Paramecium genome survey: a pilot project

Dessen, Philippe; Zagulski, Marek; Gromadka, Robert; Plattner, Helmut; Kissmehl, Roland; Meyer, Éric; Bétermier, Mireille; Schultz, Joachim E.; Linder, Jürgen; Pearlman, Ronald E.; Kung, Ching; Forney, James D.; Satir, Birgit H.; Houten, Judith L. Van; Keller, Anne Marie; Froissard, Marine; Sperling, Linda; Cohen, Jean

doi:10.1016/s0168-9525(01)02307-1

Cited by 63 publications

(53 citation statements)

References 26 publications

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“…Molecular characterization of CRC-IV-1 channels. Previously two partial sequences of CRC genes could be identified in a pilot genome survey of P. tetraurelia (21), implying two clones, M22Ho1r and M24E11u(rc), containing sequences homologous to IP 3 Rs. One gene (P. tetraurelia IP 3 R N -1), represented by M24E11u(rc), was recently characterized in detail (53) (accession no.…”

Section: Resultsmentioning

confidence: 99%

Novel Types of Ca²⁺ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Ladenburger¹,

Sehring²,

Korn³

et al. 2009

Molecular and Cellular Biology

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121

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A database search of the Paramecium genome reveals 34 genes related to Ca 2؉ -release channels of the inositol-1,4,5-trisphosphate (IP 3 ) or ryanodine receptor type (IP 3 R, RyR). Phylogenetic analyses show that these Ca 2؉ release channels (CRCs) can be subdivided into six groups (Paramecium tetraurelia CRC-I to CRC-VI), each one with features in part reminiscent of IP 3 Rs and RyRs. We characterize here the P. tetraurelia CRC-IV-1 gene family, whose relationship to IP 3 Rs and RyRs is restricted to their C-terminal channel domain. CRC-IV-1 channels localize to cortical Ca 2؉ stores (alveolar sacs) and also to the endoplasmic reticulum. This is in contrast to a recently described true IP 3 channel, a group II member (P. tetraurelia IP 3 R N -1), found associated with the contractile vacuole system. Silencing of either one of these CRCs results in reduced exocytosis of dense core vesicles (trichocysts), although for different reasons. Knockdown of P. tetraurelia IP 3 R N affects trichocyst biogenesis, while CRC-IV-1 channels are involved in signal transduction since silenced cells show an impaired release of Ca 2؉ from cortical stores in response to exocytotic stimuli. Our discovery of a range of CRCs in Paramecium indicates that protozoans already have evolved multiple ways for the use of Ca 2؉ as signaling molecule.Ca 2ϩ is an important component of cell activity in all organisms, from protozoa to mammals. Thereby Ca 2ϩ may originate from the outside medium and/or from internal stores (7, 18). Ca 2ϩ release from internal stores is mediated by various Ca 2ϩ release channels (CRCs), of which the inositol-1,4,5-trisphosphate receptor (IP 3 R) and ryanodine receptor (RyR) families have been studied most extensively (8,9,29,63 (14). IP 3 generates and maintains a Ca 2ϩ gradient in the hyphal tip of Neurospora crassa and the IP 3 -sensitive channels have been reconstituted and characterized with the planar bilayer method (87). In summary, these publications suggest that IP 3 -dependent signaling pathways are conserved among unicellular organisms, including protozoa.Despite these data, the molecular characterization of IP 3 or ryanodine receptors in low eukaryotes is currently a challenge since the identification of orthologues has not been possible thus far, probably because of evolutionary sequence divergence (66). Traynor et al. (96) identified an IP 3 receptor-like protein, IplA, in Dictyostelium discoideum, which possesses regions related to IP 3 R sequences, but thus far no evidence for IP 3 interaction exists. We have recently described an IP 3 R in the ciliated protozoa Paramecium tetraurelia (referred to here as P. tetraurelia IP 3 R N ) (53), with features characteristic of mammalian IP 3 Rs in terms of topology and ability for IP 3 binding. The expression level of P. tetraurelia IP 3 R N is modulated by extracellular Ca 2ϩ concentrations ([Ca 2ϩ ] o ) and immunofluorescence studies reveal an unexpected localization to the contractile vacuole complex (CVC), the major organelle involved in osmoregulation...

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Section: Resultsmentioning

confidence: 99%

Novel Types of Ca²⁺ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Ladenburger¹,

Sehring²,

Korn³

et al. 2009

Molecular and Cellular Biology

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121

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“…Complementation cloning, with identification of the respective gene and gene product, was another approach (Haynes et aI., 1996) that proved highly rewarding in the analysis of membrane trafficking (Skouri and Cohen, 1997;Vayssie et al, 2000;Froissard et al, 2001). In the last few years, the international Paramecium genome project (Dessen et al, 2001) has provided important input. This is currently pursued on a European scale as a GDRE project (Groupement de Recherche Europeen), headed and administered by Jean Cohen and Linda Sperling (CNRS, Gif-sur-Yvette; France).…”

Section: Results Relevant To Lviembrane Trafficking In Parameciummentioning

confidence: 99%

“…On the basis of an indexed genomic library with ......,60,000 macronuclear DNA clones (Keller and Cohen, 2000), the group of Jean Cohen (CNRS, Gif-surYvette) initiated an international Paramecium genome project (Dessen et al, 2001), which delivers a steadily increasing number of sequences, including some relevant for basic aspects of membrane trafficking (Table I). It allows us to design appropriate primers and to clone the respective genes.…”

Section: B Recent Molecular Approaches To the Study Of Membrane Trafmentioning

confidence: 99%

“…For several reasons, this review concentrates on Paramecium as a model system: (i) This cell disposes of a regular "design" (Plattner, 2002); (ii) this marks several routes not so overt in other ciliated protozoa; (iii) many more analyses on membrane traffic have been executed with this cell than in other ciliates; (iv) an international Paramecium genomics project (Dessen et al, 2001;Speeling et aI., 2002) has delivered new molecular details; (v) while the basic machinery for membrane-to-membrane interactions, i.e., docking and fusion, can now safely be assumed to be the same in Paramecium as in "higher" eukaryotes, additional proteins, not previously known from other cells, have been found; (vi) Paramecium is also attractive to study molecular diversification inside one cell; and (vii) reliable molecular information on this ciliated protozoan becomes increasingly available. The newly started Tetrahymena genome project (Turkewitz et al, 2002) may allow a stimulating comparison with basic findings from the rapidly progressing Paramecium project, which steadily develops in the direction of proteomics.…”

Section: Introductionmentioning

confidence: 99%

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Molecular Aspects of Membrane Trafficking in Paramecium

Plattner

Kissmehl

2003

International Review of Cytology

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Results achieved in tile molecular biology of Paramecium have shed new light on its elaborate membrane trafficking system. Paramecium disposes not only of the standard routes (endoplasmic reticulum ---+ Golgi ---+ Iysosomes or secretory vesicles; endo-and phagosomes ---+ Iysosomes/digesting vacuoles), but also of some unique features, e.g. and elaborate phagocytic route with the cytoproct and membrane recycling to the cytopharynx, as well as the osmoregulatory system with multiple membrane fusion sites. Exocytosis sites for trichocysts (dense-core secretory vesicles), parasomal sacs (coated pits), and terminal cisternae (early endosomes) display additional regularly arranged predetermined fusion/fission sites, which now can be discussed on a molecular basis. Considering the regular, repetitive arrangements of membrane components, availability of mutants for complementation studies, sensitivity to gene silencing, and so on, Paramecium continues to be a valuable model system for analyzing membrane interactions. This review intends to set a new baseline for ongoing work along these lines.

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“…All this also holds for the ciliated protozoan, Paramecium tetraurelia [5,6]. The ongoing Paramecium genome project [7][8][9] helps to identify molecules involved [10].…”

Section: Introductionmentioning

confidence: 99%

Molecular aspects of rapid, reversible, Ca2+-dependent de-phosphorylation of pp63/parafusin during stimulated exo-endocytosis in Paramecium cells

Plattner

Kissmehl

2005

Cell Calcium

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Calf signalling governs stimulated exocytosis and exocytosis-coupled endocytosis also in Paramecium cells. Upon stimulation, the ::s10 3 dense-core exocytotic organelles (trichocysts) can be synchronously (80 ms) released, followed by endocytotic membrane resealing (350 ms) and retrieval. Paramecium is the most synchronous dense-core exocytotic system known, allowing to dissect rapidly reversible Calf -dependent phenomena. This holds for the reversible de-/re-phosphorylation cycle of a 63 kD phosphoprotein, pp63/parafusin (pf), which we have cloned, immuno-Iocalised, and characterised as phosphoglucomutase. the enzyme funneling glucose into the glycolytic pathway. It was isolated ex vivo. followed by MALDl analysis, while X-ray structure analysis was performed after heterologous expression. We found multiple phosphorylation of superficial SerlThr residues. Although present also in exo-mutants. pp63/pf is selectively de-phosphorylated only in exo+ strains during synchronous exocytosis (80 ms) and re-phosphorylated within~20 s, Le., the time required to re-establish [Calf] homeostasis. We have isolated relevant protein phosphatases and kinases and probed their activity on pp63/pf in vitro. We consider CalfIcalmodulin-activated PP2B (calcineurin, whose subunits have been cloned) relevant for de-phosphorylation. Re-phosphorylation can be achieved by two protein kinases that also have been cloned. One is activated by cGMP (PKG) which in turn is formed by Calf-activated guanylate cyclase. Another kinase, casein kinase 2. is inhibited by Calf and. hence, activated with some delay in parallel to decreasing [Calf] after exocytosis. In total, several Calf -sensitive cycles cooperate whose protein components have been localised to the cell cortex. Regulation of the phosphorylation degree of pp63/pf may affect structure binding on a microscale and/or its enzymatic activity. All this may serve fueling substrate into glycolysis with increased ATP re-formation (compromised in exo-mutants) and NADH formation. with effects on Calf signalling including mobilisation from cortical stores (alveolar sacs) and overall effects on ATP and Ca 2 + dynamics during synchronous exo-and endocytosis.

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Paramecium genome survey: a pilot project

Cited by 63 publications

References 26 publications

Novel Types of Ca²⁺ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Novel Types of Ca²⁺ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Molecular Aspects of Membrane Trafficking in Paramecium

Molecular aspects of rapid, reversible, Ca2+-dependent de-phosphorylation of pp63/parafusin during stimulated exo-endocytosis in Paramecium cells

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Paramecium genome survey: a pilot project

Cited by 63 publications

References 26 publications

Novel Types of Ca2+ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Novel Types of Ca2+ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Molecular Aspects of Membrane Trafficking in Paramecium

Molecular aspects of rapid, reversible, Ca2+-dependent de-phosphorylation of pp63/parafusin during stimulated exo-endocytosis in Paramecium cells

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Product

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Novel Types of Ca²⁺ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Novel Types of Ca²⁺ Release Channels Participate in the Secretory Cycle of Paramecium Cells