The Components of the Saccharomyces cerevisiaeMannosyltransferase Complex M-Pol I Have Distinct Functions in Mannan Synthesis

Stolz, Jürgen; Munro, Sean

doi:10.1074/jbc.m208023200

Cited by 62 publications

(56 citation statements)

References 64 publications

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“…3 Thus, the clustering of O-linked sugars requires high efficiency sugar transfer, which might be provided by mannosyltransferase complexes. That oligomerization enhances enzyme function has been proven for other glycosyltransferases such as UDPGlcNAc:dolichol-P GlcNAc-1-P transferase (50) or the mannosyltransferase complexes M-Pol I and M-Pol II (51)(52)(53). Furthermore, even though members of the PMT1 and PMT2 subfamily act on the same protein substrate (14,25), they might actually O-mannosylate different serine and threonine residues within one and the same protein.…”

Section: Heteromeric Protein Complexes Between Pmt1 and Pmt2 Subfamilmentioning

confidence: 99%

Members of the Evolutionarily Conserved PMT Family of ProteinO-Mannosyltransferases Form Distinct Protein Complexes among Themselves

Girrbach

Strahl

2003

Journal of Biological Chemistry

134

152

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Protein O-mannosyltransferases (PMTs) initiate the assembly of O-mannosyl glycans, an essential protein modification. Since PMTs are evolutionarily conserved in fungi but are absent in green plants, the PMT family is a putative target for new antifungal drugs, particularly in fighting the threat of phytopathogenic fungi. The PMT family is phylogenetically classified into PMT1, PMT2, and PMT4 subfamilies, which differ in protein substrate specificity. In the model organism Saccharomyces cerevisiae as well as in many other fungi the PMT family is highly redundant, and only the simultaneous deletion of PMT1/PMT2 and PMT4 subfamily members is lethal. In this study we analyzed the molecular organization of PMT family members in S. cerevisiae. We show that members of the PMT1 subfamily (Pmt1p and Pmt5p) interact in pairs with members of the PMT2 subfamily (Pmt2p and Pmt3p) and that Pmt1p-Pmt2p and Pmt5p-Pmt3p complexes represent the predominant forms. Under certain physiological conditions, however, Pmt1p interacts also with Pmt3p, and Pmt5p with Pmt2p, suggesting a compensatory cooperation that guarantees the maintenance of O-mannosylation. Unlike the PMT1/PMT2 subfamily members, the single member of the PMT4 subfamily (Pmt4p) acts as a homomeric complex. Using mutational analyses we demonstrate that the same conserved protein domains underlie both heteromeric and homomeric interactions, and we identify an invariant arginine residue of transmembrane domain two as essential for the formation and/or stability of PMT complexes in general. Our data suggest that protein-protein interactions between the PMT family members offer a point of attack to shut down overall protein O-mannosylation in fungi.Protein O-mannosylation is an evolutionarily conserved protein modification of fundamental importance in many eukaryotes. In yeasts and fungi, the attachment of O-linked mannosyl residues to proteins of the secretory pathway is essential for cell viability (1). In particular it is indispensable for cell wall integrity and normal cellular morphogenesis (2-4). Impairment of O-mannosylation also affects the stability, localization, and/or proper function of individual proteins (5-10). Furthermore, aberrant O-mannosylation can interfere with the retrograde transport of misfolded proteins across the membrane of the endoplasmic reticulum (ER) 1 (11). O-mannosylation is not only important in yeast, but also in mammals. It was recently shown that in humans, O-mannosyl glycosylation represents a new pathomechanism for muscular dystrophy and neuronal migration disorders (12, 13).In yeast and fungi, O-mannosylation is initiated in the lumen of the ER by an essential family of protein O-mannosyltransferases (PMTs). These enzymes catalyze the transfer of mannose from dolichyl phosphate-activated mannose (Dol-PMan) to serine or threonine residues of secretory proteins (2). In Saccharomyces cerevisiae, a total of seven PMT family members (Pmt1-7p) have been identified, which share almost identical hydropathy profiles that predict the PMTs to b...

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Section: Heteromeric Protein Complexes Between Pmt1 and Pmt2 Subfamilmentioning

confidence: 99%

Members of the Evolutionarily Conserved PMT Family of ProteinO-Mannosyltransferases Form Distinct Protein Complexes among Themselves

Girrbach

Strahl

2003

Journal of Biological Chemistry

134

152

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“…In it, M--Pol I, a heterodimer, acts first to extend the Och1--derived Man with further α1,6--linked mannoses. Analyses of mutants in the DXD motifs of Mnn9 and Van1 indicated that Mnn9 likely adds the first α1,6--liked Man, which is extended with 10--15 α1,6 mannoses in Van1--requiring reactions (Stolz and Munro, 2002;Rodionov et al 2009). This α1,6 backbone is then elongated with 40--60 α1,6 Man by M--Pol II.…”

Section: Formation Of Core Type N--glycan and Mannan Outer Chainsmentioning

confidence: 99%

“…The protein(s) involved have not been identified, but presumably either they, or M-Pol I, can tell from the context of an N-glycan which type of structure it is to bear (Lewis and Ballou 1991;Stolz and Munro 2002;Rodionov et al 2009). Core-type structures are completed when that a1,2-Man, as well as the two other terminal a1,2-Man on the Man 8 GlcNAc 2 structure, receives a1,3 mannoses from Mnn1.…”

Section: N-glycosylationmentioning

confidence: 99%

“…M-Pol I acts first, with its Mnn9 subunit adding the first a1,6-Man to the Och1-derived Man, upon which 10-15 a1,6-Man are added in Van1-requiring reactions (Stolz and Munro 2002;Rodionov et al 2009). This a1,6 backbone is further elongated with 40-60 a1,6-Man by M-Pol II, whose Mnn10 and Mnn11 subunits are responsible for the majority of the a1,6-Man-T activity (Jungmann et al 1999).…”

Section: N-glycosylationmentioning

confidence: 99%

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Architecture and Biosynthesis of the Saccharomyces cerevisiae Cell Wall

2012

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The wall gives a Saccharomyces cerevisiae cell its osmotic integrity; defines cell shape during budding growth, mating, sporulation, and pseudohypha formation; and presents adhesive glycoproteins to other yeast cells. The wall consists of b1,3-and b1,6-glucans, a small amount of chitin, and many different proteins that may bear N-and O-linked glycans and a glycolipid anchor. These components become cross-linked in various ways to form higher-order complexes. Wall composition and degree of cross-linking vary during growth and development and change in response to cell wall stress. This article reviews wall biogenesis in vegetative cells, covering the structure of wall components and how they are cross-linked; the biosynthesis of N-and O-linked glycans, glycosylphosphatidylinositol membrane anchors, b1,3-and b1,6-linked glucans, and chitin; the reactions that cross-link wall components; and the possible functions of enzymatic and nonenzymatic cell wall proteins. TABLE OF CONTENTS Abstract 775Introduction 777

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“…An interesting model to explain the biosynthesis of either extended or core-type oligosaccharides has been recently proposed (7). In this model, the protein carrying N-glycan chains with the first ␣-1,6-mannose added by Och1p interacts with Mnn9p and depending on the nature of this interaction Mnn9 would add either an ␣-1,6-or ␣-1,2-mannose.…”

Section: Discussionmentioning

confidence: 99%

A Search for Hyperglycosylation Signals in Yeast Glycoproteins

Conde

Cueva

Pablo

et al. 2004

Journal of Biological Chemistry

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N-oligosaccharides of Saccharomyces cerevisiae glycoproteins are classified as core and mannan types. The former contain 13-14 mannoses whereas mannan-type structures consist of an inner core extended with an outer chain of up to 200 -300 mannoses, a process known as hyperglycosylation. The selection of substrates for hyperglycosylation poses a theoretical and practical question. To identify hyperglycosylation determinants, we have analyzed the influence of the second amino acid (Xaa) of the sequon in this process using the major exoglucanase as a model. Our results indicate that negatively charged amino acids inhibit hyperglycosylation, whereas positively charged counterparts promote it. On the basis of the tridimensional structure of Exg1, we propose that Xaa influences the orientation of the inner core making it accessible to mannan polymerase I in the appropriate position for the addition of ␣-1,6-mannoses. The presence of Glu in the Xaa of the second sequon of the native exoglucanase suggests that negative selection may drive evolution of these sites. However, a comparison of invertases secreted by S. cerevisiae and Pichia anomala suggests that hyperglycosylation signals are also subjected to positive selection.Protein glycosylation in eukaryotic cells is thought to play an essential role in many processes such as protein folding and transport, maintenance of protein and cell structure, and cell recognition and adhesion, as well as other functions. From the several types of protein glycosylation, N-glycosylation has received a great deal of attention not only because of its high frequency but also because several biochemical steps involved in this biosynthetic process are shared by yeast and humans, an indication that they have been conserved throughout evolution. These conserved steps occur in the membrane (i) or the lumen (ii and iii) of the ER 1 and belong to three groups: (i) assembly of the precursor oligosaccharide, GlnNAc 2 -Man 9 Glc 3 on a lipid carrier (dolichol-PP), (ii) transfer of the oligosaccharide to the nascent or recently synthesized protein acceptor, and (iii) trimming of the three glucoses and one mannose (for recent reviews, see Refs. 1 and 2).However, once the glycoprotein leaves the ER, biochemical modification by trimming and/or addition of new sugars varies enormously between species and even between individual proteins of the same cell. This suggests that individual proteins carry the precise information for the final carbohydrate composition. In Saccharomyces cerevisiae, some of the protein-attached oligosaccharides leaving the ER (GlcNAc 2 -Man 8 ) are poorly elongated with up to 13-14 mannoses (coretype), whereas many others are further elongated by the addition of an outer chain of up to 200 mannose residues in the Golgi apparatus (mannan-type), a process commonly known as hyperglycosylation. The outer chain consists of a backbone of ␣-1,6-mannoses with ␣-1,2 branches that are decorated with terminal ␣-1,3-mannose residues (1, 3). The biosynthesis of this complex is carried out...

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The Components of the Saccharomyces cerevisiaeMannosyltransferase Complex M-Pol I Have Distinct Functions in Mannan Synthesis

Cited by 62 publications

References 64 publications

Members of the Evolutionarily Conserved PMT Family of ProteinO-Mannosyltransferases Form Distinct Protein Complexes among Themselves

Members of the Evolutionarily Conserved PMT Family of ProteinO-Mannosyltransferases Form Distinct Protein Complexes among Themselves

Architecture and Biosynthesis of the Saccharomyces cerevisiae Cell Wall

A Search for Hyperglycosylation Signals in Yeast Glycoproteins

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