Characterizaton of human growth hormone receptor (hGHR) gene expression in human adipocytes

Human growth hormone (hGH) is a key regulator of postnatal growth and metabolism, exerting its effects through a specific cell-surface receptor, hGHR. Eight hGHR mRNAs are produced by the hGHR gene due to transcribing from different 5' non-coding exons. Four (V2,V3,V5,V9) are ubiquitously expressed, whereas the others (V1,V4,V7,V8) are present only in normal postnatal liver, suggesting that different promoter usage is a mechanism for developmental and tissue-specific regulation of the hGHR gene.hGH influences adipose tissue development and function. By 5'RACE screening of a human adipocyte cDNA library, I determined that the V2 transcript is the predominant (∼90%) hGHR mRNA expressed in fat cells. Although five novel mRNA variants (VA-VE) were isolated, they were demonstrated to be widely expressed minor hGHR transcripts, instead of adipocyte-specific. Using the human SGBS preadipocyte cell line, I showed that total hGHR mRNAs increase significantly during adipocyte differentiation, primarily due to increases in the V2 transcript, reaching a maximum with maturity.The predominance of the V2 variant in all cells examined to date led us to carry out an in-depth study of the V2 promoter. Two major transcriptional start sites were identified that were identical for SGBS preadipocytes, liver and kidney. Transient transfection analysis of the 5' flanking region of V2 confirmed its promoter activity in multiple human cell lines. Similar transcriptional profiles were observed in SGBS pre- and mature adipocytes, with much higher V2 promoter activity seen in mature adipocytes, suggesting that increases in adipocyte factor(s) play a role in V2 promoter regulation. The V2 proximal promoter is TATA-less, with several characteristics of a housekeeping gene promoter. Cis-elements for Ets1, CHOP and Hes1 were found in the proximal promoter and within the V2 exon. They exert either positive (Ets1, CHOP) or negative (Hes1) regulatory effects on V2 transcription when their respective transcription factors are over-expressed, via direct binding (Ets1, Hes1) or interaction with other proteins (CHOP). These sites likely regulate hGHR expression in response to extra-cellular (developmental, growth, stress) signals.Thus, transcriptional regulation of hGHR V2 in human cells, including adipocytes, is the result of a complex interplay by multiple factors, to ensure sufficient local production of hGHR throughout life