Lamellipodium extension and membrane ruffling require different SNARE-mediated trafficking pathways.
BACKGROUND
Intracellular membrane traffic is an essential component of the membrane remodeling that supports lamellipodium extension during cell adhesion. The membrane trafficking pathways that contribute to cell adhesion have not been fully elucidated, but recent studies have implicated SNARE proteins. Here, the functions of several SNAREs (SNAP23, VAMP3, VAMP4 and syntaxin13) are characterized during the processes of cell spreading and membrane ruffling.


RESULTS
We report the first description of a SNARE complex, containing SNAP23, syntaxin13 and cellubrevin/VAMP3, that is induced by cell adhesion to an extracellular matrix. Impairing the function of the SNAREs in the complex using inhibitory SNARE domains disrupted the recycling endosome, impeded delivery of integrins to the cell surface, and reduced haptotactic cell migration and spreading. Blocking SNAP23 also inhibited the formation of PMA-stimulated, F-actin-rich membrane ruffles; however, membrane ruffle formation was not significantly altered by inhibition of VAMP3 or syntaxin13. In contrast, membrane ruffling, and not cell spreading, was sensitive to inhibition of two SNAREs within the biosynthetic secretory pathway, GS15 and VAMP4. Consistent with this, formation of a complex containing VAMP4 and SNAP23 was enhanced by treatment of cells with PMA. The results reveal a requirement for the function of a SNAP23-syntaxin13-VAMP3 complex in the formation of lamellipodia during cell adhesion and of a VAMP4-SNAP23-containing complex during PMA-induced membrane ruffling.


CONCLUSIONS
Our findings suggest that different SNARE-mediated trafficking pathways support membrane remodeling during ECM-induced lamellipodium extension and PMA-induced ruffle formation, pointing to important mechanistic differences between these processes.
Skalski
Michael
M
Department of Molecular and Cellular Biology, University of Guleph, Guelph, ON N1G 2W1, Canada.
Yi
Qing
Q
Kean
Michelle J
MJ
Myers
Dennis W
DW
Williams
Karla C
KC
Burtnik
Angela
A
Coppolino
Marc G
MG
D000818:Animals; D016466:CHO Cells; D002448:Cell Adhesion / Q000235:genetics; D002465:Cell Movement / Q000235:genetics; D006224:Cricetinae; D003412:Cricetulus; D004705:Endocytosis / Q000235:genetics; D005109:Extracellular Matrix / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D015202:Protein Engineering; D017434:Protein Structure, Tertiary / Q000235:genetics; D021381:Protein Transport / Q000235:genetics; D011554:Pseudopodia / Q000235:genetics / Q000378:metabolism*; D050600:SNARE Proteins / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion / Q000235:genetics
D050600:SNARE Proteins