Originally published In Press as doi:10.1074/jbc.M408978200 on September 13, 2004
J. Biol. Chem., Vol. 279, Issue 48, 49588-49598, November 26, 2004
Molecular Basis of Anti-horseradish Peroxidase Staining in Caenorhabditis elegans*
Katharina Paschinger
,
Dubravko Rendi
,
Günter Lochnit
,
Verena Jantsch¶, and
Iain B. H. Wilson||
From the
Department für Chemie, Universität für Bodenkultur, A-1190 Wien, Austria, Institut für Biochemie, Justus-Liebig Universität, D-35292 Giessen, Germany, and ¶Institut für Botanik, Universität Wien, A-1030 Wien, Austria
Cross-reactivity with anti-horseradish peroxidase antiserum is a feature of many glycoproteins from plants and invertebrates; indeed staining with this reagent has been used to track neurons in Drosophila melanogaster and Caenorhabditis elegans. Although in insects the evidence indicates that the cross-reaction results from the presence of core 
1,3-fucosylated N-glycans, the molecular basis for anti-horseradish peroxidase staining in nematodes has been unresolved to date. By using Western blots of wild-type and mutant C. elegans extracts in conjunction with specific inhibitors, we show that the cross-reaction is due to core 1,3-fucosylation. Of the various mutants examined, one with a deletion of the fut-1 (K08F8.3) gene showed no reaction to anti-horseradish peroxidase; the molecular phenotype was rescued by injection of either the K08F8 cosmid or the fut-1 open reading frame under control of the let-858 promoter. Furthermore, expression of fut-1 cDNA in Pichia and insect cells in conjunction with antibody staining, high pressure liquid chromatography, and matrix-assisted laser desorption ionization time-of-flight mass spectrometry analyses showed that FUT-1 is a core 1,3-fucosyltransferase with an unusual substrate specificity. It is the only core fucosyltransferase in plants and animals described to date that does not require the prior action of N-acetylglucosaminyltransferase I.
Received for publication, August 5, 2004
, and in revised form, September 13, 2004.
* This work was supported by Grants P15475 (to I. B. H. W.) and P17329 (to J. Loidl, Universität Wien) from the Austrian Fonds zur Förderung der Wissenschaftlichen Forschung. 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.
The nucleotide sequence(s) reported in this paper has been submitted to the GenBankTM/EBI Data Bank with accession number(s) AJ745071, AJ505020, AJ745072, AJ745073, and AJ745074.
|| To whom correspondence should be addressed. Tel.: 43-1-36006-6541; Fax: 43-1-36006-6059; E-mail: iwilson{at}edv2.boku.ac.at.
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Copyright © 2004 by the American Society for Biochemistry and Molecular Biology.
