Endoplasmic reticulum/golgi nucleotide sugar transporters contribute to the cellular release of UDP-sugar signaling molecules.

JULIANA I. SESMA; CHARLES R. ESTHER, JR.; SILVIA M. KREDA; LISA JONES; WANDA O'NEAL; SHOKO NISHIHARA; ROBERT A. NICHOLAS ; EDUARDO R. LAZAROWSKI

JOURNAL OF BIOLOGICAL CHEMISTRY

AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC

Lugar: Bethesda, Maryland; Año: 2009 vol. 284 p. 12572 - 12583

0021-9258

Extracellular UDP-sugars promote cellular responses by interacting with widely distributed P2Y14 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 contri- bution of endoplasmic reticulum (ER)/Golgi-resident UDP- GlcNAc transporters to the cellular release of their cognate sub- strates. 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 bron- chial 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 cerevi- siae) 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 illus- trate that ER/Golgi lumen constitutes a significant source of extracellular UDP-sugars and therefore plays a critical role in nucleotide sugar-promoted cell signaling.