Gregory, LC; (2017) Investigation of new candidate genes in a cohort of patients with familial congenital hypopituitarism and associated disorders. Doctoral thesis , UCL (University College London).

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Abstract

Congenital hypopituitarism is a complex variable genetic disorder that is known to be caused by multiple mutated genes, both in isolation and in variably penetrant cases of digenic inheritance. In only <10% of cases, a mutation in a known causative gene has been identified in the patient, leaving the vast majority of patients yet to have a genetic mutation detected that is responsible for the pathogenicity and that has functional significance to their condition. This study investigates novel genes and pathways involved in hypothalamo-pituitary development. Our large cohort of consanguineous and non-consanguineous pedigrees with hypothalamo-pituitary disease are routinely screened for variants in the known causative genes. In pedigrees where there are no variants in these particular genes, exome sequencing in collaboration with GOSgene is carried out to uncover novel genes and regions of interest that are abnormal in the individual. Upon the identification of any novel variant in known or novel genes, functional assays are conducted to further show the significance of the change. Firstly this study identifies the first novel homozygous mutation in the LHX4 gene, p.T126M, in two deceased brothers from a pedigree with combined pituitary hormone deficiency with subsequent fatal consequences. Functional luciferase assays showed that there was no significant difference between mutant p.T126M and WT constructs in transactivating the αGSU and prolactin promoters, and that mutant LHX4 could synergise with POU1F1 similar to WT LHX4. Secondly, six new candidate genes; CTPS2, RNPC3, PRMT6, FASN, APEX2 and EIF2S3, were identified in phenotypically unique pedigrees submitted to GOSgene for exome sequencing. The human embryonic expression profiles of these novel candidate genes were analysed in this study in a hypothalamo-pituitary context, as well as in related tissues that are affected in the individuals. Thirdly, the role of eIF2γ, encoded by EIF2S3, which was found to be mutated in an X-linked pedigree with congenital hypopituitarism, hypothyroidism and hyperinsulinism causing hypoglycaemia, was investigated in this study. A lentiviral shRNA knock out of the EIF2S3 gene in human pancreatic cells resulted in significantly higher apoptosis compared to WT cells. This study has used both a Sanger sequencing and an exome sequencing approach to identify novel variants in known and novel candidate genes respectively.

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