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Drainin required for membrane fusion of the contractile vacuole in Dictyostelium is the prototype of a protein family also represented in man.

ABSTRACT: The contractile vacuole expels water by forming a channel with the plasma membrane and thus enables cells to survive in a hypo-osmotic environment. Here we characterize drainin, a Dictyostelium protein involved in this process, as the first member of a protein family represented in fission yeast, Caenorhabditis elegans and man. Gene replacement in Dictyostelium shows that drainin acts at a checkpoint of channel formation between the contractile vacuole and the plasma membrane. A green fluorescent protein fusion of drainin localizes specifically to the contractile vacuole and rescues its periodic discharge in drainin-null cells. Drainin is a peripheral membrane protein, requiring a short hydrophobic stretch in its C-terminal region for localization and function. We suggest that drainin acts in a signaling cascade that couples a volume-sensing device in the vacuolar membrane to the membrane fusion machinery.

SUBMITTER: Becker M

PROVIDER: S-EPMC1171411 | biostudies-other | 1999 Jun

REPOSITORIES: biostudies-other

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Drainin required for membrane fusion of the contractile vacuole in Dictyostelium is the prototype of a protein family also represented in man.

Becker M M Matzner M M Gerisch G G

The EMBO journal 19990601 12

The contractile vacuole expels water by forming a channel with the plasma membrane and thus enables cells to survive in a hypo-osmotic environment. Here we characterize drainin, a Dictyostelium protein involved in this process, as the first member of a protein family represented in fission yeast, Caenorhabditis elegans and man. Gene replacement in Dictyostelium shows that drainin acts at a checkpoint of channel formation between the contractile vacuole and the plasma membrane. A green fluorescen ...[more]

PMID: 10369671

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Project description:Contractile vacuole complexes are critical components of cell volume regulation and have been shown to have other functional roles in several free-living protists. However, very little is known about the functions of the contractile vacuole complex of the parasite Trypanosoma cruzi, the etiologic agent of Chagas disease, other than a role in osmoregulation. Identification of the protein composition of these organelles is important for understanding their physiological roles. We applied a combined proteomic and bioinfomatic approach to identify proteins localized to the contractile vacuole. Proteomic analysis of a T. cruzi fraction enriched for contractile vacuoles and analyzed by one-dimensional gel electrophoresis and LC-MS/MS resulted in the addition of 109 newly detected proteins to the group of expressed proteins of epimastigotes. We also identified different peptides that map to at least 39 members of the dispersed gene family 1 (DGF-1) providing evidence that many members of this family are simultaneously expressed in epimastigotes. Of the proteins present in the fraction we selected several homologues with known localizations in contractile vacuoles of other organisms and others that we expected to be present in these vacuoles on the basis of their potential roles. We determined the localization of each by expression as GFP-fusion proteins or with specific antibodies. Six of these putative proteins (Rab11, Rab32, AP180, ATPase subunit B, VAMP1, and phosphate transporter) predominantly localized to the vacuole bladder. TcSNARE2.1, TcSNARE2.2, and calmodulin localized to the spongiome. Calmodulin was also cytosolic. Our results demonstrate the utility of combining subcellular fractionation, proteomic analysis, and bioinformatic approaches for localization of organellar proteins that are difficult to detect with whole cell methodologies. The CV localization of the proteins investigated revealed potential novel roles of these organelles in phosphate metabolism and provided information on the potential participation of adaptor protein complexes in their biogenesis.

Dataset Information

Drainin required for membrane fusion of the contractile vacuole in Dictyostelium is the prototype of a protein family also represented in man.

Publications

Drainin required for membrane fusion of the contractile vacuole in Dictyostelium is the prototype of a protein family also represented in man.

Similar Datasets

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