title: Regulation of NEIL1 protein abundance by RAD9 is important for efficient base excision repair.
abstract: RAD9 participates in DNA damage-induced cell cycle checkpoints and DNA repair. As a member of the RAD9-HUS1-RAD1 (9-1-1) complex, it can sense DNA damage and recruit ATR to damage sites. RAD9 binding can enhance activities of members of different DNA repair pathways, including NEIL1 DNA glycosylase, which initiates base excision repair (BER) by removing damaged DNA bases. Moreover, RAD9 can act independently of 9-1-1 as a gene-specific transcription factor. Herein, we show that mouse Rad9(-/-) relative to Rad9(+/+) embryonic stem (ES) cells have reduced levels of Neil1 protein. Also, human prostate cancer cells, DU145 and PC-3, knocked down for RAD9 demonstrate reduced NEIL1 abundance relative to controls. We found that Rad9 is required for Neil1 protein stability in mouse ES cells, whereas it regulates NEIL1 transcription in the human cells. RAD9 depletion enhances sensitivity to UV, gamma rays and menadione, but ectopic expression of RAD9 or NEIL1 restores resistance. Glycosylase/apurinic lyase activity was reduced in Rad9(-/-) mouse ES and RAD9 knocked-down human prostate cancer whole cell extracts, relative to controls. Neil1 or Rad9 addition restored this incision activity. Thus, we demonstrate that RAD9 regulates BER by controlling NEIL1 protein levels, albeit by different mechanisms in human prostate cancer versus mouse ES cells.
authors: Center for Radiological Research, College of Physicians and Surgeons, Columbia University Medical Center, New York, NY 10032, USA.
authors: Sunil K
authors: SK
authors: Panigrahi
authors: Center for Radiological Research, College of Physicians and Surgeons, Columbia University Medical Center, New York, NY 10032, USA.
authors: Kevin M
authors: KM
authors: Hopkins
authors: Center for Radiological Research, College of Physicians and Surgeons, Columbia University Medical Center, New York, NY 10032, USA Department of Environmental Health Sciences, Mailman School of Public Health, Columbia University Medical Center, New York, NY 10032, USA HBL1@columbia.edu.
authors: Howard B
authors: HB
authors: Lieberman
mesh_terms: D000818:Animals; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D045744:Cell Line, Tumor; D045647:DNA Glycosylases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D004260:DNA Repair; D053595:Embryonic Stem Cells / Q000378:metabolism; D008297:Male; D051379:Mice; D011401:Promoter Regions, Genetic; D011471:Prostatic Neoplasms / Q000235:genetics / Q000378:metabolism; D054730:Protein Interaction Domains and Motifs; D012333:RNA, Messenger / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; D012333:RNA, Messenger; C069930:rad9 protein; D045647:DNA Glycosylases; C473634:NEIL1 protein, human; C481347:Neil1 protein, mouse