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The N-terminus region of Drp1, a Rint1 family protein is essential for cell survival and its interaction with Rad50 protein in fission yeast S.pombe.
Biochimica et Biophysica Acta (BBA) - General Subjects ( IF 3 ) Pub Date : 2020-09-19 , DOI: 10.1016/j.bbagen.2020.129739
Sachin Gaurav 1 , Rajeev Ranjan 1 , Jitendra Kuldeep 1 , Kanika Dhiman 1 , Pinaki Prasad Mahapatra 1 , Ashish 1 , Mohammad Imran Siddiqi 1 , Shakil Ahmed 1
Affiliation  

Background

Defects in DNA repair pathway can lead to double-strand breaks leading to genomic instability. Earlier we have shown that S.pombe Drp1, a Rint1/Tip1 family protein is required for the recovery from DNA damage.

Methods

Various truncations of Drp1 protein were constructed and their role in DNA damage response and interaction with Rad50 protein has been studied by co-immunoprecipitation and pull-down assays.

Results

The structural and functional analysis of Drp1 protein revealed that the N-terminus region of Drp1 is indispensable for the survival. The C-terminus truncation mutants, drp1C1Δ and drp1C2Δ exhibit temperature sensitive phenotype and are hypersensitive against DNA damaging agents with elevated level of Rad52-YFP foci at non-permissive temperature indicating the impairment for DNA damage repair pathway. The essential N-terminus region of Drp1 interacts with the C-terminus region of Rad50 and might be involved in influencing the MRN/X function. Small-angle X-ray (SAXS) analysis revealed three-domain like shapes in Drp1 protein while the C-terminus region of Rad50 exhibit unusual bulges. Computational docking studies revealed the amino acid residues at the C-terminus region of Rad50 that are involved in the interaction with the residues present at the N-terminal region of Drp1 indicating the importance of the N-terminal region of Drp1 protein.

Conclusions

We have identified the region of Drp1 and Rad50 proteins that are involved in the interaction and their role in the DNA damage response pathway has been analyzed.

General significance

The functional and structural aspects of fission yeast Drp1 protein and its interaction with Rad50 have been elucidated.



中文翻译:

Drp1的N端区域是Rint1家族蛋白,对于细胞存活及其与裂殖酵母S.pombe中的Rad50蛋白相互作用至关重要。

背景

DNA修复途径的缺陷可能导致双链断裂,从而导致基因组不稳定。早些时候,我们已经表明,粟酒裂殖酵母DRP1,一个Rint1 /提示1家族蛋白需要从DNA损伤的恢复。

方法

构建了多种截短的Drp1蛋白,并通过免疫共沉淀和下拉法研究了它们在DNA损伤应答中和与Rad50蛋白相互作用中的作用。

结果

Drp1蛋白的结构和功能分析表明,Drp1的N端区域对于生存是必不可少的。C端截短突变体drp1C1Δdrp1C2Δ表现出对温度敏感的表型,并且对DNA损伤剂高度敏感,在非允许温度下Rad52-YFP病灶水平升高,表明DNA损伤修复途径受损。Drp1的基本N末端区域与Rad50的C末端区域相互作用,可能参与影响MRN / X功能。小角X射线(SAXS)分析揭示了Drp1蛋白中的三个域类似的形状,而Rad50的C端区域表现出不寻常的凸起。计算对接研究揭示了Rad50 C末端区域的氨基酸残基,该残基与存在于Drp1 N末端区域的残基相互作用,表明Drp1蛋白N末端区域的重要性。

结论

我们已经确定了参与相互作用的Drp1和Rad50蛋白区域,并分析了它们在DNA损伤反应途径中的作用。

一般意义

裂变酵母Drp1蛋白的功能和结构方面及其与Rad50的相互作用已得到阐明。

更新日期:2020-09-23
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