The long-term goal of this project is to understand how basal bodies are assembled. Basal bodies are complex, microtubule-based structures that template cilia and recruit proteins needed for ciliary assembly. Cilia are found on almost every cell type of the human body and numerous human diseases are associated with ciliary defects including: respiratory infections, male infertility, situs inversus, polycystic kidney disease, retinal degeneration, liver dysplasia, and Bardet-Biedl Syndrome. Understanding the assembly of basal bodies will be invaluable in the elucidation of cilia formation and function and understanding how cilia are involved in human disease. This project focuses on the protein Stp2 and how it is involved in basal body and cilia assembly. Knockdown of Stp2 by RNA interference results in cells that have short cilia, increased numbers of cytoplasmic microtubules and an absent distal striated fiber that connects the basal bodies to each other. In addition to being required for basal body and cilia function, Stp2 may also play a role in the nucleolus. Immunofluorescence studies show that Stp2 localizes to both the basal bodies and nucleolus. A nucleolar function for Stp2 is further supported by evidence that nucleolar structure is altered in Stp2 RNA interference strains. With the goal of understanding how Stp2 functions at the basal bodies and nucleolus, this proposal seeks 1) to address how Stp2 associates with the basal bodies and nucleolus by using a combination of drug studies, real-time fluorescence microscopy and truncation analysis, and fluorescence recovery after photobleaching techniques, 2) to generate mutant alleles of stp2 by insertional, X-ray and chemical mutagenesis, and 3) to address if Stp2 has phospholipase activity as suggested by sequence analysis using a biochemical purification of Stp2 and in vitro phospholipase assays.
Cilia are finger-like projections on cells that serve motility and sensory functions. They are found on almost every cell of the human body and numerous human diseases are associated with defects in cilia including respiratory infections, male infertility, situs inversus, polycystic kidney disease, retinal degeneration, liver dysplasia, and Bardet- Biedl Syndrome. The goal of this project is to understand how cilia form and in turn how they are related to human disease.
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|Albee, Alison J; Kwan, Alan L; Lin, Huawen et al. (2013) Identification of cilia genes that affect cell-cycle progression using whole-genome transcriptome analysis in Chlamydomonas reinhardtti. G3 (Bethesda) 3:979-91|
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