The cytoplasmic tail of mouse hepatitis virus M protein is essential but not sufficient for its retention in the Golgi complex.
The M protein of mouse hepatitis virus (MHV) is a triple-spanning membrane glycoprotein that is exclusively O-glycosylated. When expressed independently, it accumulates in late Golgi and the trans-Golgi network (TGN) (Locker, J. K., Griffiths, G., Horzinek, M. C., and Rottier, P. J. M. (1992) (J. Biol. Chem. 267, 14094-14101). To analyze the domains of this protein responsible for its localization, we have generated deletion mutants by site-directed mutagenesis and analyzed their intracellular transport. The intracellular distribution of the mutant proteins was determined by following the extent of O-glycosylation in pulse-chase experiments, by electron microscopic immunocytochemistry, and by surface immunoprecipitation. Mutant proteins lacking the first or the first and second transmembrane domains were not efficiently retained in the Golgi complex or TGN. The latter mutant proteins also localized to endocytic compartments but were not subject to rapid lysosomal degradation. Deletion of the COOH-terminal 22 amino acids, including a tyrosine residue in the context of a potential internalization signal, resulted in plasma membrane exposure of the respective mutant protein. We show that the wild-type MHV-M protein does not recycle between the plasma membrane and the TGN, but rather behaves as a late Golgi/TGN resident in our assays. We propose that the MHV-M protein is retained in the Golgi by two signals, one contained in the cytoplasmic tail and the other determined by the transmembrane domains.
Department of Infectious Diseases and Immunology, Faculty of Veterinary Medicine, University of Utrecht, The Netherlands.
J K
JK
Locker
J
J
Klumperman
V
V
Oorschot
M C
MC
Horzinek
H J
HJ
Geuze
P J
PJ
Rottier
D000818:Animals; D002240:Carbohydrate Sequence; D006528:Carcinoma, Hepatocellular; D002460:Cell Line; D002522:Chlorocebus aethiops; D011992:Endosomes / Q000378:metabolism; D006031:Glycosylation; D006056:Golgi Apparatus / Q000378:metabolism*; D006801:Humans; D007668:Kidney; D008113:Liver Neoplasms; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D006517:Murine hepatitis virus / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D014162:Transfection; D014763:Viral Matrix Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*
D014763:Viral Matrix Proteins