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J. Biol. Chem., Vol. 281, Issue 14, 9393-9399, April 7, 2006

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Protein O-Fucosyltransferase 2 Adds O-Fucose to Thrombospondin Type 1 Repeats^*

Yi Luo, Kate Koles, Wendy Vorndam, Robert S. Haltiwanger¹, and Vladislav M. Panin²

From the Department of Biochemistry and Cell Biology, Institute for Cell and Developmental Biology, Stony Brook University, Stony Brook, New York 11794-5215 and the Department of Biochemistry and Biophysics, Texas A&M University, College Station, Texas 77843-2128

O-Fucose is an unusual form of glycosylation found on epidermal growth factor-like (EGF) repeats and thrombospondin type 1 repeats (TSRs) in many secreted and transmembrane proteins. Recently O-fucose on EGF repeats was shown to play important roles in Notch signaling. In contrast, physiological roles for O-fucose on TSRs are unknown. In the accompanying paper (Luo, Y., Nita-Lazar, A., and Haltiwanger, R. S. (2006) J. Biol. Chem. 281, 9385–9392), we demonstrated that an enzyme distinct from protein O-fucosyltransferase 1 adds O-fucose to TSRs. A known homologue of O-fucosyltransferase 1 is putative protein O-fucosyltransferase 2. The cDNA sequence encoding O-fucosyltransferase 2 was originally identified during a data base search for fucosyltransferases in Drosophila. Like O-fucosyltransferase 1, O-fucosyltransferase 2 is conserved from Caenorhabditis elegans to humans. Although O-fucosyltransferase 2 was assumed to be another protein O-fucosyltransferase, no biochemical characterization existed supporting this contention. Here we show that RNAi-mediated reduction of the O-fucosyltransferase 2 message significantly decreased TSR-specific O-fucosyltransferase activity in Drosophila S2 cells. We also found that O-fucosyltransferase 2 is predominantly localized in the endoplasmic reticulum compartment of these cells. Furthermore, we expressed recombinant Drosophila O-fucosyltransferase 2 and showed that it O-fucosylates TSRs but not EGF repeats in vitro. These results demonstrate that O-fucosyltransferase 2 is in fact a TSR-specific O-fucosyltransferase.

Received for publication, November 7, 2005 , and in revised form, February 7, 2006.

^* This work was supported in part by grants from the Mizutani Foundation for Glycosciences (to R. S. H.), the Carol M. Baldwin Breast Cancer Foundation (to R. S. H.), National Institutes of Health Grant GM 61126 (to R. S. H.), ARP/ATP Grant 000517-0069-2001 from the Texas Higher Education Coordinating Board and National Institutes of Health Grant GM069952 (to V. M. P.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ To whom correspondence may be addressed. Tel.: 631-632-7336; Fax: 631-632-8575; E-mail: rhaltiwanger{at}ms.cc.sunysb.edu. ² To whom correspondence may be addressed. Tel.: 979-458-4630; Fax: 979-845-9274; E-mail: panin{at}tamu.edu.

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