Transcriptional regulation involving the intronic heat shock element of the rat hsp27 gene.
While sequencing the first intron of the rat heat shock protein 27 gene (hsp27), we identified a consensus heat shock regulatory element (HSE). This intronic HSE (i-HSE) is conserved among mammalian hsp27 genes. The aim of this study was to investigate possible effects of this intronic HSE (i-HSE) on transcription from the rat hsp27 promoter. Gel mobility shift assays indicated that the i-HSE bound heat shock transcription factor 1 (HSF1) in a manner equivalent to that of HSE present in hsp27 promoter (p-HSE). The effect of i-HSE on transcription from the hsp27 promoter was evaluated using reporter constructs transiently transfected in the osteosarcoma cell line ROS17/2.8. When inserted 5' to a 145 bp fragment of the hsp27 promoter not containing p-HSE, a 215 bp fragment of hsp27 intron 1 containing i-HSE enhanced CAT activity and conferred heat shock-inducible activity to the construct. This intronic fragment containing i-HSE also enhanced CAT activity in either normal or heat-shocked culture conditions when inserted 3' to the CAT open reading frame. However, in chimeric reporter constructs with a 273 bp hsp27 promoter containing p-HSE directly 5' to CAT reporter, and with a 215 bp fragment containing i-HSE inserted 3' to the CAT open reading frame, transcription from hsp27 promoter was reduced under normal and heat-stressed culture conditions. Mutation of the i-HSE reversed this effect. Further study is required to define the mechanism by which the i-HSE-containing region of the hsp27 promoter may mediate negative regulation of hsp27 transcription.
Cooper
L F
LF
Department of Biochemistry and Biophysics, School of Medicine, 308 Brauer Hall, CB #7450, University of North Carolina, Chapel Hill, NC 27599, USA. lyndon_cooper@dentistry.unc.edu
Uoshima
K
K
Guo
Z
Z
D000818:Animals; D002460:Cell Line; D017930:Genes, Reporter; D055551:HSP27 Heat-Shock Proteins; D006360:Heat-Shock Proteins / Q000235:genetics* / Q000378:metabolism; D007438:Introns; D009363:Neoplasm Proteins / Q000235:genetics*; D009687:Nuclear Proteins / Q000378:metabolism; D011401:Promoter Regions, Genetic; D051381:Rats; D015183:Restriction Mapping; D013696:Temperature; D014158:Transcription, Genetic; D014162:Transfection
D055551:HSP27 Heat-Shock Proteins; D006360:Heat-Shock Proteins; C486114:Hspb1 protein, rat; D009363:Neoplasm Proteins; D009687:Nuclear Proteins