title: Endoplasmic reticulum/golgi nucleotide sugar transporters contribute to the cellular release of UDP-sugar signaling molecules.
abstract: Extracellular UDP-sugars promote cellular responses by interacting with widely distributed P2Y(14) receptors, but the mechanisms by which these molecules are released from cells are poorly understood. Given the active role of UDP-sugars in glycosylation reactions within the secretory pathway, we hypothesized that UDP-sugar release includes an exocytotic component. This hypothesis was tested by assessing the contribution of endoplasmic reticulum (ER)/Golgi-resident UDP-GlcNAc transporters to the cellular release of their cognate substrates. A sensitive and highly selective assay for UDP-GlcNAc mass was developed using purified AGX2, an isoenzyme of human UDP-GlcNAc pyrophosphorylase. Robust constitutive release of UDP-GlcNAc was observed in yeast as well as in well differentiated human airway epithelial cells. The human UDP-GlcNAc transporter HFRC1 was overexpressed in human bronchial epithelial cells and was shown to localize in the Golgi and to enhance the surface expression of N-acetylglucosamine-rich glycans. HFRC1-overexpressing cells also displayed increased constitutive and hypotonic stress-stimulated release of UDP-GlcNAc. Yeast mutants lacking Yea4 (the ER UDP-GlcNAc transporter endogenously expressed in Saccharomyces cerevisiae) showed reduced UDP-GlcNAc release. Yea4-deficient cells complemented with Yea4 showed UDP-GlcNAc release rates at levels similar to or higher than wild type cells. Our results illustrate that ER/Golgi lumen constitutes a significant source of extracellular UDP-sugars and therefore plays a critical role in nucleotide sugar-promoted cell signaling.
authors.affiliation: Department of Medicine, University of North Carolina, Chapel Hill, North Carolina 27599-7248, USA.
authors.forename: Juliana I
authors.initials: JI
authors.lastname: Sesma
authors.forename: Charles R
authors.initials: CR
authors.lastname: Esther
authors.forename: Silvia M
authors.initials: SM
authors.lastname: Kreda
authors.forename: Lisa
authors.initials: L
authors.lastname: Jones
authors.forename: Wanda
authors.initials: W
authors.lastname: O'Neal
authors.forename: Shoko
authors.initials: S
authors.lastname: Nishihara
authors.forename: Robert A
authors.initials: RA
authors.lastname: Nicholas
authors.forename: Eduardo R
authors.initials: ER
authors.lastname: Lazarowski
mesh_terms: D002461:Cell Line, Transformed; D004721:Endoplasmic Reticulum / Q000235:genetics / Q000378:metabolism*; D005700:Galactosyltransferases / Q000235:genetics / Q000378:metabolism*; D006056:Golgi Apparatus / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D008297:Male; D033704:Nucleotide Transport Proteins / Q000235:genetics / Q000378:metabolism*; D009997:Osmotic Pressure / Q000502:physiology; D018048:Receptors, Purinergic P2 / Q000235:genetics / Q000378:metabolism; D020545:Respiratory Mucosa / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D014536:Uridine Diphosphate N-Acetylgalactosamine / Q000235:genetics / Q000378:metabolism*
chemical_list: D007527:Isoenzymes; D033704:Nucleotide Transport Proteins; C518528:P2Y14 receptor, human; D018048:Receptors, Purinergic P2; C487823:SLC35D2 protein, human; D029701:Saccharomyces cerevisiae Proteins; D014536:Uridine Diphosphate N-Acetylgalactosamine; D005700:Galactosyltransferases; C511281:UDP-GalNAc pyrophosphorylase AGX1, human