title: Deficiency in the catalytic subunit of DNA-dependent protein kinase causes down-regulation of ATM.
abstract: Previous reports have suggested a connection between reduced levels of the catalytic subunit of DNA-dependent protein kinases (DNA-PKcs), a component of the nonhomologous DNA double-strand breaks end-joining system, and a reduction in ATM. We studied this possible connection in other DNA-PKcs-deficient cell types, and following knockdown of DNA-PKcs with small interfering RNA, Chinese hamster ovary V3 cells, lacking DNA-PKcs, had reduced levels of ATM and hSMG-1, but both were restored after transfection with PRKDC. Atm levels were also reduced in murine scid cells. Reduction of ATM in a human glioma cell line lacking DNA-PKcs was accompanied by defective signaling through downstream substrates, post-irradiation. A large reduction of DNA-PKcs was achieved in normal human fibroblasts after transfection with two DNA-PKcs small interfering RNA sequences. This was accompanied by a reduction in ATM. These data were confirmed using immunocytochemical detection of the proteins. Within hours after transfection, a decline in PRKDC mRNA was seen, followed by a more gradual decline in DNA-PKcs protein beginning 1 day after transfection. No change in ATM mRNA was observed for 2 days post-transfection. Only after the DNA-PKcs reduction occurred was a reduction in ATM mRNA observed, beginning 2 days post-transfection. The amount of ATM began to decline, starting about 3 days post-treatment, then it declined to levels comparable to DNA-PKcs. Both proteins returned to normal levels at later times. These data illustrate a potentially important cross-regulation between the nonhomologous end-joining system for rejoining of DNA double-strand breaks and the ATM-dependent damage response network of pathways, both of which operate to maintain the integrity of the genome.
authors.affiliation: Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, Colorado 80523, USA.
authors.forename: Yuanlin
authors.initials: Y
authors.lastname: Peng
authors.forename: Rick G
authors.initials: RG
authors.lastname: Woods
authors.forename: Heather
authors.initials: H
authors.lastname: Beamish
authors.forename: Ruiqiong
authors.initials: R
authors.lastname: Ye
authors.forename: Susan P
authors.initials: SP
authors.lastname: Lees-Miller
authors.forename: Martin F
authors.initials: MF
authors.lastname: Lavin
authors.forename: Joel S
authors.initials: JS
authors.lastname: Bedford
mesh_terms: D000818:Animals; D000954:Antigens, Surface; D001260:Ataxia Telangiectasia / Q000378:metabolism; D064007:Ataxia Telangiectasia Mutated Proteins; D016466:CHO Cells; D020134:Catalytic Domain; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D006224:Cricetinae; D004247:DNA / Q000235:genetics* / Q000378:metabolism / Q000528:radiation effects; D004249:DNA Damage / Q000235:genetics; D004260:DNA Repair / Q000235:genetics; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D015536:Down-Regulation; D005347:Fibroblasts / Q000378:metabolism / Q000473:pathology; D005786:Gene Expression Regulation; D005910:Glioma / Q000378:metabolism / Q000473:pathology; D006801:Humans; D007124:Immunoenzyme Techniques; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D021122:Protein Subunits; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D014162:Transfection; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D000954:Antigens, Surface; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D008565:Membrane Proteins; D009687:Nuclear Proteins; D021122:Protein Subunits; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C099014:SPAG8 protein, human; D025521:Tumor Suppressor Proteins; D004247:DNA; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D017346:Protein Serine-Threonine Kinases