THE LIGHT-CHAIN SUBUNIT IS REQUIRED FOR CLATHRIN FUNCTION IN SACCHAROMYCES-CEREVISIAE

Clathrin, a multimeric protein involved in intracellular protein traff icking, is composed of three heavy chains (Chc) and three light chains (Clc), Upon disruption (clc1 Delta) of the single Clc-encoding gene ( CLC1) in yeast, the steady state protein levels of Chc decreased 5-10- fold compared with wild type cells; consequently, phenotypes exhibited by clc1 Delta cells may result indirectly from the loss of Chc as opp osed to the absence of Clc, As an approach to directly examine Clc fun ction, clc1 Delta strains were generated that carry a multicopy plasmi d containing the clathrin heavy chain gene (CHC1), resulting in levels of Chc 5-10-fold elevated over wild-type levels, As with deletion of CHC1, deletion of CLC1 results in defects in growth, receptor-mediated endocytosis, and maturation of the mating pheromone alpha-factor, How ever, elevated Chc expression in clc1 Delta cells partially suppresses the growth and alpha-factor maturation defects displayed by clc1 Delt a cells alone, Biochemical analyses indicate that trimerization and as sembly of Chc are perturbed in the absence of Clc, resulting in vesicu lation defects, Our results demonstrate that the light chain subunit o f clathrin is required for efficient Chc trimerization, proper formati on of clathrin coats, and the generation of clathrin-coated vesicles.