Endocrine Abstracts

Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant disorder characterised by the occurrence of parathyroid, pancreatic and pituitary tumours, and is due to mutations of the MEN1 gene, which encodes menin. We have investigated identical twins with MEN1, one of whom developed primary hyperparathyroidism (PHPT) and a prolactinoma that caused pubertal arrest, and the other had PHPT only. DNA sequence analysis of the MEN1 coding region had not identified any abnormalities and we hypothesised that deletions and mutations involving the untranslated regions may be involved, and we investigated these using patients’ leukocyte DNA and combined Sanger DNA sequence and multiplex ligation-dependent probe amplification (MLPA) analysis. This revealed a heterozygous 596bp deletion between nucleotides −1,088 and −493 upstream of the translation start site, located within the MEN1 5′ untranslated region (UTR), and includes the core promoter and multiple cis-regulatory regions. To investigate the effects of this 5′UTR deletion on MEN1 promoter activity, we generated luciferase reporter constructs, containing a wild-type 842bp or mutant 246bp MEN1 promoter, and transfected them into human insulinoma BON-1 cells and human embryonic kidney HEK293 cells. This revealed the mutation to result in significant reductions by 17-fold (P<0.0001) and 30-fold (P<0.0001) in luciferase expression in BON-1 and HEK293 cells, respectively, when compared to the wild-type. The effects of this 5′UTR deletion on MEN1 transcription and translation were assessed using qRT-PCR and Western blot analysis, respectively, of mRNA and protein lysates obtained from Epstein-Barr-virus transformed lymphoblastoid cells derived from the affected twins and unrelated controls (N=4). This demonstrated the 5′UTR deletion to result in significant reductions of 82% (P<0.05) and 88% (P<0.05) in MEN1 mRNA and menin protein, respectively. Thus, our results report the first germline MEN1 5′UTR mutation and highlight the importance of investigating UTRs in MEN1 patients who do not have coding region mutations.