title: Separate cis-acting DNA elements control cell type- and tissue-specific expression of collagen binding molecular chaperone HSP47.
abstract: HSP47 is a collagen-binding heat shock protein and is assumed to act as a molecular chaperone in the biosynthesis and secretion of procollagen. As the synthesis of HSP47 is closely correlated with that of collagen in various cell lines and tissues, we performed a promoter/reporter assay using HSP47-producing and nonproducing cells. 280 base pairs (bp(s)) of upstream promoter were shown to be necessary for the basal expression but not to be enough for the cell type-specific expression. When the first and the second introns were introduced downstream of this 280-bp region, marked up-regulation of the reporter activity was observed in HSP47-producing cells but not in nonproducing cells. This was confirmed in transgenic mice by staining the lacZ gene product under the control of the 280-bp upstream promoter and the introns. Staining was observed in skin, chondrocytes, precursor of bone, and other HSP47/collagen-producing tissues. A putative Sp1-binding site at -210 bp in the promoter, to which Sp3 and an unidentified protein bind, was shown to be responsible for this up-regulation when combined with the introns. However no difference in the binding to this probe was observed between HSP47-producing and nonproducing cells. The responsible region for cell type-specific up-regulation was found to be located in a 500-bp segment in the first intron. On electrophoresis mobility shift assay using this 500-bp probe, specific DNA-protein complexes were only observed in HSP47-producing cell extracts. These results suggest that two separate elements are necessary for the cell type-specific expression of the hsp47 gene; one is a putative Sp1-binding site at -210 bp necessary for basal expression, and the other is a 500-bp region within the first intron, required for cell type-specific expression.
authors.affiliation: Department of Molecular and Cellular Biology, Institute for Frontier Medical Sciences, Kyoto University, Kyoto 606-8507, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Hirata
authors.forename: I
authors.initials: I
authors.lastname: Yamamura
authors.forename: K
authors.initials: K
authors.lastname: Yasuda
authors.forename: A
authors.initials: A
authors.lastname: Kobayashi
authors.forename: N
authors.initials: N
authors.lastname: Tada
authors.forename: M
authors.initials: M
authors.lastname: Suzuki
authors.forename: K
authors.initials: K
authors.lastname: Hirayoshi
authors.forename: N
authors.initials: N
authors.lastname: Hosokawa
authors.forename: K
authors.initials: K
authors.lastname: Nagata
mesh_terms: D016475:3T3 Cells; D000818:Animals; D003094:Collagen / Q000378:metabolism*; D005314:Embryonic and Fetal Development; D005786:Gene Expression Regulation; D018507:Gene Expression Regulation, Developmental; D015698:Genomic Library; D050958:HSP47 Heat-Shock Proteins; D006367:HeLa Cells; D006360:Heat-Shock Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007438:Introns; D008156:Luciferases / Q000032:analysis / Q000235:genetics; D051379:Mice; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D009928:Organ Specificity; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D015183:Restriction Mapping; D014162:Transfection; D014407:Tumor Cells, Cultured; D001616:beta-Galactosidase / Q000235:genetics
chemical_list: D050958:HSP47 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D011993:Recombinant Fusion Proteins; C494660:SERPINH1 protein, human; C494661:Serpinh1 protein, mouse; D003094:Collagen; D008156:Luciferases; D001616:beta-Galactosidase