Differential expression of growth hormone receptor messenger RNA from a second promoter.
Multiple alternatively spliced 5' untranslated regions (5'UTRs) have been identified in growth hormone (GH) receptor mRNA isolated from hepatic and adipocyte tissue. In the present study, the preferential utilisation of a GC-rich 5'UTR, designated exon 1B, was observed following the isolation of ovine (o) GH receptor cDNA clones from a skeletal muscle cDNA library. Although exon 1B-oGH receptor mRNA was expressed in all tissues examined, marked differences in the level of expression relative to the whole GH receptor transcript pool were observed between tissues. A single genomic clone (lambda 9) was isolated that encompassed exon 1B, together with 6 kilobase pairs of 5' and 12 kilobase pairs of 3' flanking sequence. Multiple transcription initiation sites were identified using RNase protection analysis on skeletal muscle poly(A)+ RNA, a result consistent with the absence of a proximal TATA box element. A CAAT box (-37 to -33) and a putative binding site for SP1 (a GC box -68 to -63) were found in the sense orientation immediately upstream of major transcription initiation site. Transfection of a series of overlapping promoter fragments linked to the luciferase reporter gene into HuH7, CHO and HeLa cells defined a core promoter element of 134 base pairs that was sufficient for maximum promoter activity. The emerging complexity of the 5' regulatory region of the GH receptor gene was emphasised by the observation that probes derived from exon 1B and the distal 3' intron boundary do not hybridise with previously cloned genomic sequences that span the liver-specific P1 promoter and exon 2.
Adams
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Centre for Animal Biotechnology, University of Melbourne, Parkville, Victoria, Australia.
D000273:Adipose Tissue / Q000737:chemistry; D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D016466:CHO Cells; D006528:Carcinoma, Hepatocellular; D006224:Cricetinae; D004247:DNA / Q000032:analysis / Q000737:chemistry / Q000235:genetics; D005091:Exons / Q000235:genetics; D018507:Gene Expression Regulation, Developmental; D006367:HeLa Cells; D006801:Humans; D008099:Liver / Q000737:chemistry; D008113:Liver Neoplasms; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000737:chemistry; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics*; D011986:Receptors, Somatotropin / Q000032:analysis / Q000235:genetics*; D012756:Sheep; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D012333:RNA, Messenger; D011986:Receptors, Somatotropin; D004247:DNA