Heat-induced phosphorylation of NBS1 in human skin fibroblast cells.
NBS1 is known to be involved in DNA damage-induced cellular responses after exposure to ionizing radiation (IR). Phosphorylation of NBS1 contributes to cell-cycle checkpoints. The aim of this study was to determine whether heat exposure induces or stimulates cellular responses mediated by the phosphorylation of NBS1 in human skin fibroblast cell lines. The results of immunofluorescent staining and Western blot analysis showed that NBS1 proteins are phosphorylated after exposure to heat in the nucleus of a normal skin fibroblast cell line (82-6 cells). This suggests that the NBS1-mediated signal transduction could be induced by heat. We further examined whether a deficiency in the NBS1 protein modifies heat sensitivity in human skin fibroblast cell lines. A skin fibroblast cell line (Gmtert), derived from a Nijmegen breakage syndrome (NBS) patient containing mutant NBS1, showed higher sensitivity to heat than the same cell line transfected with the wild-type copy of the NBS1 gene. We also showed that transfection of a DNA cassette expressing small interference RNA (siRNA) targeted to NBS1 into 82-6 cells enhanced cell sensitivity to heat. These results suggest that NBS1 is involved in cellular responses to DNA damage which is induced by heat exposure as well as by radiation exposure in human skin fibroblast cells.
Ohnishi
Ken
K
Department of Biology, Nara Medical University School of Medicine, 840 Shijo-cho, Kashihara, Nara 634-8521, Japan.
Scuric
Zorica
Z
Yau
David
D
Schiestl
Robert H
RH
Okamoto
Noritomo
N
Takahashi
Akihisa
A
Ohnishi
Takeo
T
D001483:Base Sequence; D015153:Blotting, Western; D018797:Cell Cycle Proteins / Q000378:metabolism*; D017931:DNA Primers; D005347:Fibroblasts / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D006358:Hot Temperature; D006801:Humans; D009687:Nuclear Proteins / Q000378:metabolism*; D010766:Phosphorylation; D021381:Protein Transport; D012867:Skin / Q000166:cytology / Q000378:metabolism*
D018797:Cell Cycle Proteins; D017931:DNA Primers; C112156:NBN protein, human; D009687:Nuclear Proteins