Cytological and biochemical requirements for the establishment of a polar cell.

SUMMARY Our research is aimed at understanding the biochemical and cytological basis of cell polarity in zygotes of the brown alga, Fucus distichus L. Powell. One manifestation of this polar cell is the localization of a sulphated fucan polysaccharide (F2) in only one region of the zygote cell surface, the rhizoid cell wall. The focus of this paper is centered around the mechanism responsible for the directional transport of Golgi vesicles containing F2 and the biochemical properties of F2 that might specify its localized fate. Recent findings indicate that the various sulphated polysaccharides in the brown algae are complexes resulting from linkages of two basic polymers: an α-(1→2)-linked fucan that contains high levels of ester sulphate (F3), and a uronic acid-rich polymer (F1). The fucan complex F2, which is localized in the rhizoid wall, is composed of a fucan sulphate core (F3) to which uronic acid polymers (similar to F1) are attached. Our results, using a purified endoguluronate lyase, indicate that guluronate bridges link these subunits of F2. The carbon backbone of F2 is not synthesized de novo after fertilization. However, F2 is sulphated, and possibly assembled, beginning 10 h after fertilization, after which it is locally inserted into the rhizoid wall, and held in the wall structure only by calcium ionic bonds. Although sulphation is required for localization of F2, it is not known if the uronic acid side-chains are also assembled at the time of sulphation, and/or required for localization. The fact that F3 (F2 without the side-chains) is secreted uniformly into the zygote wall suggests that the uronic acid chains of F2 may play a critical role in its localization. A sulphated F2 alone is not sufficient for its localization since in the presence of cytochalasin, vesicles containing F2 are not transported to the rhizoid. Recent studies point to a central role for a cytoskeletal element, possibly microfilaments, in the directional transport of these vesicles. We have used the techniques of isoelectric focusing and electrophoretic mobility to study surface charge of these Golgi vesicles to determine if charge might be one factor that specifies their localization. Vesicles that contain the sulphated fucan F3 are secreted randomly and have the same surface charge as those containing F2 that are directionally transported. However, there is no stable endogenous electrical current at the time when F3 vesicles are randomly secreted, whereas a current is detectable when F2 vesicles are localized. The same vesicle population without a sulphated F2 is not localized and has a significantly lower negative surface charge than the vesicles containing F2 or F3. Hence, surface charge is related to sulphation and ability to localize and is consistent with a self-electrophoretic mechanism of localization. The relationships between the sulphation, assembly and packaging of F2 into the Golgi vesicles, as well as the surface properties of the localized vesicles (other than charge) and a cytoskeletal component, must now be investigated. A discussion of our previous model for the establishment of a polar cell is presented along with various modifications in the light of more recent evidence.