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Title: A novel mutation in SLC4A1 causing distal renal tubular acidosis : an investigation of the AE1 C-terminal domain
Author: Fry, A. C.
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2011
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This research identifies a novel AE1 mutation in a newly-referred family where dRTA segregates with disease through a four-generation pedigree, as determined by DNA sequencing and specific restriction enzyme digestion. This mutation – AE1-M909T – lies within the last four residues of the tail, in a potential PDZ-binding domain. AE1-M909T expression in Xenopus oocytes confirmed preserved anion exchange function. To investigate the mechanism of disease, N-terminal GFP-tagged AE1 was stably expressed in a variety of cell types in HEK293 and non-polarised MDCK cells, wild-type (AE1wt) and AE1-M909T both reach the cell surface. In polarised MDCK cells, GFP-tagged AE1wt has normal basolateral residency. In marked contrast, the mutant protein is aberrantly targeted, appearing at the apical membrane in addition to preserved basolateral presence. Antibody-labelling assays demonstrated post-synthetic AE1wt traffic direct to the basolateral membrane without any apical passage. In contrast, AE1-M909T traffics directly to both cell surfaces, thus implying gain of an apical targeting signal (plus persistent basolateral signals): this non-polarised expression will cause failure of α-IC function and dRTA. This novel SLC41 mutation causes AE1 mistargeting through acquisition of an apical targeting signal. The results suggest that basolateral targeting motifs are concentrated upstream of the extreme C-terminus of AE1, but that this micro-domain is important for normal biosynthetic traffic to the cell surface, potentially through PDZ-based interactions.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available