The mammalian exocyst, a complex required for exocytosis, inhibits tubulin polymerization.
The exocyst is a 734-kDa complex essential for development. Perturbation of its function results in early embryonic lethality. Extensive investigation has revealed that this complex participates in multiple biological processes, including protein synthesis and vesicle/protein targeting to the plasma membrane. In this article we report that the exocyst may also play a role in modulating microtubule dynamics. Using monoclonal antibodies, we observed that endogenous exocyst subunits co-localized with microtubules and mitotic spindles in normal rat kidney cells. To test for a functional relationship between the exocyst complex and microtubules, we established an in vitro exocyst reconstitution assay and studied exocyst effect on microtubule dynamics. We found that the exocyst complex reconstituted from eight recombinant exocyst subunits inhibited tubulin polymerization in vitro. Deletion of exocyst subunit sec5, sec6, sec15, or exo70 diminished its tubulin polymerization inhibition activity. Surprisingly, exocyst subunit exo70 itself was also capable of inhibiting tubulin polymerization, although exocyst complex with exo70 deletion did not lose its activity completely. Overexpression of exo70 in NRK cells resulted in microtubule network disruption and the formation of filopodia-like plasma membrane protrusions. The formation of these membrane protrusions was greatly hampered by stabilizing microtubules with taxol. Overexpression of exo84, an exocyst subunit that did not show tubulin polymerization inhibition activity, did not cause this phenotype. Results shown in this article, along with a previous report that localized microtubule instability induces plasma membrane addition, implicates a novel role for the exocyst in modulating microtubule dynamics underlying exocytosis.
Wang
Sheng
S
Department of Cell Biology and Neuroscience, Rutgers University, Piscataway, New Jersey 08854, USA.
Liu
Yan
Y
Adamson
Crista L
CL
Valdez
Gregorio
G
Guo
Wei
W
Hsu
Shu C
SC
D000818:Animals; D002460:Cell Line; D019281:Dimerization; D005089:Exocytosis; D007668:Kidney / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D008870:Microtubules / Q000502:physiology*; D016716:PC12 Cells; D051381:Rats; D014404:Tubulin / Q000737:chemistry / Q000378:metabolism*; D033921:Vesicular Transport Proteins
C109901:Exoc7 protein, rat; D008565:Membrane Proteins; D014404:Tubulin; D033921:Vesicular Transport Proteins