Activation of Tel1ATM kinase requires Rad50 ATPase and long nucleosome-free DNA but no DNA ends

Sarem Hailemariam,Sandeep Kumar,Peter M Burgers

doi:10.1074/jbc.ra119.008410

Sarem Hailemariam, Sandeep Kumar + Show 1 more

Open Access

https://doi.org/10.1074/jbc.ra119.008410

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Abstract

In Saccharomyces cerevisiae, Tel1 protein kinase, the ortholog of human ataxia telangiectasia-mutated (ATM), is activated in response to DNA double-strand breaks. Biochemical studies with human ATM and genetic studies in yeast suggest that recruitment and activation of Tel1ATM depends on the heterotrimeric MRXMRN complex, composed of Mre11, Rad50, and Xrs2 (human Nbs1). However, the mechanism of activation of Tel1 by MRX remains unclear, as does the role of effector DNA. Here we demonstrate that dsDNA and MRX activate Tel1 synergistically. Although minimal activation was observed with 80-mer duplex DNA, the optimal effector for Tel1 activation is long, nucleosome-free DNA. However, there is no requirement for DNA double-stranded termini. The ATPase activity of Rad50 is critical for activation. In addition to DNA and Rad50, either Mre11 or Xrs2, but not both, is also required. Each of the three MRX subunits shows a physical association with Tel1. Our study provides a model of how the individual subunits of MRX and DNA regulate Tel1 kinase activity.

Highlights

In Saccharomyces cerevisiae, Tel1 protein kinase, the ortholog of human ataxia telangiectasia–mutated (ATM), is activated in response to DNA double-strand breaks
Eukaryotes respond to Double-strand breaks (DSBs) by promptly initiating a multipronged DNA damage response, which involves the initiation of a cell cycle checkpoint and DNA repair
Cryo-EM studies of human ATM and S. pombe Tel1 show that these orthologs are homodimeric in structure [10, 39]

Summary

Edited by Patrick Sung

In Saccharomyces cerevisiae, Tel protein kinase, the ortholog of human ataxia telangiectasia–mutated (ATM), is activated in response to DNA double-strand breaks. The yeast Saccharomyces cerevisiae Tel and Mec protein kinases are orthologs of mammalian ataxia telangiectasia-mutated (ATM) and ATM- and Rad3-related (ATR), respectively They are essential regulators of cell cycle checkpoint initiation in yeast [3, 4]. MRX has several enzymatic and scaffolding properties and plays major roles in DNA break metabolism It is a crucial player in all aspects of DSB processing: damage detection, cell cycle checkpoint initiation, and facilitating and catalyzing repair of the lesion [2, 21, 22]. Extensive genetic studies of S. cerevisiae Tel and the MRX complex have given us a comprehensive understanding of where and how these factors function in various DNA metabolic pathways and which activities and/or domains are essential in each of these pathways [2, 21]. This stimulatory effect of Rad is absolutely dependent on its ATPase activity

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Discussion

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