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Advisory Committee Chair

Bradley Yoder

Advisory Committee Members

Kent Keyser

Michael Miller

Michael Crowley

Alecia Gross

Document Type

Dissertation

Date of Award

2015

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Nephronophthisis (NPHP) is a ciliopathy with diverse clinical features likely caused by genetic modifiers. To identify NPHP modifiers, a screen was conducted on nphp-4(tm925) mutant C. elegans to reveal mutations that exacerbate the NPHP ciliary defects. Ten loci were generated, five of which have now been identified. Three are mutations in ciliopathy genes mks-1, mks-2, and mks-5. The fourth allele (yhw66) is a missense mutation (S316F) in OSM-3, a kinesin required for cilia distal segment assembly. As in osm-3 null mutants, nphp-4(tm925);osm-3(yhw66) mutants lack distal segments, are dye-filing defective (Dyf), and have osmotic avoidance defects (OSM). The osm-3(yhw66) mutant alone has no overt cilia phenotype but do have reduced IFT rates. To evaluate the possibility S316F is a phosphorylation site, we compared the localization and function of phospho-null OSM-3(S316F), phospho-mimetic OSM-3(S316D), and wild type OSM-3. OSM-3(S316F) is enriched at the cilia base compared to wild type OSM-3. In contrast, OSM-3(S316D) accumulates in the cilia distal tip. While OSM-3(S316F) rescues all phenotypes in osm-3(mn357) nulls and OSM-3(S316D) restores distal cilia segments, OSM-3(S316D) does not rescue the Dyf phenotype with or without NPHP-4. Thus, there are NPHP-4 dependent and independent phenotypes associated with these OSM-3 mutations. These data reveal that NPHP-4 can exert regulatory influences on trafficking and function of a ciliary kinesin. The fifth allele from the screen was cca-1, a T-type calcium channel. The genetic interaction between cca-1 and nphp-4 is not critical for building the cilia but is necessary to maintain the cilium and affects its localization in an NPHP-4 dependent manner. Without this interaction calcium signaling and adaptation responses in the AWC neuron is impaired. Moreover, the data suggest that the human OSM-3 (Kif17) and CCA-1 (CACNA1I) homologs are candidate modifying locus affecting disease penetrance or expressivity in NPHP patients.

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