Characterization of an intron splice enhancer that regulates alternative splicing of human GH pre-mRNA

Splicing of pre-mRNA transcripts is regulated by consensus sequences at intron (intervening sequence, IVS) boundaries and the branch site. In vitro studies have shown that the small introns of some genes also require intron splice enhancers (ISE) to modulate splice site selection. An autosomal dominant form of isolated GH deficiency (IGHD-II) is caused by mutations in IVS3 of the GH-1 gene that cause exon 3 (E3) skipping, resulting in truncated hGH products that prevent secretion of normal hGH. Interestingly, some of these IGHD-II mutations perturb an ISE that is buried in IVS3. We localized this ISE by quantitating the effects of deletions within IVS3 on E3 skipping. The importance of individual nucleotides to ISE function was determined by analyzing the effects of point mutants and additional deletions. Our results show that (i) an ISE with a G2X1-4G3motif resides in IVS3 of GH-1; (ii) both runs of Gs are required for ISE function; (iii) a single copy of the ISE regulates E3 skipping and (iv) ISE function can be modified by an adjacent AC element. Our findings reveal a new mechanism by which mutations can cause inherited human endocrine disorders and suggest that (i) ISEs may regulate splicing of transcripts of other genes and (ii) mutations of these ISEs or of the trans -acting factors that bind them may cause other genetic disorders.