Sulfation of fucoidin in focus embryos: III. Required for localization in the rhizoid wall

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Abstract
Zygotes of the brown alga F. distichus L. Powell accumulate a sulfated polysaccharide (fucoidin) in the cell wall at the site of rhizoid formation. Previous work indicated that zygotes grown in seawater minus sulfate do not sulfate the preformed fucan (an unsulfated fucoidin) but form rhizoids. Whether sulfation of the fucan is required for its localization in the rhizoid wall was determined. This was accomplished by developing a specific stain for both the fucan and fucoidin. Using a precipitin assay in vitro, the lectin ricin (RCAI) was found to specifically complex with both the sulfated and desulfated polysaccharide. No precipitate is observed when either is incubated in 0.1 M D-galactose or when RCAI is mixed with laminarin or alginic acid, the other major polysaccharides in Fucus. RCAI conjugated with fluorescein isothiocyanate (FITC) is also shown to bind specifically to fucoidin using a filter paper (DE81) assay. When added to zygotes, RCAI-FITC binds only to the site of fucoidin localization, i.e., the rhizoid cell wall. However, RCAI-FITC is not observed in the rhizoid wall of zygotes grown in the absence of sulfate. This observation is not due to the inability of RCAI-FITC to bind to the fucan in vivo. Chemically desulfated cell walls that contained fucoidin in the rhizoid wall bind RCAI-FITC only in the rhizoid region. Also, the concentration of fucose-containing polymers and polysaccharides that form precipitates with RCAI is the same in embryos grown in the presence or absence of sulfate. If sulfate is added back to cultures of zygotes grown without sulfate, fucoidin is detected at the rhizoid tip by RCAI-FITC several hours later. The enzymatic sulfation of the fucan is apparently a modification of the polysaccharide required for its localization and/or assembly into a specific region of the cell wall.