MNN6, A MEMBER OF THE KRE2 MNT1 FAMILY, IS THE GENE FOR MANNOSYLPHOSPHATE TRANSFER IN SACCHAROMYCES-CEREVISIAE/

In yeast Saccharomyces cerevisiae the N-linked sugar chain is modified at different positions by the addition of mannosylphosphate, The mnn6 mutant is deficient in the mannosylphosphate transferase activity tow ard mannotetraose (Karson, E. M., and Ballou, C. E. (1978) J. Biol. Ch em. 253, 6484-6492). We have cloned the MNN6 gene by complementation. It has encoded a 446-amino acid polypeptide with the characteristics o f type II membrane protein, The deduced Mnn6p showed a significant sim ilarity to Kre2p/Mnt1p, a Golgi alpha-1,2-mannosyltransferase involved in O-glycosylation, The null mutant of MNN6 showed a normal cell grow th, less binding to Alcian blue, hypersensitivity to Calcoflour White and hygromycin B, and diminished mannosylphosphate transferase activit y toward the endoplasmic reticulum core oligosaccharide accepters (Man 8GlcNAc2-PA and Man5GlcNAc2-PA) in vitro, suggesting the involvement o f the MNNG gene in the endoplasmic reticulum core oligosaccharide phos phorylation. However, no differences were observed in N-linked mannopr otein oligosaccharides between Delta och1 Delta mnn1 cells and Delta o ch1 Delta mnn1 Delta mnn6 cells, indicating the existence of redundant genes required for the core oligosaccharide phosphorylation. Based on a dramatic decrease in polymannose outer chain phosphorylation by MNN 6 gene disruption and a determination of the mannosylphosphorylation s ite in the acceptor, it is postulated that the MNNG gene may be a stru ctural gene encoding a mannosylphosphate transferase, which recognizes any oligosaccharides with at least one alpha-1,2-linked mannobiose un it.