title: PHLP2 is essential and plays a role in ciliogenesis and microtubule assembly in Tetrahymena thermophila.
abstract: Recent studies have implicated the phosducin-like protein-2 (PHLP2) in regulation of CCT, a chaperonin whose activity is essential for folding of tubulin and actin. However, the exact molecular function of PHLP2 is unclear. Here we investigate the significance of PHLP2 in a ciliated unicellular model, Tetrahymena thermophila, by deleting its single homolog, Phlp2p. Cells lacking Phlp2p became larger and died within 96 h. Overexpressed Phlp2p-HA localized to cilia, basal bodies, and cytosol without an obvious change in the phenotype. Despite similar localization, overexpressed GFP-Phlp2p caused a dominant-negative effect. Cells overproducing GFP-Phlp2p had decreased rates of proliferation, motility and phagocytosis, as compared to wild type cells or cells overproducing a non-tagged Phlp2p. Growing GFP-Phlp2p-overexpressing cells had fewer cilia and, when deciliated, failed to regenerate cilia, indicating defects in cilia assembly. Paclitaxel-treated GFP-Phlp2p cells failed to elongate cilia, indicating a change in the microtubules dynamics. The pattern of ciliary and cytosolic tubulin isoforms on 2D gels differed between wild type and GFP-Phlp2p-overexpressing cells. Thus, in Tetrahymena, PhLP2 is essential and under specific experimental conditions its activity affects tubulin and microtubule-dependent functions including cilia assembly.
authors.affiliation: Department of Cell Biology, Nencki Institute of Experimental Biology, Warsaw, Poland.
authors.forename: Cezary
authors.initials: C
authors.lastname: Bregier
authors.forename: Lucja
authors.initials: L
authors.lastname: Krzemień-Ojak
authors.forename: Dorota
authors.initials: D
authors.lastname: Włoga
authors.forename: Maria
authors.initials: M
authors.lastname: Jerka-Dziadosz
authors.forename: Ewa
authors.initials: E
authors.lastname: Joachimiak
authors.forename: Katarzyna
authors.initials: K
authors.lastname: Batko
authors.forename: Iwona
authors.initials: I
authors.lastname: Filipiuk
authors.forename: Urszula
authors.initials: U
authors.lastname: Smietanka
authors.forename: Jacek
authors.initials: J
authors.lastname: Gaertig
authors.forename: Stanisław
authors.initials: S
authors.lastname: Fabczak
authors.forename: Hanna
authors.initials: H
authors.lastname: Fabczak
mesh_terms: D002923:Cilia / Q000378:metabolism* / Q000648:ultrastructure; D055786:Gene Knockout Techniques; D005799:Genes, Dominant; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008870:Microtubules / Q000378:metabolism*; D038081:Organogenesis; D010802:Phylogeny; D020033:Protein Isoforms / Q000378:metabolism; D015800:Protozoan Proteins / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D016808:Tetrahymena thermophila / Q000166:cytology / Q000378:metabolism* / Q000648:ultrastructure; D014404:Tubulin / Q000378:metabolism
chemical_list: D020033:Protein Isoforms; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; D014404:Tubulin; D049452:Green Fluorescent Proteins