Fertilisation in the brown alga Fucus involves the interaction of motile, biflagellate sperm with spherical eggs. The eggs differ from those of animals in not having the equivalent of a vitelline layer, jelly coat or zona pellucida outside the plasma membrane, and in addition they are not surrounded by a cell wall. Previous studies on Fucus eggs have shown that the lectin Concanavalin A (ConA) binds in patches on the egg surface, suggesting that there is a non-uniform distribution of plasma membrane glycoproteins. In this paper we have further investigated the occurrence of domains on the Fucus egg plasma membrane using monoclonal antibodies (mAbs) and the lectins ConA and Fucose Binding Protein (FBP). Confocal laser scanning microscopy (CLSM) has been used to observe the binding of probes to the Fucus egg cell surface. Four mAbs (FS2, FS4, FS5 and FS9) raised to Fucus serratus sperm have previously been shown to cross-react with crude egg membrane vesicles. Three of these mAbs (FS2, FS4 and FS5) have now been shown to bind to the egg cell surface and they recognise glycoproteins which are organised into domains. mAb FS4 labels large areas of the egg surface, whereas mAbs FS2 and FS5 bind to smaller patches. The lectins ConA and FBP also bind to smaller, discrete domains. Western blotting results and competition binding assays have shown that mAbs FS2 and FS5 compete for binding to the same set of glycoproteins, and FS5 is used in subsequent experiments; FS4 also binds to several glycoproteins but produces a different pattern of labelling on Western blots compared to FS5, though there may be some common components. ConA labels a subset of the glycoproteins recognised by mAb FS5, and FBP recognises one major glycoprotein which is also recognised by ConA and FS5. Double labelling experiments using the CLSM, with FITC- and Au-labelled probes, have shown that the regions on the egg surface labelled by FS4 and FS5 are mainly exclusive, with small areas of overlap. There are also areas which are not labelled by either of these antibodies. The domains recognised by mAb FS5 contain smaller areas which are labelled by ConA. Overall the results show that the Fucus egg surface is heterogeneous with different sets of glycoproteins being organised into domains. With the probes used it is possible to distinguish between FS4+ FS5−, FS4− FS5+, FS4+ FS5+ and FS4− FS5− regions. Within the FS5+ domains smaller sets of glycoproteins are recognised by ConA and within these latter regions there are glycoproteins recognised by FBP.
