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Title: Characterisation of transmembrane protein 114 (TMEM114), a protein associated with juvenile onset cataract.
Author: Maher, Geoffrey
ISNI:       0000 0004 2707 5811
Awarding Body: University of Manchester
Current Institution: University of Manchester
Date of Award: 2011
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Transmembrane protein 114 (TMEM114) is an uncharacterised predicted transmembrane protein expressed in the lens epithelium. A balanced translocation that transects the putative promoter of TMEM114 is associated with autosomal dominant congenital cataract (ADCC), however, coding sequence variants in TMEM114 were not identified in a panel of ADCC patients. Subsequent to the identification of TMEM114 a similar novel transmembrane protein named TMEM114-like protein 1 (TMLP1) was identified. This study aimed to functionally characterise the two novel proteins TMEM114 and TMLP1.TMEM114 and TMLP1 showed homology to voltage dependent calcium channel gamma (γ) subunits, but TMEM114 and TMLP1 lacked some of the key domains present in these proteins. Expression of TMEM114 and TMLP1 in the developing human eye was identified. TMLP1 was also expressed in developing neural tissue. To aid functional characterisation, the murine orthologues of Tmem114 and Tmlp1 were cloned and polyclonal antibodies were generated. Bioinformatic tools predicted co-and post-translational modifications. The predicted plasma membrane localisation of Tmem114 was confirmed in vitro in polarised MDCK II cells and the membrane localisation was shown to be dependent on the presence of N-linked oligosaccharides. Murine Tmlp1 was localised in the endoplasmic reticulum (ER) in MDCK II cells, possibly due to the fact that murine Tmlp1 lacks an N-glycosylation site present in other species.The knockdown of Tmem114 in Xenopus tropicalis using antisense morpholinos results in microphthalmia (small eye) confirming the protein's role in eye development and growth. Sequencing of patients with microphthalmia and anophthalmia identified a novel heterozygous missense mutation (p.R2Q) not present in controls. The p.R2Q variant was not found to affect the expression or localisation of the protein. The p.A147V variant, previously reported as a SNP, mis-localises to the endoplasmic reticulum.In conclusion, this study identifies that TMEM114 is a transmembrane glycoprotein with an important role in ocular development in vertebrates, although its precise functional role remains to be elucidated. Knockdown of Tmem114 in X. tropicalis results in microphthalmia, suggesting that loss of function mutations in TMEM114 may be associated with human ocular disease. Further characterisation of TMLP1 is required to identify its role in human biology.
Supervisor: Manson, Forbes Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Oral
EThOS ID:  DOI: Not available