Podoplanin requires sialylated O-glycans for stable expression on lymphatic endothelial cells and for interaction with platelets

Abstract

Key Points• Sialylated O-glycans protects PDPN from proteolytic degradation.• Sialylated core 1 O-glycans of PDPN on lymphatic endothelial cells (LECs) are required for interacting with platelets.O-glycosylation of podoplanin (PDPN) on lymphatic endothelial cells is critical for the separation of blood and lymphatic systems by interacting with platelet C-type lectin-like receptor 2 during development. However, how O-glycosylation controls endothelial PDPN function and expression remains unclear. In this study, we report that core 1 O-glycan-deficient or desialylated PDPN was highly susceptible to proteolytic degradation by various proteases, including metalloproteinases (MMP)-2/9. We found that the lymph contained activated MMP-2/9 and incubation of the lymph reduced surface levels of PDPN on core 1 O-glycan-deficient endothelial cells, but not on wild-type ECs. The lymph from mice with sepsis induced by cecal ligation and puncture, which contained bacteriaderived sialidase, reduced PDPN levels on wild-type ECs. The MMP inhibitor, GM6001, rescued these reductions. Additionally, GM6001 treatment rescued the reduction of PDPN level on lymphatic endothelial cells in mice lacking endothelial core 1 O-glycan or cecal ligation and puncture-treated mice. Furthermore, core 1 O-glycan-deficient or desialylated PDPN impaired platelet interaction under physiological flow. These data indicate that sialylated O-glycans of PDPN are essential for platelet adhesion and prevent PDPN from proteolytic degradation primarily mediated by MMPs in the lymph. (Blood. 2014;124(24):3656-3665)

Metastatic Progression of Prostate Cancer Is Mediated by Autonomous Binding of Galectin-4-O-Glycan to Cancer Cells

“…Glycan modification of particular proteins can contribute to quality-control in protein-folding, stability and oligomerization (29,30). For example, core 1 O-glycan-deficient podoplanin, a type 1 transmembrane mucin-type O-glycoprotein, was highly susceptible to proteolytic degradation by various proteases, such as MMP-2/9 (31). Therefore, deglycosylated receptors possibly results in protein instability or degradation.…”

Plasma soluble podoplanin is a novel marker for the diagnosis of tumor occurrence and metastasis

“…In addition, our results implied that PDPN on the surface of tumor cells can be hydrolyzed, spontaneously resulting in the release of the extracellular domain into the plasma. PDPN is known to possess multiple cleavage sites for different types of proteolytic enzymes, such as serine protease, trypsin, elastase, cysteine protease calpain‐2, MMP2/9 and presenilin‐1/gamma‐secretase in the extracellular and cytoplasmic domains . Therefore, the extracellular domain of PDPN may be shed through the action of these proteolytic enzymes, especially in the tumor environment where O ‐glycosylation of PDPN is known to be misregulated.…”

Analysis of Mammalian O-Glycopeptides—We Have Made a Good Start, but There is a Long Way to Go

“…Covalent modifications of these mucintype glycans have been reported, such as the O-acetylation of neuraminic acid (1,23,34), and the sulfation of different Gal and GlcNAc units (23,35,36). Mucin-type O-glycosylation has been implicated in a wide variety of biological processes, such as interaction with pathogens (37,38), cell adhesion (39 -41), and proteolytic processing (40,(42)(43)(44)(45). Altered glycosylation patterns have been linked to different diseases.…”