RPA70 depletion induces hSSB1/2-INTS3 complex to initiate ATR signaling.
The primary eukaryotic single-stranded DNA-binding protein, Replication protein A (RPA), binds to single-stranded DNA at the sites of DNA damage and recruits the apical checkpoint kinase, ATR via its partner protein, ATRIP. It has been demonstrated that absence of RPA incapacitates the ATR-mediated checkpoint response. We report that in the absence of RPA, human single-stranded DNA-binding protein 1 (hSSB1) and its partner protein INTS3 form sub-nuclear foci, associate with the ATR-ATRIP complex and recruit it to the sites of genomic stress. The ATRIP foci formed after RPA depletion are abrogated in the absence of INTS3, establishing that hSSB-INTS3 complex recruits the ATR-ATRIP checkpoint complex to the sites of genomic stress. Depletion of homologs hSSB1/2 and INTS3 in RPA-deficient cells attenuates Chk1 phosphorylation, indicating that the cells are debilitated in responding to stress. We have identified that TopBP1 and the Rad9-Rad1-Hus1 complex are essential for the alternate mode of ATR activation. In summation, we report that the single-stranded DNA-binding protein complex, hSSB1/2-INTS3 can recruit the checkpoint complex to initiate ATR signaling.
National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi-110067, India.
Ananya
A
Kar
National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi-110067, India.
Manpreet
M
Kaur
National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi-110067, India.
Tanushree
T
Ghosh
National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi-110067, India.
Md Muntaz
MM
Khan
National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi-110067, India.
Aparna
A
Sharma
National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi-110067, India.
Ritu
R
Shekhar
National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi-110067, India.
Akhil
A
Varshney
National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi-110067, India sandeep@nii.res.in.
Sandeep
S
Saxena
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D064007:Ataxia Telangiectasia Mutated Proteins / Q000378:metabolism; D002460:Cell Line; D000071877:Checkpoint Kinase 1; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D024101:Mitochondrial Proteins / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism; D034622:RNA Interference; D051716:Replication Protein A / Q000037:antagonists & inhibitors* / Q000235:genetics; D015398:Signal Transduction; D013312:Stress, Physiological / Q000235:genetics
C521194:ATRIP protein, human; D048868:Adaptor Proteins, Signal Transducing; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; C543505:INTS3 protein, human; D024101:Mitochondrial Proteins; C497167:RPA1 protein, human; D051716:Replication Protein A; C504091:NABP2 protein, human; D011494:Protein Kinases; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1