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Vol. 10, Issue 11, 3643-3659, November 1999
and§
*Department of Biological Chemistry, Clathrin-associated adaptor protein (AP) complexes are major
structural components of clathrin-coated vesicles, functioning in
clathrin coat assembly and cargo selection. We have carried out a
systematic biochemical and genetic characterization of AP complexes in
Saccharomyces cerevisiae. Using coimmunoprecipitation, the subunit composition of two complexes, AP-1 and AP-2R, has been
defined. These results allow assignment of the 13 potential AP subunits
encoded in the yeast genome to three AP complexes. As assessed by in
vitro binding assays and coimmunoprecipitation, only AP-1 interacts
with clathrin. Individual or combined disruption of AP-1 subunit genes
in cells expressing a temperature-sensitive clathrin heavy chain
results in accentuated growth and Molecular Biology Institute, University of California,
Los Angeles, California 90095
-factor pheromone maturation
defects, providing further evidence that AP-1 is a clathrin adaptor
complex. However, in cells expressing wild-type clathrin, the same AP
subunit deletions have no effect on growth or -factor maturation.
Furthermore, gel filtration chromatography revealed normal elution
patterns of clathrin-coated vesicles in cells lacking AP-1. Similarly,
combined deletion of genes encoding the 
subunits of the three AP
complexes did not produce defects in clathrin-dependent sorting in the
endocytic and vacuolar pathways or alterations in gel filtration
profiles of clathrin-coated vesicles. We conclude that AP complexes are dispensable for clathrin function in S. cerevisiae under
normal conditions. Our results suggest that alternative factors assume key roles in stimulating clathrin coat assembly and cargo selection during clathrin-mediated vesicle formation in yeast.
Present address: Sugen, Inc., 230 East
Grand Avenue, South San Francisco, CA 94080.
§
Corresponding author. E-mail address:
gpayne{at}mednet.ucla.edu.
Molecular Biology of the Cell
Vol. 10, 3643-3659, November 1999
Copyright © 1999 by The American Society for Cell Biology
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