Differential gene expression in primary human skin keratinocytes and fibroblasts in response to ionizing radiation.
Although skin is usually exposed during human exposures to ionizing radiation, there have been no thorough examinations of the transcriptional response of skin fibroblasts and keratinocytes to radiation. The transcriptional response of quiescent primary fibroblasts and keratinocytes exposed to from 10 cGy to 5 Gy and collected 4 h after treatment was examined. RNA was isolated and examined by microarray analysis for changes in the levels of gene expression. Exposure to ionizing radiation altered the expression of 279 genes across both cell types. Changes in RNA expression could be arranged into three main categories: (1) changes in keratinocytes but not in fibroblasts, (2) changes in fibroblasts but not in keratinocytes, and (3) changes in both. All of these changes were primarily of p53 target genes. Similar radiation-induced changes were induced in immortalized fibroblasts or keratinocytes. In separate experiments, protein was collected and analyzed by Western blotting for expression of proteins observed in microarray experiments to be overexpressed at the mRNA level. Both Q-PCR and Western blot analysis experiments validated these transcription changes. Our results are consistent with changes in the expression of p53 target genes as indicating the magnitude of cell responses to ionizing radiation.
Warters
Raymond L
RL
Department of Radiation Oncology, University of Utah Health Sciences Center, Salt Lake City, Utah 84132, USA.
Packard
Ann T
AT
Kramer
Gwen F
GF
Gaffney
David K
DK
Moos
Philip J
PJ
D015153:Blotting, Western; D015816:Cell Adhesion Molecules, Neuronal / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000378:metabolism; D004307:Dose-Response Relationship, Radiation; D005347:Fibroblasts / Q000378:metabolism / Q000528:radiation effects*; D015870:Gene Expression / Q000528:radiation effects*; D016158:Genes, p53; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D015603:Keratinocytes / Q000378:metabolism / Q000528:radiation effects*; D009414:Nerve Growth Factors / Q000235:genetics / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011839:Radiation, Ionizing; D012867:Skin / Q000378:metabolism / Q000528:radiation effects*; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism; D000084622:Sestrins
D015816:Cell Adhesion Molecules, Neuronal; D050759:Cyclin-Dependent Kinase Inhibitor p21; D006360:Heat-Shock Proteins; D009414:Nerve Growth Factors; D012313:RNA; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; C493956:CDKN1A protein, human; C484438:NINJ1 protein, human; C117338:SESN1 protein, human; D000084622:Sestrins