Evidence for hSNM1B/Apollo functioning in the HSP70 mediated DNA damage response.
The hSNM1B/Apollo protein is involved in the cellular response to DNA-damage as well as in the maintenance of telomeres during S-phase. TRF2 has been shown to interact physically with hSNM1B. As a core component of shelterin, TRF2 functions in organization and protection of telomeres. However, TRF2 was also shown to have a role in the early DNA-damage response, suggesting that hSNM1B and TRF2 cooperate in this dual function. Here we have used Tandem-Affinity-Purification in combination with mass spectrometry to identify additional binding partners of hSNM1B. This revealed HSC70, HSP72, HSP60 and beta-Tubulin to be hSNM1B-interactors. We have confirmed the interaction of hSNM1B and HSP70 in co-immunoprecipitation assays and found that hSNM1B binds to a C-terminal fragment of HSP72, known to contain the substrate binding domain. Depletion of HSP72 in human fibroblasts resulted in a significant reduction of nuclear hSNM1B foci. We also found the phosphorylation of CHK1 at serine 317 to be attenuated in response to UVC irradiation as a consequence of hSNM1B depletion, a result which extends our previous findings on the DNA-damage response function of hSNM1B. HSP70 chaperones have been implicated in the maintenance of genome stability and their expression is often aberrant in cancer. Our results presented here, suggest that the role in genome stability might not be specific to HSP70 but rather can be attributed, at least in part, to hSNM1B. This, together with its stimulating effect on ATM and ATR substrate phosphorylation in response to DNA-damage qualify hSNM1B as a putative target in cancer therapy.
Anders
Marco
M
Institut für Humangenetik, Charité-Universitätsmedizin Berlin, Berlin, Germany.
Mattow
Jens
J
Digweed
Martin
M
Demuth
Ilja
I
D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000378:metabolism; D002460:Cell Line; D000071877:Checkpoint Kinase 1; D004249:DNA Damage; D045643:DNA Repair Enzymes / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism; D005090:Exodeoxyribonucleases; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D050884:HSP72 Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D009687:Nuclear Proteins / Q000378:metabolism*; D011485:Protein Binding; D011494:Protein Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D016196:S Phase; D014404:Tubulin / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000378:metabolism; D014466:Ultraviolet Rays
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D018840:HSP70 Heat-Shock Proteins; D050884:HSP72 Heat-Shock Proteins; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D014404:Tubulin; D025521:Tumor Suppressor Proteins; D011494:Protein Kinases; C576324:ATM protein, human; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D017346:Protein Serine-Threonine Kinases; C493741:DCLRE1B protein, human; D005090:Exodeoxyribonucleases; D045643:DNA Repair Enzymes