Journal of Cellular Biochemistry 82:246±259 (2001) Cloning and Characterization of Human Syndecan-3 Christine Berndt, Ricardo P. Casaroli-Marano, Sene Ân Vilaro Â, and Manuel Reina* Department of Cell Biology, University of Barcelona, Diagonal 645, 08028 Barcelona, Spain Abstract Syndecans are cell-surface heparan sulfate proteoglycans, which perform a variety of functions in the cell. Most important, they are co-receptors for growth factors and mediate cell±cell and cell±matrix interactions. Four syndecans (syndecan 1±4) have been described in different species. The aim of this work was the cloning and characterization of human syndecan-3. The human syndecan-3 sequence has high homology to the rat and mouse sequences, with the exception of the 5 0 -region. Syndecan-3 mRNA is mostly expressed in the nervous system, the adrenal gland, and the spleen. When different cell lines were transiently transfected with full-length syndecan-3 cDNA, it was localized to the membrane and induced the formation of long ®lopodia-like structures, microspikes, and varicosities. Consequently, the actin cytoskeleton was re-organized, since actin staining was mostly found in the cellular extensions and at the cell periphery, co-localizing with the syndecan-3 staining. The development of the phenotype depended on the presence of sugar chains, as transfected glycosaminoglycan-de®cient Chinese hamster ovary (CHO) 745 cells did not show these structural changes, nor did transfected CHO K1 cells in the presence of heparin. The similarity of the cloned DNA sequence with that of other mammalian species and the high expression in the nervous system led us to the assumption that human syndecan-3 could perform comparable functions to those described for syndecan-3 in rat and mouse. Additionally, transient transfection experiments suggest a role of human syndecan-3 in the organization of cell shape by affecting the actin cytoskeleton, possibly by transferring signals from the cell surface in a sugar-dependent mechanism. J. Cell. Biochem. 82: 246±259, 2001. ß 2001 Wiley-Liss, Inc. Key words: syndecan-3; actin cytoskeleton; ®lopodia Mammalian cells are endowed with a variety of cell-surface receptors, which mediate cell± matrix and cell ± cell adhesion. The most promi- nent members of these families are the integrins [Howe et al., 1998] and cadherins [Takeichi, 1991], but many groups have recently focused on the syndecans. Syndecans are heparan sulfate proteoglycans (HSPGs) belonging to the family of type I transmembrane proteins [Bern®eld et al., 1992; Carey, 1997a; Woods and Couchman, 1998; Zimmermann and David, 1999]. Four syndecans have been described so far: syndecan-1, -2 (®broglycan), -3 (N-synde- can), and -4 (ryudocan, amphiglycan), which are expressed by four different genes in a tissue- speci®c and developmentally regulated manner [Kim et al., 1994]. Syndecan-1 is most abundant in epithelial tissue, syndecan-2 is mainly found in ®broblasts, syndecan-3 is predominant in the central nervous system (CNS), while syndecan- 4 is expressed by multiple cell types [Zimmer- mann and David, 1999]. The main function of the syndecans is un- known. They have been described as co-recep- tors for growth factors (e.g., basic ®broblast growth factor, (bFGF)), low af®nity receptors for enzymes (such as lipoprotein lipase, LPL), mediators of cell±cell and cell±matrix interac- tions (co-operating with integrins), co-receptors ß 2001 Wiley-Liss, Inc. Abbreviations used: BFGF, basic ®broblast growth factor; bp, base pair; CHO, chinese hamster ovary; CLSM, confocal laser scanning microscopy; CNS, central nervous system; DEAE-dextran, diethylaminoethyl-dextran; ECM, extra- cellular matrix; ERM proteins, Ezrin-Radixin-Moesin pro- teins; GAG, glycosaminoglycan; GFP, green ¯uorescent protein; HB-GAM, heparin binding growth-associated molecule; HSPGs, heparan sulfate proteoglycans; LPL, lipoprotein lipase; LSC, laser scanning cytometer; LTP, long term potentiation, WASP, Wiskott-Aldrich syndrome protein; ORF, open reading frame. The nucleotide sequence reported in this paper has been submitted to the Genbank/EMBL Data Bank with regis- tration number AF248634. Grant sponsor: European Commission; Grant number: BMH4-CT98-5142; Grant sponsor: Studienstiftung des Deutschen Volkes; Fundacio  Marato  TV3. *Correspondence to: Dr. Manuel Reina, Department of Cell Biology, School of Biology, University of Barcelona, Avda. Diagonal 645, E-08028-Barcelona, Spain. E-mail: mreina@porthos.bio.ub.es Received 14 July 2000; Accepted 10 January 2001