Abstract: SA-PO483
Evidence for Genetic Compensation for Cilia Membrane Delivery Defects in cep290/NPHP6 Mutants
Session Information
- Cystic Kidney Diseases: Basic/Translational
November 09, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Genetic Diseases of the Kidneys
- 1001 Genetic Diseases of the Kidneys: Cystic
Authors
- Cardenas-Rodriguez, Magdalena, Massachusetts General Hospital, Charlestown, Massachusetts, United States
- Drummond, Iain A., Massachusetts General Hospital, Charlestown, Massachusetts, United States
Background
Apical cilia move fluid and participate in sensing the extracellular environment. Cilia dysfunction or "ciliopathy" results from disruptions in cilia structure or failed localization of ciliary proteins. Mutations in the centrosomal protein of 290 kDa (CEP290/NPHP6) gene are linked to ciliopathy syndromes including Juvenile nephronophthisis. CEP290 has been proposed to form a "ciliary gate" at the transition zone in Chlamydomonas reinhardtii however its roles in cilia appear to be more complex.
Methods
We generated antibodies to the cep290 N terminus and C terminus to assay localization in different cell types. cep290 was acutely disrupted using antisense morpholino oligos and phenotypes were compared with Enu or Crispr/Cas9-generated genetic cep290 mutants. RNA Seq was performed to test for upregulation of compensatory gene expression in mutants.
Results
Cep290 was localized in different cell types to the transition zone or pericentriolar satellites. Pericentriolar satellite Cep290 correlated with axoneme length defects in photoreceptors and KV cilia of morphants and mutants. In cep290 mutants, photoreceptors showed an abnormal accumulation of cytoplasmic vesicles. Cilia length defects in the KV could be rescued by over-expression of exogenous cilia membrane protein mRNAs (pkd2 or sstr3). cep290 mutants, which show a mild phenotype compared to morphants, exhibited upregulation of multiple mRNAs encoding proteins essential for cilia membrane transport (Arl3, unc119b, Arl13b). Overexpression of upregulated mRNAs partially rescued cep290 deficiency phenotypes in cep290 morphants.
Conclusion
Cep290 plays a key role in facilitating cilia membrane transport, particularly in cells where it localizes to pericentriolar satellites. Long term vs. acute loss of cep290 induces upregulation of genes encoding proteins involved in cilia membrane transport, suggesting genetic compensation in cep290 mutants. Overexpression of cilia GTPases (Arl3, unc119b, Arl13b) in cep290 human patients may provide a new route to ciliopathy gene therapy.
Funding
- NIDDK Support