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KMID : 0604220060130020043
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Korean Journal Investigative Dermatology
2006 Volume.13 No. 2 p.43 ~ p.52
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Analysis of TPA-inducible Genes in Keratinocytes using cDNA Microarray
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Kim Yeong-Ho
Kang Dong-Wook
Park Mi-Ja
Seo Eun-Young
Kim Bo-Joong
Kim Chang-Duk
Lee Jeung-Hoon
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Abstract
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The tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) is a strong inducer of differentiation in keratinocyte. This effect of TPA is thought to be due to the activation of PKC pathway. To identify the genes relating to TPA induced tumor promotion and differentiation in keratinocytes, we adopted suppression subtractive hybridization (SSH) and cDNA microarray. Messenger RNAs were isolated from the primary skin keratinocytes cultured in vitro after calcium, and then SSH was performed. A total of about 3,000 clones were obtained, and the sequencing and BLAST searching analyses revealed 840 independent cDNA clones, which were used for making the microarray slides. Time-course cDNA microarray analysis (3, 6, 12, 24, and 48 hours after TPA treatment) revealed the pictures of global gene expression profile in keratinocytes after TPA treatment. Of 840 genes tested, 336 genes showed the changes in their expression level, at least in one occasion over five time points. The genes were clustered into six groups according to their expression pattern using Self-Organizing Map analysis, and showed the global feature of function-related regulation. The genes related to keratinocyte differentiation were markedly up-regulated by TPA treatment, and a similar pattern of increase was seen in the expression of genes related to ribosomal protein. On the other hand, transcripts involved in metabolism, DNA repair, transcription, and translation were generally down-regulated. These results demonstrate the complexity of the gene expression profile that contributes to the spatiotemporal regulation of keratinocyte differentiation. cDNA microarray analysis in the present study allowed efficient identifying and validating of the genes thought to play some roles in tumor promotion and differentiation in keratinocytes.
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KEYWORD
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Keratinocyte, Differentiation, TPA, Calcium, Microarray
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