The Yeast RER2 Gene, Identified by Endoplasmic Reticulum Protein Localization Mutations, Encodes cis-Prenyltransferase, a Key Enzyme in Dolichol Synthesis

Abstract

As an approach to understand the molecular mechanisms of endoplasmic reticulum (ER) protein sorting, we have isolated yeast rer mutants that mislocalize a Sec12-Mfα1p fusion protein from the ER to later compartments of the secretory pathway (S. Nishikawa and A. Nakano, Proc. Natl. Acad. Sci. USA 90:8179–8183, 1993). The temperature-sensitive rer2 mutant mislocalizes different types of ER membrane proteins, suggesting that RER2 is involved in correct localization of ER proteins in general. The rer2 mutant shows several other characteristic phenotypes: slow growth, defects in N and O glycosylation, sensitivity to hygromycin B, and abnormal accumulation of membranes, including the ER and the Golgi membranes.RER2 and SRT1, a gene whose overexpression suppresses rer2, encode novel proteins similar to each other, and their double disruption is lethal.RER2 homologues are found not only in eukaryotes but also in many prokaryote species and thus constitute a large gene family which has been well conserved during evolution. Taking a hint from the phenotype of newly established mutants of the Rer2p homologue of Escherichia coli, we discovered that the rer2 mutant is deficient in the activity of cis-prenyltransferase, a key enzyme of dolichol synthesis. This and other lines of evidence let us conclude that members of the RER2 family of genes encode cis-prenyltransferase itself. The difference in phenotypes between the rer2 mutant and previously obtained glycosylation mutants suggests a novel, as-yet-unknown role of dolichol.

ACKNOWLEDGMENTS

We are grateful to M. Rose of the Massachusetts Institute of Technology; D. Gallwitz of the Max Planck Institute of Biophysical Chemistry, Göttingen, Germany; H. Riezman of the University of Basel; Y. Amaya of Niigata University; Y. Wada of Osaka University; and Suntory Limited, Osaka, Japan, for antibodies. We also thank A. Ohta of the University of Tokyo, K. Hosaka of Gunma University, M. Wachi of Tokyo Institute of Technology, H. Hara of Saitama University, and S. Fujisaki of Toho University for helpful discussions on the functions of the Rer2p family. We are particularly indebted to S. Fujisaki for valuable advice on the assay of cis-prenyltransferase. Thanks are also due to T. Koyama of Tohoku University for providing information prior to publication. Finally, we appreciate valuable discussions with the members of the Nakano laboratory.

This work was supported by grants-in-aid from the Ministry of Education, Science, Sports and Culture of Japan, by a research grant from the Human Frontier Science Program, and by funds from the Inamori Foundation and from the Biodesign Project of RIKEN. M. Sato and K. Sato are recipients of the Research Fellowship for Young Scientists from the Japan Society for the Promotion of Science.