Journal of Biological Chemistry
Volume 286, Issue 38, 23 September 2011, Pages 33632-33640
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Glycobiology and Extracellular Matrices
Cellular Content of UDP-N-acetylhexosamines Controls Hyaluronan Synthase 2 Expression and Correlates with O-Linked N-Acetylglucosamine Modification of Transcription Factors YY1 and SP1

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Hyaluronan, a high molecular mass polysaccharide on the vertebrate cell surface and extracellular matrix, is produced at the plasma membrane by hyaluronan synthases using UDP-GlcNAc and UDP-GlcUA as substrates. The availability of these UDP-sugar substrates can limit the synthesis rate of hyaluronan. In this study, we show that the cellular level of UDP-HexNAc also controls hyaluronan synthesis by modulating the expression of HAS2 (hyaluronan synthase 2). Increasing UDP-HexNAc in HaCaT keratinocytes by adding glucosamine down-regulated HAS2 gene expression, whereas a decrease in UDP-HexNAc, realized by mannose treatment or siRNA for GFAT1 (glutamine:fructose-6-phosphate amidotransferase 1), enhanced expression of the gene. Tracing the UDP-HexNAc-initiated signal to the HAS2 promoter revealed no change in the binding of STAT3, NF-κB, and cAMP response element-binding protein, shown previously to mediate growth factor and cytokine signals on HAS2 expression. Instead, altered binding of SP1 and YY1 to the promoter correlated with cellular UDP-HexNAc content and inhibition of HAS2 expression. siRNA silencing of YY1 and SP1 confirmed their inhibitory effects on HAS2 expression. Reduced and increased levels of O-GlcNAc-modified SP1 and YY1 proteins were associated with stimulation or inhibition of HAS2 expression, respectively. Our data are consistent with the hypothesis that, by regulating the level of protein O-GlcNAc modifications, cellular UDP-HexNAc content controls HAS2 transcription and decreases the effects on hyaluronan synthesis that would result from cellular fluctuations of this substrate.

Cell Signaling
Glucose Metabolism
Hyaluronate
O-GlcNAcylation
siRNA
Transcription Regulation
SP1
UDP-glucuronic Acid
UDP-N-acetylglucosamine
YY1

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The on-line version of this article (available at http://www.jbc.org) contains supplemental Fig. S1 and Tables S1–S3.

This work was supported by the Academy of Finland (to M. I. T. and C. C.), the Sigrid Juselius Foundation and the Cancer Center of the University of Eastern Finland (to M. I. T. and R. H. T.), the EVO (Erityisvaltionosuus) Funds of the Kuopio University Hospital and the Mizutani Foundation (to M. I. T.), and the National Glycoscience Graduate School.

1

Both authors contributed equally to this work.