Document Type

Dissertation

Date of Degree

Spring 2014

Degree Name

PhD (Doctor of Philosophy)

Degree In

Genetics

First Advisor

Val C. Sheffield

Abstract

Bardet-Biedl syndrome (BBS) is a pleiotropic and genetically heterogeneous disorder, and a well-known ciliopathy. Nineteen different genes have been reported for BBS, mutations in which cause characteristic phenotypes including retinal degeneration, obesity, polydactyly, renal abnormalities, hypogenitalism and cognitive impairment. Protein products of eleven BBS genes are part of two major complexes: the BBSome complex and a CCT/CTRiC/BBS complex. The CCT/CTRiC/BBS complex assists in the formation of the BBSome complex, which in turn traffics numerous receptor proteins to the cilia. However, the precise mechanism by which BBSome ciliary trafficking activity is regulated is not fully understood. In fact, a complete picture of the cellular functions of BBS proteins is still missing, and gaps remain in our understanding of the pleiotropy and heterogeneity of the disease. With the aim of bridging those gaps, this thesis project was designed to identify tissue specific cargoes of the BBSome and to characterize their BBS-related functions. To this end, we generated a transgenic LAP-BBS4 mouse, which expresses the transgene in various tissues including brain, eye, testis, heart, kidney, and adipose tissue. We found that despite tissue specific variable expression, LAP-BBS4 was able to complement the deficiency of Bbs4 and rescue all the BBS phenotypes in the Bbs4 null mice. The finding provides an encouraging prospective for gene therapy for BBS related phenotypes and potentially for other ciliopathies. We also utilized the transgenic mice to search for tissue specific BBSome cargo proteins and identified CEP131 as a novel BBSome interacting protein. Using in vitro cell culture models we show that CEP131 interacts with the BBSome through BBS4. CEP131 is not involved in BBSome assembly, but accumulation of the BBSome in cilia is enhanced upon CEP131 depletion. Our in vitro data implicate CEP131 as a negative regulator of ciliary BBSome trafficking. Finally, we show that cep131 knockdown in zebrafish embryos results in typical BBS phenotypes including Kupffer's vesicle abnormalities and melanosome transport delay. This finding confirms the association of CEP131 with the BBS pathway. Overall, the work performed for this thesis provides further insight into the regulation of BBSome ciliary trafficking and suggests CEP131 as a BBS candidate gene.

Keywords

Bardet-Biedl Syndrome, BBS4, BBSome, CEP131, Ciliopathy, primary cilia

Pages

xiii, 116 pages

Bibliography

Includes bibliographical references (pages 100-116).

Copyright

Copyright 2014 Xitiz Chamling

Included in

Genetics Commons

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