A ZYGOTE‐SPECIFIC PROTEIN WITH HYDROXYPROLINE‐RICH GLYCOPROTEIN DOMAINS AND LECTIN‐LIKE DOMAINS INVOLVED IN THE ASSEMBLY OF THE CELL WALL OF CHLAMYDOMONAS REINHARDTII (CHLOROPHYTA)

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Abstract
The cell wall of Chlamydomonas reinhardtii zygotes, which forms rapidly after the fusion of wall-free gametes, provides a tractable system for studying the properties and assembly of hydroxyproline-rich glycoproteins, the major proteinaceous components of green algal and plant cell walls. We report the cloning of the zsp2 gene and the analysis of its ZSP-2 product, a 58.9 kDa polypeptide that is synthesized exclusively by zygotes. The protein contains two (SP) repeats, establishing it as a member of the cell wall hydroxyproline-rich glycoproteins family. It also contains a 4-fold iteration of an amino acid sequence centered around cysteine residues, a configuration found in both plant and animal lectins. Furthermore, we report four observations on pellicle composition and production. First, cell-free preparations of the pellicle matrix are rich in hydroxyproline, arabinose, and galactose and contain bundles of very long fibrils. Second, glutathione blocks pellicle formation and results in the accumulation of long fibrils in the growth medium. Third, antibody to ZSP-2 also blocks pellicle formation. Fourth, ZSP-2 immunolocalizes to the boundary between the outer layers of the wall proper and the pellicle matrix. These observations are consistent with the possibility that the Cys-rich (glutathione-sensitive) lectin-like domains of ZSP-2 may bind to sugar residues on the long fibrils and anchor them to the cell wall, thereby initiating and maintaining pellicle formation.