title: The membrane skeleton in Paramecium: Molecular characterization of a novel epiplasmin family and preliminary GFP expression results.
abstract: Previous attempts to identify the membrane skeleton of Paramecium cells have revealed a protein pattern that is both complex and specific. The most prominent structural elements, epiplasmic scales, are centered around ciliary units and are closely apposed to the cytoplasmic side of the inner alveolar membrane. We sought to characterize epiplasmic scale proteins (epiplasmins) at the molecular level. PCR approaches enabled the cloning and sequencing of two closely related genes by amplifications of sequences from a macronuclear genomic library. Using these two genes (EPI-1 and EPI-2), we have contributed to the annotation of the Paramecium tetraurelia macronuclear genome and identified 39 additional (paralogous) sequences. Two orthologous sequences were found in the Tetrahymena thermophila genome. Structural analysis of the 43 sequences indicates that the hallmark of this new multigenic family is a 79 aa domain flanked by two Q-, P- and V-rich stretches of sequence that are much more variable in amino-acid composition. Such features clearly distinguish members of the multigenic family from epiplasmic proteins previously sequenced in other ciliates. The expression of Green Fluorescent Protein (GFP)-tagged epiplasmin showed significant labeling of epiplasmic scales as well as oral structures. We expect that the GFP construct described herein will prove to be a useful tool for comparative subcellular localization of different putative epiplasmins in Paramecium.
authors.affiliation: Laboratoire de Biologie des Protistes, UMR CNRS 6023, Université Blaise Pascal, 63177 Aubière cedex, France.
authors.forename: Sébastien
authors.initials: S
authors.lastname: Pomel
authors.forename: Marie
authors.initials: M
authors.lastname: Diogon
authors.forename: Philippe
authors.initials: P
authors.lastname: Bouchard
authors.forename: Lydie
authors.initials: L
authors.lastname: Pradel
authors.forename: Viviane
authors.initials: V
authors.lastname: Ravet
authors.forename: Gérard
authors.initials: G
authors.lastname: Coffe
authors.forename: Bernard
authors.initials: B
authors.lastname: Viguès
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003598:Cytoskeletal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016054:DNA, Protozoan / Q000032:analysis; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D005810:Multigene Family; D016805:Paramecium tetraurelia / Q000235:genetics / Q000254:growth & development / Q000378:metabolism / Q000648:ultrastructure*; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D003598:Cytoskeletal Proteins; D016054:DNA, Protozoan; D008565:Membrane Proteins; D015800:Protozoan Proteins; D049452:Green Fluorescent Proteins