Characterization of coat proteins of COPI- and clathrin-coated vesicles in the unicellular green alga Botryococcus braunii

Unicellular green algae are a useful system for studying intracellular protein transport in plant cells because they possess various plant-specific features such as cell wall formation and isolated Golgi bodies with distinct multilayered cisternae. In this study, two genes encoding the β-COP subunit of COPI-coated vesicles and the σ1 subunit of the AP-1 complex of clathrin-coated vesicles were cloned from the green alga Botryococcus braunii. The β-COP gene contains an open reading frame (ORF) that codes for a 947-amino acid protein with a deduced molecular mass of 104 kDa, while the σ1 gene contains an ORF for a 159-amino acid protein with a deduced molecular mass of 19 kDa. Polyclonal antibodies raised against these proteins recognized proteins with predicted molecular masses in cell lysates prepared from B. braunii and the green alga Chlamydomonas reinhardtii. Cell fractionation studies revealed that both β-COP and σ1 were associated with membranous compartments. By immunoelectron microscopy, β-COP was found mainly on Golgi cisternae and to a lesser extent on Golgi-associated vesicles at all levels of the Golgi bodies, consistent with a proposed role of COPI-coated vesicles in inter-compartmental transport between Golgi cisternae.