Characterisation of a functional intronic polymorphism in the human growth hormone (GH1) gene

Abstract

The +1169A allele of the A/T single nucleotide polymorphism (SNP; rs2665802), located within intron 4 of the human growth hormone 1 ( GH1 ) gene, has been associated with reduced levels of circulating GH and insulin-like growth factor 1, a reduced risk of colorectal cancer and a predisposition to osteoporosis. Whether this intronic SNP is itself the functional polymorphism responsible for exerting a direct effect on GH1 gene expression, however, or whether it is instead in linkage disequilibrium with the functional SNP, has been an open question. The evolutionary conservation of the +1169T allele (and the surrounding intronic sequence) in the bovine genome, as well as in primate genomes, is, however, suggestive of its functionality. Although a potential alternative splice site spans the location of the +1169 SNP, polymerase chain reaction-based assays failed to yield any evidence for alternative splicing associated with either allele. To determine whether the +1169 SNP, in different allelic combinations with SNPs at -278 (G/T), -57 (T/G) and +2103 (C/T), exerts a direct effect on gene expression and/or GH secretion, we performed a series of transfections of various GH1 haplotype-expressing constructs into rat GC (somatotroph) cells. The results obtained provided evidence to support the contention that the +1169A allele contributes directly to the observed reduction in both GH1 gene expression and GH secretion. Part of the apparent influence of the +1169A-bearing allele on GH1 gene expression and GH secretion may still, however, be attributable to alleles of additional SNPs in cis to +1169A and located within either the promoter or the 3'-flanking region.

Figure 1

Intron 4 sequence from the orthologous GH genes of human, five other primates and Bos taurus (cow). The polymorphic nucleotide at +1169 in the human GH1 gene is marked in green. Mismatched positions are highlighted in yellow. GenBank accession nos: Homo sapiens J03071; Pan troglodytes, chimpanzee (AF374232.1); Gorilla gorilla, gorilla (FP245407.5); Nomascus leucogenys, white-cheeked gibbon (AY621637.1); Pygathrix roxellana, Sichuan-snub-nosed monkey (AY621647.1); Macaca mulatta, rhesus macaque (DQ002799.1); Bos taurus, cow (Bos taurus release Btau_4.0). The location of the consensus sequence of the p53-responsive element (RRRCWWGYYY) is shown in red [16].