title: Hsc70 is required for endocytosis and clathrin function in Drosophila.
abstract: By screening for Drosophila mutants exhibiting aberrant bride of sevenless (Boss) staining patterns on eye imaginal disc epithelia, we have recovered a point mutation in Hsc70-4, the closest homologue to bovine clathrin uncoating ATPase. Although the mutant allele was lethal, analysis of mutant clones generated by FLP/FRT recombination demonstrated that the Sevenless-mediated internalization of Boss was blocked in mutant Hsc70-4 eye disc epithelial cells. Endocytosis of other probes was also greatly inhibited in larval Garland cells. Immunostaining and EM analysis of the mutant cells revealed disruptions in the organization of endosomal/lysosomal compartments, including a substantial reduction in the number of clathrin-coated structures in Garland cells. The Hsc70-4 mutation also interacted genetically with a dominant-negative mutant of dynamin, a gene required for the budding of clathrin-coated vesicles (CCVs). Consistent with these phenotypes, recombinant mutant Hsc70 proteins exhibited diminished clathrin uncoating activity in vitro. Together, these data provide genetic support for the long-suspected role of Hsc70 in clathrin-mediated endocytosis, at least in part by inhibiting the uncoating of CCVs.
authors.affiliation: Department of Cell Biology, Ludwig Institute for Cancer Research, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520-8002, USA.
authors.forename: Henry C
authors.initials: HC
authors.lastname: Chang
authors.forename: Sherri L
authors.initials: SL
authors.lastname: Newmyer
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Hull
authors.forename: Melanie
authors.initials: M
authors.lastname: Ebersold
authors.forename: Sandra L
authors.initials: SL
authors.lastname: Schmid
authors.forename: Ira
authors.initials: I
authors.lastname: Mellman
mesh_terms: D000818:Animals; D001360:Avidin / Q000737:chemistry / Q000378:metabolism; D002417:Cattle; D002966:Clathrin / Q000378:metabolism*; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism; D004331:Drosophila melanogaster / Q000033:anatomy & histology / Q000235:genetics / Q000502:physiology*; D034281:Dynamins / Q000235:genetics / Q000378:metabolism; D004705:Endocytosis / Q000502:physiology*; D011992:Endosomes / Q000378:metabolism; D005136:Eye Proteins / Q000235:genetics / Q000378:metabolism*; D017344:Genes, Insect; D049452:Green Fluorescent Proteins; D050883:HSC70 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008247:Lysosomes / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D017956:Photoreceptor Cells, Invertebrate / Q000254:growth & development / Q000378:metabolism / Q000648:ultrastructure; D011485:Protein Binding; D018000:Receptors, Peptide; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D000090802:rab7 GTP-Binding Proteins
chemical_list: D002966:Clathrin; D029721:Drosophila Proteins; D005136:Eye Proteins; D050883:HSC70 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D008164:Luminescent Proteins; D008562:Membrane Glycoproteins; D018000:Receptors, Peptide; D011993:Recombinant Fusion Proteins; C072141:boss protein, Drosophila; D000090802:rab7 GTP-Binding Proteins; D001360:Avidin; D049452:Green Fluorescent Proteins; D020691:rab GTP-Binding Proteins; D034281:Dynamins; C439628:shi protein, Drosophila