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* Howard Hughes Medical Institute and Department of Pharmacology and Department of Cell Biology, Yale University School
of Medicine, New Haven, Connecticut 06510; and Cellubrevin is a ubiquitously expressed
membrane protein that is localized to endosomes
throughout the endocytotic pathway and functions in
constitutive exocytosis. We report that cellubrevin binds with high specificity to BAP31, a representative
of a highly conserved family of integral membrane proteins that has recently been discovered to be binding
proteins of membrane immunoglobulins. The interaction between BAP31 and cellubrevin is sensitive to high
ionic strength and appears to require the transmembrane regions of both proteins. No other proteins of
liver membrane extracts copurified with BAP31 on immobilized recombinant cellubrevin, demonstrating that
the interaction is specific. Synaptobrevin I bound to
BAP31 with comparable affinity, whereas only weak
binding was detectable with synaptobrevin II. Furthermore, a fraction of BAP31 and cellubrevin was complexed when each of them was quantitatively immunoprecipitated from detergent extracts of fibroblasts
(BHK 21 cells). During purification of clathrin-coated
vesicles or early endosomes, BAP31 did not cofractionate with cellubrevin. Rather, the protein was enriched
in ER-containing fractions. When BHK cells were analyzed by immunocytochemistry, BAP31 did not overlap
with cellubrevin, but rather colocalized with resident proteins of the ER. In addition, immunoreactive vesicles were clustered in a paranuclear region close to the
microtubule organizing center, but different from the
Golgi apparatus. When microtubules were depolymerized with nocodazole, this accumulation disappeared and BAP31 was confined to the ER. Truncation of the
cytoplasmic tail of BAP31 prevented export of cellubrevin, but not of the transferrin receptor from the ER.
We conclude that BAP31 represents a novel class of
sorting proteins that controls anterograde transport of
certain membrane proteins from the ER to the Golgi
complex.
Max-Planck-Institut für Immunologie, D-79108 Freiburg, Germany
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