Clathrin heavy chain is required for pinocytosis, the presence of large vacuoles, and development in Dictyostelium.

O’Halloran, Theresa J.; Anderson, Richard G. W.

doi:10.1083/jcb.118.6.1371

Cited by 121 publications

(102 citation statements)

References 45 publications

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“…24 and data not shown). We found that clathrin-minus cells were not as efficient at capping (Ϸ5% of mutant cells formed caps vs. Ϸ50% of wild-type cells), possibly due to swelling of the mutants in the hypotonic buffer used in the capping assay (19). Nonetheless, the ability of clathrin-minus cells to form Con A caps contrasted markedly with the complete failure of myosin II-null mutants to form caps.…”

Section: Resultsmentioning

confidence: 51%

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A novel role for clathrin in cytokinesis

Niswonger

O’Halloran

1997

Proc. Natl. Acad. Sci. U.S.A.

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Using clathrin-minus Dictyostelium cells, we identified a novel requirement for clathrin during cytokinesis. In suspension culture, clathrin-minus cells failed to divide and became large and multinucleate. This cytokinesis deficiency was not attributable to a pleiotropic effect on the actomyosin cytoskeleton, since other cellular events driven by myosin II (e.g., cortical contraction and capping of concanavalin A receptors) remained intact in clathrin-minus cells. Examination of cells expressing myosin II tagged with green f luorescent protein showed that clathrin-minus cells failed to assemble myosin II into a functional contractile ring. This inability to localize myosin II to a particular location was specific for cytokinesis, since clathrin-minus cells moving across a substrate localized myosin II properly to their posterior cortexes. These results demonstrate that clathrin is essential for construction of a functional contractile ring during cell division.Cytokinesis, which follows nuclear division in a coordinated fashion, is the process by which one cell divides into two (1). During late anaphase/early telophase, as condensed chromosomes move toward opposite poles of a cell, a contractile ring assembles in the center of the cell. As the contractile ring pinches the cell in half, two cells are formed. While many of the cellular properties involved in cytokinesis have been studied extensively, central questions remain. Especially lacking is information about how the plasma membrane and associated proteins integrate with the contractile ring to form a cleavage furrow during cytokinesis.The best characterized proteins of the contractile ring are actin and myosin II. Actin filaments form the scaffold for the contractile ring, whereas myosin II is the motor that drives constriction of the ring. Additional proteins, both membranebound and cytosolic, are certainly involved in contractile ring formation and function. Some proteins identified recently include racE, a small GTPase required for cytokinesis in Dictyostelium (2), and septins, identified first in Saccharomyces cerevisiae (3) and then in Drosophila (4). In addition, the actin-binding protein anillin (5, 6) and the formin-like protein diaphanous (7) are required for cytokinesis in Drosophila. In Dictyostelium, the actin-binding protein cortexillin (8) and the cytoskeletal protein coronin (9) both function in cytokinesis.

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Section: Resultsmentioning

confidence: 51%

“…We recently engineered Dictyostelium cell lines that are devoid of clathrin heavy chain and identified defects in specific membrane traffic events such as endocytosis and regulated secretion (15,19). Clathrin-minus cells also were blocked in development (17).…”

Section: Resultsmentioning

confidence: 99%

A novel role for clathrin in cytokinesis

Niswonger

O’Halloran

1997

Proc. Natl. Acad. Sci. U.S.A.

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“…Clathrin-deficient Dictyostelium cells show dramatic morphological and functional defects of the CV (O'Halloran and Anderson, 1992;Wang et al, 2003). Deletion of the apm1 gene encoding the medium subunit of the clathrin adaptorprotein AP-1 also results in the absence of a characteristic CV (Lefkir et al, 2003).…”

Section: Introductionmentioning

confidence: 99%

Acidic clusters target transmembrane proteins to the contractile vacuole inDictyosteliumcells

Mercanti

Blanc

Lefkir

et al. 2006

Journal of Cell Science

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The mechanisms responsible for the targeting of transmembrane integral proteins to the contractile vacuole (CV) network in Dictyostelium discoideum are unknown. Here we show that the transfer of the cytoplasmic domain of a CV-resident protein (Rh50) to a reporter transmembrane protein (CsA) is sufficient to address the chimera (CsA-Rh50) to the CV. We identified two clusters of acidic residues responsible for this targeting, and these motifs interacted with the gamma-adaptin AP-1 subunit in a yeast protein-protein interaction assay. For the first time we report the existence of an indirect transport pathway from the plasma membrane to the CV via endosomes. Upon internalization, the small fraction of CsA-Rh50 present at the cell surface was first concentrated in endosomes distinct from early and late p80-positive endosomes and then slowly transported to the CV. Together our results suggest the existence of an AP-1-dependent selective transport to the contractile vacuole in Dictyostelium

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“…Essentially all of the internalized fluid phase is exocytosed from cells from postlysosomes, beginning approximately 45 min after internalization; no early endosomal recycling compartment has been identified (Aubry et al, 1993;Padh et al, 1993). Endocytosis is regulated by a number of proteins in Dictyostelium including the clathrin heavy chain (O'Halloran and Anderson, 1992;Ruscetti et al, 1994), the actin cytoskeleton (Temesvari and Cardelli, unpublished results), the vacuolar H+-ATPase (Temesvari et al, 1996a), select myosin I molecules (Novak et al, 1995;Jung et al, 1996;Temesvari et al, 1996b), and phosphatidylinositol 3-kinases (Buczynski et al, 1997). It is currently not known how much the clathrin-dependent and -independent pathways individually contribute to pinocytosis.…”

Section: Introductionmentioning

confidence: 99%

Evidence for a recycling role for Rab7 in regulating a late step in endocytosis and in retention of lysosomal enzymes in Dictyostelium discoideum.

Buczynski

Bush

Zhang

et al. 1997

MBoC

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The mammalian small molecular weight GTPase Rab7 (Ypt7 in yeast) has been implicated in regulating membrane traffic at postinternalization steps along the endosomal pathway. A cDNA encoding a protein 85% identical at the amino acid level to mammalian Rab7 has been cloned from Dictyostelium discoideum. Subcellular fractionation and immunofluorescence microscopy indicated that Rab7 was enriched in lysosomes, postlysosomes, and maturing phagosomes. Cell lines were generated that overexpressed Rab7 wild-type (WT), Rab7 Q67L (constitutively active form), and Rab7 T22N (dominant negative form) proteins. The Rab7 T22N cell line internalized fluid phase markers and latex beads (phagocytosis) at one-third the rate of control cells, whereas Rab7 WT and Rab7 Q67L cell lines were normal in uptake rates but exocytosed fluid phase faster than control cells. In contrast, fluid phase markers resided in acidic compartments for longer periods of time and were more slowly exocytosed from Rab7 T22N cells as compared with control cells. Light microscopy indicated that Rab7-expressing cell lines contained morphologically altered endosomal compartments. Compared with control cells, Rab7 WT-and Rab7 Q67L-expressing cells contained a reduced number of vesicles the size of postlysosomes (>2.5 ,um) and an increased number of smaller vesicles, many of which were nonacidic; in control cells, >90% of the smaller vesicles were acidic. In contrast, Rab7 T22N cells contained an increased proportion of large acidic vesicles relative to nonacidic vesicles. Radiolabel pulse-chase experiments indicated that all of the cell lines processed and targeted lysosomal a-mannosidase normally, indicating the lack of a significant role for Rab7 in the targeting pathway; however, retention of mature lysosomal hydrolases was affected in Rab7 WT and Rab7 T22N cell lines. Contrary to the results observed for the fluid phase efflux experiments, Rab7 T22N cells oversecreted a-mannosidase, whereas Rab7 WT cells retained this hydrolase as compared with control cells. These data support a model that Rab7 may regulate retrograde transport of lysosomal enzymes and the V-type H+-ATPase from postlysosomes to lysosomes coupled with the efficient release of fluid phase from cells.

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Clathrin heavy chain is required for pinocytosis, the presence of large vacuoles, and development in Dictyostelium.

Cited by 121 publications

References 45 publications

A novel role for clathrin in cytokinesis

A novel role for clathrin in cytokinesis

Acidic clusters target transmembrane proteins to the contractile vacuole inDictyosteliumcells

Evidence for a recycling role for Rab7 in regulating a late step in endocytosis and in retention of lysosomal enzymes in Dictyostelium discoideum.

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