Molecular characterization of collaborator of ARF (CARF) as a DNA damage response and cell cycle checkpoint regulatory protein.
CARF is an ARF-binding protein that has been shown to regulate the p53-p21-HDM2 pathway. CARF overexpression was shown to cause growth arrest of human cancer cells and premature senescence of normal cells through activation of the p53 pathway. Because replicative senescence involves permanent withdrawal from the cell cycle in response to DNA damage response-mediated signaling, in the present study we investigated the relationship between CARF and the cell cycle and whether it is involved in the DNA damage response. We demonstrate that the half-life of CARF protein is less than 60 min, and that in cycling cells CARF levels are highest in G2 and early prophase. Serially passaged normal human skin and stromal fibroblasts showed upregulation of CARF during replicative senescence. Induction of G1 growth arrest and senescence by a variety of drugs was associated with increase in CARF expression at the transcriptional and translational level and was seen to correlate with increase in DNA damage response and checkpoint proteins, ATM, ATR, CHK1, CHK2, γH2AX, p53 and p21. Induction of growth arrest by oncogenic RAS and shRNA-mediated knockdown of TRF2 in cancer cells also caused upregulation of CARF. We conclude that CARF is associated with DNA damage response and checkpoint signaling pathways.
Cell Proliferation Research Group and DBT-AIST International Laboratory for Advanced Biomedicine, National Institute of Advanced Industrial Science and Technology (AIST), Central 4, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8562, Japan.
Rumani
R
Singh
Cell Proliferation Research Group and DBT-AIST International Laboratory for Advanced Biomedicine, National Institute of Advanced Industrial Science and Technology (AIST), Central 4, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8562, Japan.
Rajkumar S
RS
Kalra
Cell Proliferation Research Group and DBT-AIST International Laboratory for Advanced Biomedicine, National Institute of Advanced Industrial Science and Technology (AIST), Central 4, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8562, Japan.
Kamrul
K
Hasan
Children׳s Medical Research Institute, 214 Hawkesbury Road, Westmead, New South Wales 2145, Australia; Department of Molecular Virology, Immunology and Medical Genetics, 960 Biomedical Research Tower, The Ohio State University, Columbus, OH 43210, USA.
Zeenia
Z
Kaul
Cell Proliferation Research Group and DBT-AIST International Laboratory for Advanced Biomedicine, National Institute of Advanced Industrial Science and Technology (AIST), Central 4, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8562, Japan.
Caroline T
CT
Cheung
Children׳s Medical Research Institute, 214 Hawkesbury Road, Westmead, New South Wales 2145, Australia.
Lily
L
Huschtscha
Children׳s Medical Research Institute, 214 Hawkesbury Road, Westmead, New South Wales 2145, Australia.
Roger R
RR
Reddel
Cell Proliferation Research Group and DBT-AIST International Laboratory for Advanced Biomedicine, National Institute of Advanced Industrial Science and Technology (AIST), Central 4, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8562, Japan. Electronic address: s-kaul@aist.go.jp.
Sunil C
SC
Kaul
Cell Proliferation Research Group and DBT-AIST International Laboratory for Advanced Biomedicine, National Institute of Advanced Industrial Science and Technology (AIST), Central 4, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8562, Japan. Electronic address: renu-wadhwa@aist.go.jp.
Renu
R
Wadhwa
D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D059447:Cell Cycle Checkpoints / Q000502:physiology*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D016922:Cellular Senescence / Q000502:physiology; D004249:DNA Damage / Q000502:physiology*; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006801:Humans; D009369:Neoplasms / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D011401:Promoter Regions, Genetic / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012867:Skin / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D017154:Stromal Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D035341:Telomeric Repeat Binding Protein 2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
D000970:Antineoplastic Agents; D051017:Apoptosis Regulatory Proteins; C539004:CDKN2AIP protein, human; D018797:Cell Cycle Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; C504753:TERF2 protein, human; D035341:Telomeric Repeat Binding Protein 2
CARF; DNA damage response; Mechanism; Senescence; Upregulation