IDENTIFICATION OF CIS-ELEMENTS AND TRANSCRIPTION FACTORS THAT REGULATE THE TISSUE SPECIFIC EXPRESSION OF HSP47

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HSP47 is a collagen-binding heat shock protein assumed to act as molecular chaperone in the biosynthesis and secretion of procollagen. Synthesis of HSP47 is closely correlated with those of various types of collagens in various cell types and tissues. Hirata et. al reported that 280 base pairs (bp(s)) of upstream promoter were necessary for the basal expression and 500 bps within the first intron, called BS region, were necessary for the cell type-specific expression using promoter analysis and transgenic mice. And more, we identified the EPm region (480bps) within the second intron, which was correlated with cell type-specific expression of HSP47, like BS region. To narrow down the responsive element in both BS and EPm region, we performed EMSA assay using nuclear extracts of BALB3T3 cells expressing both HSP47 and collagen. Finally, we identified two 30 bps regions, BS-5B and EP7-D, which were localized within BS and EPm region, respectively. The shift pattern in EMSA assay using the 32-P labeled BS5-B was similar to that of EP7-D. In addition, the shift bands of the 32P-labeled BS5-B disappeared in the presence of cold EP7-D DNA as a competitor, and vice versa. Thus, it is suggested that the same factor(s) bind to both BS5-B and EP7-D regions. To investigate the transcription activity of BS5-B region, we performed promoter analysis using deletion and mutation constructs of BS5-B. Luciferase activities by these constructs were shown to be reduced to the similar extent of the deletion construct of whole BS region. It is suggested that BS5-B is a responsible cis-element responsible for the cell type-specific expression of HSP47 in the collaboration with 280 bps of upstream promoter. We have identified the transcription factor binding to the BS5-B and EP7-D regions adopting the one-hybrid assay using these BS5-B and EP7-D regions as baits. W are now characterizing these factors in the transcriptional regulation of HSP47..
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IDENTIFICATION OF CIS-ELEMENTS AND TRANSCRIPTION FACTORS THAT REGULATE THE TISSUE SPECIFIC EXPRESSION OF HSP47

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