title: Regulation of Chlamydomonas flagella and ependymal cell motile cilia by ceramide-mediated translocation of GSK3.
abstract: Cilia are important organelles formed by cell membrane protrusions; however, little is known about their regulation by membrane lipids. We characterize a novel activation mechanism for glycogen synthase kinase-3 (GSK3) by the sphingolipids phytoceramide and ceramide that is critical for ciliogenesis in Chlamydomonas and murine ependymal cells, respectively. We show for the first time that Chlamydomonas expresses serine palmitoyl transferase (SPT), the first enzyme in (phyto)ceramide biosynthesis. Inhibition of SPT in Chlamydomonas by myriocin led to loss of flagella and reduced tubulin acetylation, which was prevented by supplementation with the precursor dihydrosphingosine. Immunocytochemistry showed that (phyto)ceramide was colocalized with phospho-Tyr-216-GSK3 (pYGSK3) at the base and tip of Chlamydomonas flagella and motile cilia in ependymal cells. The (phyto)ceramide distribution was consistent with that of a bifunctional ceramide analogue UV cross-linked and visualized by click-chemistry-mediated fluorescent labeling. Ceramide depletion, by myriocin or neutral sphingomyelinase deficiency (fro/fro mouse), led to GSK3 dephosphorylation and defective flagella and cilia. Motile cilia were rescued and pYGSK3 localization restored by incubation of fro/fro ependymal cells with exogenous C24:1 ceramide, which directly bound to pYGSK3. Our findings suggest that (phyto)ceramide-mediated translocation of pYGSK into flagella and cilia is an evolutionarily conserved mechanism fundamental to the regulation of ciliogenesis.
authors.affiliation: Department of Neuroscience and Regenerative Medicine, Medical College of Georgia, Georgia Regents University, Augusta, GA 30912.
authors.forename: Ji Na
authors.initials: JN
authors.lastname: Kong
authors.affiliation: Department of Neuroscience and Regenerative Medicine, Medical College of Georgia, Georgia Regents University, Augusta, GA 30912.
authors.forename: Kara
authors.initials: K
authors.lastname: Hardin
authors.affiliation: Department of Neuroscience and Regenerative Medicine, Medical College of Georgia, Georgia Regents University, Augusta, GA 30912.
authors.forename: Michael
authors.initials: M
authors.lastname: Dinkins
authors.affiliation: Department of Neuroscience and Regenerative Medicine, Medical College of Georgia, Georgia Regents University, Augusta, GA 30912.
authors.forename: Guanghu
authors.initials: G
authors.lastname: Wang
authors.affiliation: Department of Neuroscience and Regenerative Medicine, Medical College of Georgia, Georgia Regents University, Augusta, GA 30912.
authors.forename: Qian
authors.initials: Q
authors.lastname: He
authors.affiliation: Department of Neuroscience and Regenerative Medicine, Medical College of Georgia, Georgia Regents University, Augusta, GA 30912.
authors.forename: Tarik
authors.initials: T
authors.lastname: Mujadzic
authors.affiliation: Department of Neuroscience and Regenerative Medicine, Medical College of Georgia, Georgia Regents University, Augusta, GA 30912.
authors.forename: Gu
authors.initials: G
authors.lastname: Zhu
authors.affiliation: Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, SC 29425.
authors.forename: Jacek
authors.initials: J
authors.lastname: Bielawski
authors.affiliation: Department of Medicine, Medical University of South Carolina, Charleston, SC 29425.
authors.forename: Stefka
authors.initials: S
authors.lastname: Spassieva
authors.affiliation: Department of Neuroscience and Regenerative Medicine, Medical College of Georgia, Georgia Regents University, Augusta, GA 30912 ebieberich@gru.edu.
authors.forename: Erhard
authors.initials: E
authors.lastname: Bieberich
mesh_terms: D000107:Acetylation; D000818:Animals; D002478:Cells, Cultured; D002518:Ceramides / Q000378:metabolism*; D002696:Chlamydomonas / Q000201:enzymology / Q000378:metabolism*; D002923:Cilia / Q000201:enzymology / Q000378:metabolism*; D004805:Ependyma / Q000166:cytology / Q000378:metabolism; D005407:Flagella / Q000201:enzymology / Q000378:metabolism*; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D051102:Serine C-Palmitoyltransferase / Q000378:metabolism; D014404:Tubulin / Q000378:metabolism
chemical_list: D002518:Ceramides; D014404:Tubulin; D051102:Serine C-Palmitoyltransferase; D038362:Glycogen Synthase Kinase 3