New SHOX mutations in patients with idiopathic short stature

Mutations of SHOX (Short Stature Homeobox) gene are associated with the short stature in Turner syndrome, Leri-Weill dyschondrosteosis and in some patients (2 to 15%) with idiopathic short stature (ISS). Nevertheless, recently it has been characterized the SHOX gene enhancer, whose deletions has been shown to be responsible for the Leri-Weill syndrome.We report the results of a study carried out on patients with ISS, investigated for the presence of SHOX gene and SHOX enhancer mutations.Fifty-three patients of both sexes (2-18 years) entered this study. The inclusion criteria were: 1) height <3° centile; 2) normal stimulated GH values 3) absence of obvious skeletal anomalies 4) exclusion of chronic disorders causing short stature.All 53 samples were examined for the presence of deletions or duplications within the SHOX gene and the PAR1 region by Multiplex Ligation-dependent Probe Amplification (MLPA), using the SHOX salsa P018B kit (MRC-Holland); they were also examined for the presence of point mutations and small deletions and insertions by direct sequencing of all the coding exons and the intron/exon boundaries of the gene isoform A (from exon 2 to exon 6a) and of its enhancer.Sequencing analysis revealed no point mutations and small deletions or insertions that could account for the phenotype. On the contrary the molecular analysis of the entire PAR1 region by MLPA surprisingly revealed two large duplications. One patient presented a duplication of about 500Kb extending from exon 1 of the SHOX gene and encompassing its enhancer; another patient presented a smaller duplication involving only the enhancer. Both the duplications were further confirmed by using specific panels of microsatellites markers that resulted in triallelic patterns.For all we know, these mutations had never been described before and they could have a role in the short stature of these patients. In fact, in both cases, the duplication of PAR1 could disrupt the normal cis/regulation of transcription caused by the closeness of the duplicated regions. Such proximity could impede the regular interaction of each enhancer to each appropriate promoter, resulting in the alteration, most likely a reduction, of the normal transcriptional activity. This could be the reason why the duplication involving also a regulative element such as the enhancer is associated to a short stature phenotype while an extra copy of only the SHOX gene seems to determine tall stature.