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Na Luo, Michael Conwell, Akhilesh Kumar, Ryan Anderson, Yang Sun; Inositol phosphatase INPP5E in primary cilia and eye development. Invest. Ophthalmol. Vis. Sci. 2013;54(15):5014.
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© ARVO (1962-2015); The Authors (2016-present)
To investigate the function of inositol 5-phosphatase INPP5E in cilia development in the retina. Mutations of INPP5E cause Joubert Syndrome, which is characterized by retinal degeneration, renal cysts, polydactyly, and mental retardation. Previous studies have implicated primary cilia defects in Joubert Syndrome, but the function and mechanisms of INPP5E in cilia formation and retina development are not well understood.
Using antisense morpholino oligonucleotides, knockdown of INPP5E was performed in zebrafish and rescue experiments were done using wild type and mutant human mRNA. INPP5E mRNAs mutations of R378C and R435Q were generated using QuikChangeII and Ambion mMessage mMachine®. Immunofluorescence was performed to study the functional aspect of cilia in zebrafish embryos. mTOR inhibitors treatments were performed in zebrafish and cell lines to examine the role of INPP5E in cilia signaling.
In zebrafish INPP5E morphants, embryos showed a dose-dependent phenotype of retinal degeneration, microphthalmia, and body axis asymmetry. The Inpp5e morphant zebrafish exhibited shortened and decreased cilia formation in the Kupffer’s vesicle and pronephric ducts as compared to control zebrafish. Epinephrine-stimulated melanosome trafficking was delayed in the Inpp5e zebrafish morphants. The phenotypes were rescued by co-injection of human wide type INPP5E but not R378C or R435Q hINPP5E mRNA. Rapamycin was found to slow the development of the retinal phenotype in the zebrafish morphants.
Our data supports an important role of INPP5E in ciliary development and maintenance and provides a novel model to evaluate small molecules in the function of retinal development.
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