Involvement of DNA-dependent protein kinase in down-regulation of cell cycle progression.
The catalytic polypeptide of DNA-dependent protein kinase (p470) is encoded by the gene responsible for murine severe combined immunodeficiency (SCID) devoid of DNA double-strand break repair and V(D)J recombination. Here, we have characterized the role of p470 in cell proliferation using SCID mice and the cell lines. In accord with DNA histogram patterns, SCID cell lines (SD/SD-eA and SC3VA2) expressing extremely low level of DNA-PK activity grew faster than a normal mouse cell line (CB/CB-eB) and SC3VA2 complemented with human p470 gene (RD13B2). In regenerating liver after partial hepatectomy, de novo DNA synthesis determined by [(3)H]thymidine incorporation started at 30h in C.B-17/Icr-SCID (SCID) mice and at around 36h in C.B-17/Icr (C.B-17) mice. Compared with normal cells, SCID cells contained slightly higher levels of transcripts of cyclin A, cyclin E, B-Myb and dihydrofolate reductase, which are regulated by E2F-1. E2F-1 playing a key role in G1- to S-phase progression was phosphorylated in vitro by DNA-PK. Importantly, the E2F-1 promoter transcriptional activity in SCID cell lines (SD/SD-eA and SC3VA2) was 4-5-fold higher than that in CB/CB-eB and RD13B2. These results suggest that p470 is involved in down-regulation of cell cycle progression through E2F-1-responsible genes.
Watanabe
Fumiaki
F
Department of Pathological Biochemistry, Medical Research Institute, Tokyo Medical and Dental University, 2-3-10 Kandasurugadai, Chiyoda-ku, 101-0062, Tokyo, Japan.
Shinohara
Ken-ichi
K
Teraoka
Hirobumi
H
Komatsu
Kenshi
K
Tatsumi
Kouichi
K
Suzuki
Fumio
F
Imai
Takashi
T
Sagara
Masashi
M
Tsuji
Hideo
H
Ogiu
Toshiaki
T
D000818:Animals; D034961:Antigens, Nuclear / Q000378:metabolism; D002453:Cell Cycle / Q000502:physiology*; D018797:Cell Cycle Proteins; D002460:Cell Line; D004247:DNA / Q000096:biosynthesis; D004265:DNA Helicases; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins / Q000378:metabolism; D015536:Down-Regulation / Q000502:physiology*; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; D004622:Embryo, Mammalian; D005347:Fibroblasts; D006801:Humans; D000072200:Ku Autoantigen; D008099:Liver; D008297:Male; D051379:Mice; D016513:Mice, SCID; D009687:Nuclear Proteins; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism
D034961:Antigens, Nuclear; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; C493754:E2f1 protein, mouse; D009687:Nuclear Proteins; D014157:Transcription Factors; D004247:DNA; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D017346:Protein Serine-Threonine Kinases; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; C000606510:Xrcc6 protein, mouse; D000072200:Ku Autoantigen