rgnef (p190rhogef) knockout inhibits rhoa activity, focal adhesion establishment, and cell motility downstream of integrinsrgnef(p190rhogef)淘汰赛抑制rhoa活动,粘着斑,和细胞活性整合蛋白的下游.pdfVIP
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rgnef (p190rhogef) knockout inhibits rhoa activity, focal adhesion establishment, and cell motility downstream of integrinsrgnef(p190rhogef)淘汰赛抑制rhoa活动,粘着斑,和细胞活性整合蛋白的下游
Rgnef (p190RhoGEF) Knockout Inhibits RhoA Activity,
Focal Adhesion Establishment, and Cell Motility
Downstream of Integrins
¤
Nichol L. G. Miller, Christine Lawson, Xiao Lei Chen, Ssang-Taek Lim , David D. Schlaepfer*
Moores Cancer Center, University of California San Diego, La Jolla, California, United States of America
Abstract
Background: Cell migration is a highly regulated process that involves the formation and turnover of cell-matrix contact
sites termed focal adhesions. Rho-family GTPases are molecular switches that regulate actin and focal adhesion dynamics in
cells. Guanine nucleotide exchange factors (GEFs) activate Rho-family GTPases. Rgnef (p190RhoGEF) is a ubiquitous 190 kDa
GEF implicated in the control of colon carcinoma and fibroblast cell motility.
Principal Findings: Rgnef exon 24 floxed mice (Rgnefflox) were created and crossed with cytomegalovirus (CMV)-driven Cre
recombinase transgenic mice to inactivate Rgnef expression in all tissues during early development. Heterozygous RgnefWT/flox
(Cre+) crosses yielded normal Mendelian ratios at embryonic day 13.5, but Rgnefflox/flox (Cre+) mice numbers at 3 weeks of age were
significantly less than expected. Rgnefflox/flox (Cre+) (Rgnef2/ 2) embryos and primary mouse embryo fibroblasts (MEFs) were
isolated and verified to lack Rgnef protein expression. When compared to wildtype (WT) littermate MEFs, loss of Rgnef significantly
inhibited haptotaxis migration, wound closure motility, focal adhesion number, and RhoA GTPase activation after fibronectin-
integrin stimulation. In WT MEFs, Rgnef activation occurs within 60 minutes upon fibronectin plating of cells associated with RhoA
activation. Rgnef2/ 2 MEF phenotypes were rescued by epitope-tagged Rgnef re-expression.
Conclusions: Rg
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