NDLI: DOCK5 and DOCK1 Regulate Caco-2 Intestinal Epithelial Cell Spreading and Migration on Collagen IV*

Content Provider	PubMed Central
Author	Sanders, Matthew A. Ampasala, Dinakar Basson, Marc D.
Abstract	We observed previously that combined small interfering RNAs (siRNAs) targeting CrkII and CrkL, known activators of guanine nucleotide exchange factor DOCK1, strongly inhibit Caco-2 intestinal epithelial cell spreading and migration on collagen IV. DOCK1 siRNA reduced its expression >95% in Caco-2 cells but inhibited spreading much less than combined CrkII/CrkL siRNAs, suggesting that CrkII/CrkL interact with additional DOCK proteins. siRNA targeting DOCK5, a closely related DOCK1 family member, inhibited Caco-2 spreading similarly to DOCK1 siRNA, and the combined siRNAs synergistically inhibited spreading. Similar results were observed in human umbilical vein endothelial cells, and reverse transcriptase PCR demonstrated DOCK5 siRNA reduction of DOCK5 expression in both cell types. Combined DOCK1/DOCK5 siRNAs also inhibited Caco-2 migration and lamellipodial extension. Expression of DOCK5 cDNA, with silent mutations in the siRNA target region allowing expression simultaneously with DOCK5 siRNA, required CrkII/CrkL to restore cell spreading and DOCK5 coimmunoprecipitated with CrkII and CrkL. DOCK5 association with CrkII and CrkL was greatly reduced by mutations in their NH2-terminal SH3 domains. Expression of the DOCK5 COOH-terminal region (Met1738–Gln1870), containing potential Src homology 3 domain-binding proline-rich sites but lacking other functional regions, inhibited Caco-2 spreading and coimmunoprecipitated with CrkL. Coimmunoprecipitation of full-length DOCK5 with CrkL was strongly reduced by deletion of DOCK5 COOH-terminal amino acids 1832–1870. Green fluorescent protein-tagged DOCK5 localized to the membrane of Caco-2 cells spreading on collagen IV. In these studies, we describe human DOCK5 cloning and expression, our results indicating that, along with DOCK1, DOCK5 is an important mediator of CrkII/CrkL regulation of Caco-2 spreading and migration on collagen IV.
Related Links	http://dx.doi.org/10.1074/jbc.m808010200
Ending Page	35
Page Count	9
Starting Page	27
File Format	PDF
ISSN	00219258
e-ISSN	1083351X
Journal	The Journal of Biological Chemistry
Issue Number	1
Volume Number	284
Language	English
Publisher	American Society for Biochemistry and Molecular Biology
Publisher Date	2009-01-02
Access Restriction	Open
Rights Holder	American Society for Biochemistry and Molecular Biology
Subject Keyword	Cell Biology Biochemistry Molecular Biology Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Cell Biology Biochemistry Molecular Biology

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DOCK5 and DOCK1 Regulate Caco-2 Intestinal Epithelial Cell Spreading and Migration on Collagen IV

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