CELL WALL CARBOHYDRATE EPITOPES IN THE GREEN ALGA OEDOGONIUM BHARUCHAE F. MINOR (OEDOGONIALES, CHLOROPHYTA) 1 Jose´ M. Estevez 2,3 Carnegie Institution, Plant Biology, Stanford University, Stanford, California 94305, USA Patricia I. Leonardi Departamento de Biologı ´a, Bioquı ´mica y Farmacia, Universidad Nacional del Sur, CERZOS-CONICET, 8000 Bahı ´a Blanca, Argentina and Josefina S. Alberghina Departamento de Biodiversidad y Biologı ´a Experimental, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria – Pabello ´n 2, 1428 Buenos Aires, Argentina Cell wall changes in vegetative and suffultory cells (SCs) and in oogonial structures from Oedogonium bharuchae N. D. Kamat f. minor Ve ´lez were charac- terized using monoclonal antibodies against several carbohydrate epitopes. Vegetative cells and SCs develop only a primary cell wall (PCW), whereas mature oogonial cells secrete a second wall, the oogonium cell wall (OCW). Based on histochemical and immunolabeling results, (1 fi 4)-b-glucans in the form of crystalline cellulose together with a vari- able degree of Me-esterified homogalacturonans (HGs) and hydroxyproline-rich glycoprotein (HRGP) epitopes were detected in the PCW. The OCW showed arabinosides of the extensin type and low levels of arabinogalactan-protein (AGP) glycans but lacked cellulose, at least in its crystalline form. Surprisingly, strong colabeling in the cytoplasm of mature oogonia cells with three different antibodies (LM-5, LM-6, and CCRC-M2) was found, suggesting the presence of rhamnogalacturonan I (RG-I)–like structures. Our results are discussed relating the possible functions of these cell wall epitopes with polysaccharides and O-glycoproteins during oogo- nium differentiation. This study represents the first attempt to characterize these two types of cell walls in O. bharuchae, comparing their similarities and dif- ferences with those from other green algae and land plants. This work represents a contribution to the understanding of how cell walls have evolved from simple few-celled to complex multicelled organisms. Key index words: cell wall; cellulose; green algae; HRGP; Oedogonium bharuchae f. minor; oogonium development ; pectins Abbreviations: AGP, arabinogalactan protein; BF, basic fuchsine; CW, calcofluor white; HG, homo- galacturonan; HRGP, hydroxyproline-rich glyco- protein; LSCM, laser scanning confocal microscopy; MB, methylene blue; OCW, oogo- nium cell wall; PCW, primary cell wall; RG-I, rhamnogalacturonan I; RR, ruthenium red; SC, suffultory cell, TBO, toluidine blue ortochromat- ic; XG, xyloglucan The order Oedogoniales is a monophyletic group within the Chlorophyceae lineage made up of green algae with an unusual form of cytokinesis and cell elongation by ring formation, stephanokont type of zooids, and a complex oogamous sexual reproduc- tion (Pickett-Heaps 1975, Graham and Wilcox 2000, Alberghina et al. 2006). Oogenesis begins with a highly asymmetrical division of the oogonial initial cell (Ohashi 1930, Coss and Pickett-Heaps 1974, Pickett-Heaps 1975). Following mitosis, cytokinesis cuts off the oogonial initial cell into the small basal SC and an apical daughter cell, which becomes the swollen oogonium. The SC may undergo further divisions to form more oogonial structures. Cell wall changes during oogonial development are crucial for the fertilization process (Pickett- Heaps 1975). During the expansion of the cell wall, a period of active biosynthesis of polysaccha- rides and glycoproteins takes place in the wall as the cell increases in volume several times (Ohashi 1930, Pickett-Heaps and Fowke 1970, Pickett-Heaps 1975). In addition, the cell wall in the already formed oogonium is weakened to permit the opening of the fertilization pore or circumcision (Hoffman 1971, Coss and Pickett-Heaps 1974, Mrozinska 1985). To study changes in cell wall epi- topes from vegetative to reproductive states during cell differentiation, we have chosen Oedogonium because it is an adequate model organism due to its simple thallus structure composed of a single row of cells. 1 Received 16 May 2007. Accepted 14 March 2008. 2 Author for correspondence: e-mail jestevez@berkeley.edu and jestevez@fbmc.fcen.uba.ar. 3 Present address: Departamento de Fisiologı ´a, Biologı ´a Molecular y Celular, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad Universitaria – Pabello ´n 2, 1428 Buenos Aires, Argentina. J. Phycol. 44, 1257–1268 (2008) Ó 2008 Phycological Society of America DOI: 10.1111/j.1529-8817.2008.00568.x 1257