Yeast Dop1p is an essential protein that is highly conserved in evolution and whose function is largely unknown. Here, we provide evidence that Dop1p localizes to endosomes and exists in a complex with two other conserved proteins: Neo1p, a P(4)-ATPase and putative flippase, and the scaffolding protein Ysl2p/Mon2p. The latter operates during membrane budding at the tubular endosomal network/trans-Golgi network (TEN/TGN) in a process that includes clathrin recruitment via adaptor proteins. Consistent with a role for Dop1p during this process, temperature-sensitive dop1-3 cells accumulate multivesicular, elongated tubular and ring-like structures similar to those displayed by neo1 and ysl2 mutants. In further agreement with the concept of Dop1p-Neo1p-Ysl2p complex formation and co-operation, we show that dop1-3 cells exhibit reduced levels of Neo1p and Ysl2p at steady state. Conversely, mutations or deletions in NEO1 and YSL2 lead to a decrease in Dop1p levels. In addition to binding to Neo1p and Ysl2p, Dop1p can form dimers or multimers. A critical region for dimerization resides in the C-terminus with leucine zipper-like domains. Dop1p's membrane association is largely mediated by its internal region, but Ysl2p might not be crucial for membrane recruitment.

• PMID: 20477991
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Journal Article | Research Support, Non-U.S. Gov't

Authors

Barbosa S, Pratte D, Schwarz H, Pipkorn R, Singer-Krüger B

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