O-glycosylation is essential for intracellular targeting of synaptotagmins I and II in non-neuronal specialized secretory cells

Atiya-Nasagi, Yafit; Cohen, Hila; Medalia, Ora; Fukuda, Mitsunori; Sagi‐Eisenberg, Ronit

doi:10.1242/jcs.01710

Cited by 15 publications

(15 citation statements)

References 40 publications

(60 reference statements)

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“…O-Glycosylation has been implicated in intracellular sorting of membrane proteins (30,(51)(52)(53), however, the molecular mechanisms behind these remain unclear. Other studies have shown that O-glycosylation may affect endocytosis (54,55). In none of these studies have a single GalNAc-transferase isoform been implicated, which may be related to redundancy in GalNAc-transferases in cells.…”

Section: Discussionmentioning

confidence: 96%

Polypeptide GalNAc-transferase T3 and Familial Tumoral Calcinosis

Kato

Jeanneau

Tarp

et al. 2006

Journal of Biological Chemistry

372

168

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Mutations in the gene encoding the glycosyltransferase polypeptide GalNAc-T3, which is involved in initiation of O-glycosylation, were recently identified as a cause of the rare autosomal recessive metabolic disorder familial tumoral calcinosis (OMIM 211900). Familial tumoral calcinosis is associated with hyperphosphatemia and massive ectopic calcifications. Here, we demonstrate that the secretion of the phosphaturic factor fibroblast growth factor 23 (FGF23) requires O-glycosylation, and that GalNAc-T3 selectively directs O-glycosylation in a subtilisin-like proprotein convertase recognition sequence motif, which blocks processing of FGF23. The study suggests a novel posttranslational regulatory model of FGF23 involving competing O-glycosylation and protease processing to produce intact FGF23.The UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase (GalNAc-transferase) 2 isoform GalNAc-T3 was recently implicated in familial tumoral calcinosis (FTC) (1, 2). GalNAc-T3 is a member of the large polypeptide GalNAc transferase gene family containing 20 genes of which 15 have been shown to encode functional enzymes. The GalNAc-transferase gene family is the largest mammalian family of glycosyltransferases, which are involved in catalysis of a single glycosidic linkage, GalNAc␣1-O-Ser/Thr, and collectively the GalNAc-transferase isoforms control the initiation of mucin-type O-glycosylation in the Golgi complex (3, 4). Mucin-type O-glycosylation is one of the most abundant forms of glycosylation of proteins, and is found on a large variety of cell membrane and secreted glycopeptides and glycoproteins. O-Glycosylation imparts unique physicochemical features to glycoproteins and O-glycans have been shown to play important functions in almost all known biological processes including intracellular sorting, cell-cell adhesion, and microbial adhesion events (5).Inactivating mutations in GALNT3 were originally discovered in FTC (1, 2, 6, 7), but more recently mutations in the gene encoding the phosphaturic factor FGF23 have also been identified in FTC (8 -10). Thus, FGF23 mutations affecting folding and secretion were identified in FTC patients without mutations in GalNAc-T3. Furthermore, ablation of FGF23 in mice leads to hyperphosphatemia resembling FTC (11). FTC patients with mutations in GalNAc-T3 or FGF23 exhibit hyperphosphatemia and have reduced serum intact FGF23 levels, which may suggest that GalNAc-T3 and FGF23 act in a common pathway. GalNAc-T3 and FGF23 were found to be co-expressed in a number of tissues (2).FGF23 is a key regulator of phosphate homeostasis (12, 13), and is an O-glycosylated glycoprotein of ϳ32 kDa (14). FGF23 is partially processed intracellularly by subtilisin-like proprotein convertases (SPC) at the consensus sequence RXXR2 (RHTR 179 ) between Arg 179 and Ser 180 (14 -16). This processing step appears to be essential in the regulation of phosphate homeostasis, because mutations in the SPC cleavage sequence prevent processing and result in autosomal dominant hypophosphatemic rickets (12). Ma...

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Section: Discussionmentioning

confidence: 96%

Polypeptide GalNAc-transferase T3 and Familial Tumoral Calcinosis

Kato

Jeanneau

Tarp

et al. 2006

Journal of Biological Chemistry

372

168

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“…Specifically, we monitored the spatiotemporal characteristics of NPY-mRFP on its route from the Golgi to the SG in living GFP-CA Rab5A-expressing cells. For this purpose, cells were placed at 20˚C 2.5 h after their cotransfection with NPY-mRFP and CA Rab5A and kept at this temperature for an additional 2.5 h to prevent protein exit from the Golgi (42,43). This period allowed the cells to recover from the transfection and to accumulate sufficient NPY- mRFP in the Golgi for further analysis.…”

Section: Rab5a Interacts With Newly Formed Sgsmentioning

confidence: 99%

Rab5 Is a Novel Regulator of Mast Cell Secretory Granules: Impact on Size, Cargo, and Exocytosis

Azouz

Zur

Efergan

et al. 2014

The Journal of Immunology

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Secretion of inflammatory mediators prestored in mast cells secretory granules (SGs) enhances immune responses such as in allergy and host defense. However, the mechanisms underlying the biogenesis of the SGs remain largely unresolved. By combining high-resolution live cell imaging and quantitative morphometric analyses, we show that the small GTPase Rab5 controls the SG size and cargo composition by a VAMP8-dependent fusion mechanism. Knockdown of the endogenous Rab5, or expression of constitutively negative mutants, significantly reduces the size of SGs and increases their number. Conversely, expression of constitutively active Rab5 mutants induces few, but giant, SGs. Both the small and giant SGs maintain their exocytosis competence. Finally, we show that Rab5-mediated fusion between Golgi-derived SGs and early endosomes precedes the maturation of the SGs, as reflected by the recruitment of Rab27B, and allows the incorporation of cargo, such as CD63, that traffics through endosomes. Collectively, our results assign Rab5 a key role in mediating mast cell SG fusion during biogenesis, thereby controlling the amount and composition of the SGs content and maintaining the communication between new and pre-existing SGs.

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“…The modification is initiated by O-linked N-acetylgalactosamine (GalNAc) to Ser or Thr residues on a peptide backbone and is involved in a variety of important biological processes, such as processing of hormones (1), endocytosis (2), and sorting of apical proteins in the Drosophila embryo (3). Mucin-type glycans are sometimes clustered, forming the ''mucin domain'' found on both membrane-bound (4) and secreted mucins (5), which function as a selective molecular barrier at the epithelial surface (6) and are involved in morphogenetic signal transduction (7).…”

mentioning

confidence: 99%

Engineering of mucin-type human glycoproteins in yeast cells

Amano

Chiba

Kasahara

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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Mucin-type O-glycans are the most typical O-glycans found in mammalian cells and assume many different biological roles. Here, we report a genetic engineered yeast strain capable of producing mucintype sugar chains. Genes encoding Bacillus subtilis UDP-Gal/GalNAc 4-epimerase, human UDP-Gal/GalNAc transporter, human ppGalNAc-T1, and Drosophila melanogaster core1 ␤1-3 GalT were introduced into Saccharomyces cerevisiae. The engineered yeast was able to produce a MUC1a peptide containing O-glycan and also a mucin-like glycoprotein, human podoplanin (hPod; also known as aggrus), which is a platelet-aggregating factor that requires a sialyl-core1 structure for activity. After in vitro sialylation, hPod from yeast could induce platelet aggregation. Interestingly, substitution of ppGalNAc-T1 for ppGalNAc-T3 caused a loss of platelet aggregation-inducing activity, despite the fact that the sialyl-core1 was detectable in both hPod proteins on a lectin microarray. Most of O-mannosylation, a common modification in yeast, to MUC1a was suppressed by the addition of a rhodanine-3-acetic acid derivative in the culture medium. The yeast system we describe here is able to produce glycoproteins modified at different glycosylation sites and has the potential for use in basic research and pharmaceutical applications.glycosylation engineering ͉ mucin-type glycan ͉ podoplanin M ucin-type glycosylation is one of the most abundant posttranslational modifications. The modification is initiated by O-linked N-acetylgalactosamine (GalNAc) to Ser or Thr residues on a peptide backbone and is involved in a variety of important biological processes, such as processing of hormones (1), endocytosis (2), and sorting of apical proteins in the Drosophila embryo (3). Mucin-type glycans are sometimes clustered, forming the ''mucin domain'' found on both membrane-bound (4) and secreted mucins (5), which function as a selective molecular barrier at the epithelial surface (6) and are involved in morphogenetic signal transduction (7). It is also known that changes in expression of mucin and in their glycosylation state are closely associated with the development of cancer and cancer-related processes such as cell growth, differentiation, adhesion, invasion, and immune surveillance (8). Immunohistochemical studies have identified several tumor-associated antigens (TAAs) of adenocarcinoma (9), and most TAAs on mucins were originally found as sialylated mucin-type glycans (10). Podoplanin (also called aggrus) (11, 12), one of mucin-type glycoproteins, acts as a platelet-aggregating factor for cancer cells, a finding that has attracted recent attention because it is well known that the platelet-aggregating activity of cancer cells influences tumor metastasis. In fact, the anti-human podoplanin antibody NZ-1 has an inhibitory effect on lung colonization of human podoplanintransfected cells (13). Additionally, podoplanin is a potential diagnostic marker for many tumors, including testicular tumors, several squamous cell carcinomas, and brain tumors, and may be ...

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O-glycosylation is essential for intracellular targeting of synaptotagmins I and II in non-neuronal specialized secretory cells

Cited by 15 publications

References 40 publications

Polypeptide GalNAc-transferase T3 and Familial Tumoral Calcinosis

Polypeptide GalNAc-transferase T3 and Familial Tumoral Calcinosis

Rab5 Is a Novel Regulator of Mast Cell Secretory Granules: Impact on Size, Cargo, and Exocytosis

Engineering of mucin-type human glycoproteins in yeast cells

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