Cilia and centrioles are evolutionarily conserved eukaryotic organelles. Cilia sense the environment through a growing list of ciliary membrane localized receptors and proteins and/or move cells or fluids. Centrioles are also microtubule-based organelles that perform several functions. A centriole is matured into a basal body, which functions as a platform for templating/anchoring cilia to the membrane. Centrioles organize both cytoplasmic and spindle microtubules. During maturation of centrioles, a distal structure known as the transition zone is assembled. It consists of morphologically defined structures known as transition fibers, the Y- linkers, and the membrane-embedded ciliary necklace. Little is known about how the ciliary necklace is assembled onto the Y-linkers, or how the transition zone serves as a barrier between the cytoplasm and cilia. Many of the human ciliopathies affect proteins in the transition zone. Our newly discovered cnc1 mutant lacks the ciliary necklace, but has transition fibers and the Y- linkers. The cnc1 mutant provides a unique tool to dissect how proteins are assembled into the ciliary necklace, the identify of these ciliary necklace proteins and the role of this structure. Mutations in the Nek1 kinase cause retinal degeneration and abnormal bone growth in patients. Our analysis of the Nek1 homolog in Chlamydomonas suggests that it affects the transition zone, IFT regulation, and spindle microtubules. Analysis of its targets in the basal body and flagella as well as interacting proteins will help to dissect the role of this conserved kinase in ciliary and microtubule biology. Using electron tomography, our recent finding that the duplication of new centrioles initiates at a particular triplet microtubuleon the old centriole is really exciting, and we would like to know what molecules mark this site. Centrioles are built of triplet microtubules, but few proteins that affect triplet microtubule assembly are known. To address these and other questions, an unbiased forward genetic screen for new basal body duplication defective mutants together with whole genome sequencing and light and electron microscopy will identify the roles of new proteins in these two important events. In addition, the mutants from the genetic screen will be used to ask if there are assembly factors that are needed for centriole duplication, but are not structural components of centrioles.

Public Health Relevance

Basal bodies are cylindrical structures that serve as templates for building cilia, which are hair- like structures that move fluids and sense environmental signals. People with defects in basal bodies and cilia show a wide range of symptoms that include microcephaly, neural tube closure, blindness, and kidney disease. Basal bodies also serve as organizers of the spindle and cleavage furrow for cell division. This research will provide new information about the roles of basal body proteins and how their loss is pathogenic.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM032843-32
Application #
9432510
Study Section
Nuclear and Cytoplasmic Structure/Function and Dynamics Study Section (NCSD)
Program Officer
Gindhart, Joseph G
Project Start
1983-12-01
Project End
2019-02-28
Budget Start
2018-03-01
Budget End
2019-02-28
Support Year
32
Fiscal Year
2018
Total Cost
Indirect Cost
Name
Washington University
Department
Genetics
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
Dutcher, Susan K; O'Toole, Eileen T (2016) The basal bodies of Chlamydomonas reinhardtii. Cilia 5:18
Xu, Gang; Wilson, Kate S; Okamoto, Ruth J et al. (2016) Flexural Rigidity and Shear Stiffness of Flagella Estimated from Induced Bends and Counterbends. Biophys J 110:2759-68
Wilson, Kate S; Gonzalez, Olivia; Dutcher, Susan K et al. (2015) Dynein-deficient flagella respond to increased viscosity with contrasting changes in power and recovery strokes. Cytoskeleton (Hoboken) 72:477-90
Lin, Huawen; Dutcher, Susan K (2015) Genetic and genomic approaches to identify genes involved in flagellar assembly in Chlamydomonas reinhardtii. Methods Cell Biol 127:349-86
Cao, Muqing; Ning, Jue; Hernandez-Lara, Carmen I et al. (2015) Uni-directional ciliary membrane protein trafficking by a cytoplasmic retrograde IFT motor and ciliary ectosome shedding. Elife 4:
Mittelmeier, Telsa M; Thompson, Mark D; Lamb, Mary Rose et al. (2015) MLT1 links cytoskeletal asymmetry to organelle placement in chlamydomonas. Cytoskeleton (Hoboken) 72:113-23
Lin, Huawen; Zhang, Zhengyan; Guo, Suyang et al. (2015) A NIMA-Related Kinase Suppresses the Flagellar Instability Associated with the Loss of Multiple Axonemal Structures. PLoS Genet 11:e1005508
Viswanadha, Rasagnya; Hunter, Emily L; Yamamoto, Ryosuke et al. (2014) The ciliary inner dynein arm, I1 dynein, is assembled in the cytoplasm and transported by IFT before axonemal docking. Cytoskeleton (Hoboken) 71:573-86
O'Toole, Eileen T; Dutcher, Susan K (2014) Site-specific basal body duplication in Chlamydomonas. Cytoskeleton (Hoboken) 71:108-18
Dutcher, Susan K (2014) The awesome power of dikaryons for studying flagella and basal bodies in Chlamydomonas reinhardtii. Cytoskeleton (Hoboken) 71:79-94

Showing the most recent 10 out of 56 publications