Mutation of the yeast ε-COP gene ANU2 causes abnormal nuclear morphology and defects in intracellular vesicular transport

Abstract

Previously we reported an original method of visualizing the shape of yeast nuclei by the expression of green fluorescent protein (GFP)-tagged Xenopus nucleoplasmin in Saccharomyces cerevisiae. To identify components that determine nuclear structure, we searched for mutants exhibiting abnormal nuclear morphology from a collection of temperature-sensitive yeast strains expressing GFP-tagged nucleoplasmin. Four anu mutant strains (anu1-1, 2-1, 3-1 and 4-1; ANU = abnormal nuclear morphology) that exhibited strikingly different nuclear morphologies at the restrictive temperature as compared to the wild-type were isolated. The nuclei of these mutants were irregularly shaped and often consisted of multiple lobes. ANU1, 3 and 4 were found to encode known factors Sec24p, Sec13p and Sec18p, respectively, all of which are involved in the formation or fusion of intracellular membrane resides of protein transport between the endoplasmic reticulum (ER) and the Golgi apparatus. On the other hand, ANU2 was not well characterized. Disruption of ANU2 (Δanu2) was not lethal but conferred temperature-sensitivity for growth. Electron microscopic analysis of anu2-1 cells revealed not only the abnormal nuclear morphology but also excessive accumulation of ER membranes. In addition, both anu2-1 and Δanu2 cells were defective in protein transport between the ER and the Golgi, suggesting that Anu2p has an important role in vesicular transport in the early secretory pathway. Here we show that ANU2 encodes a 34 kDa polypeptide, which shares a 20% sequence identity with the mammalian ε-COP. Our results suggest that Anu2p is the yeast homologue of mammalian ε-Cop and the abrupt accumulation of the ER membrane caused by a blockage of the early protein transport pathway leads to alteration of nuclear morphology of the budding yeast cells.