P4-ATPase requirement for AP-1/clathrin function in protein transport from the trans-Golgi network and early endosomes

Mol Biol Cell. 2008 Aug;19(8):3526-35. doi: 10.1091/mbc.e08-01-0025. Epub 2008 May 28.

Abstract

Drs2p is a resident type 4 P-type ATPase (P4-ATPase) and potential phospholipid translocase of the trans-Golgi network (TGN) where it has been implicated in clathrin function. However, precise protein transport pathways requiring Drs2p and how it contributes to clathrin-coated vesicle budding remain unclear. Here we show a functional codependence between Drs2p and the AP-1 clathrin adaptor in protein sorting at the TGN and early endosomes of Saccharomyces cerevisiae. Genetic criteria indicate that Drs2p and AP-1 operate in the same pathway and that AP-1 requires Drs2p for function. In addition, we show that loss of AP-1 markedly increases Drs2p trafficking to the plasma membrane, but does not perturb retrieval of Drs2p from the early endosome back to the TGN. Thus AP-1 is required at the TGN to sort Drs2p out of the exocytic pathway, presumably for delivery to the early endosome. Moreover, a conditional allele that inactivates Drs2p phospholipid translocase (flippase) activity disrupts its own transport in this AP-1 pathway. Drs2p physically interacts with AP-1; however, AP-1 and clathrin are both recruited normally to the TGN in drs2Delta cells. These results imply that Drs2p acts independently of coat recruitment to facilitate AP-1/clathrin-coated vesicle budding from the TGN.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Alleles
  • Biological Transport
  • Calcium-Transporting ATPases / metabolism*
  • Cell Membrane / metabolism
  • Clathrin / metabolism*
  • Endosomes / metabolism*
  • Exocytosis
  • Gene Expression Regulation, Fungal*
  • Golgi Apparatus / metabolism*
  • Green Fluorescent Proteins / metabolism
  • Lipid Bilayers / chemistry
  • Models, Biological
  • Saccharomyces cerevisiae / metabolism*
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Transcription Factor AP-1 / metabolism*
  • trans-Golgi Network / metabolism

Substances

  • Clathrin
  • DRS2 protein, S cerevisiae
  • Lipid Bilayers
  • Saccharomyces cerevisiae Proteins
  • Transcription Factor AP-1
  • Green Fluorescent Proteins
  • Calcium-Transporting ATPases