors of Hypochlorite Tolerance in the Hypersaline Archaeon Haloferax volcanii

高盐古菌 Haloferax volcanii 次氯酸盐耐受性的分子因素

• DOI: 10.3390/genes9110562
• 发表时间: 2018-11-20
• 期刊: Genes
• 影响因子: 3.5
• 作者: Gomez M;Leung W;Dantuluri S;Pillai A;Gani Z;Hwang S;McMillan LJ;Kiljunen S;Savilahti H;Maupin-Furlow JA
• 通讯作者: Maupin-Furlow JA

Multiplex quantitative SILAC for analysis of archaeal proteomes: a case study of oxidative stress responses: Multiplex SILAC for archaeal proteomics

用于古菌蛋白质组分析的多重定量 SILAC：氧化应激反应的案例研究：用于古菌蛋白质组学的多重 SILAC

• DOI: 10.1111/1462-2920.14014
• 发表时间: 2018-01
• 期刊: Environmental Microbiology
• 影响因子: 5.1
• 作者: McMillan LJ;Hwang S;Farah RE;Koh J;Chen S;Maupin-Furlow JA
• 通讯作者: Maupin-Furlow JA