TY - JOUR
T1 - SGEF, a RhoG guanine nucleotide exchange factor that stimulates macropinocytosis
AU - Ellerbroek, Shawn M
AU - Wennerberg, Krister
AU - Arthur, William T
AU - Dunty, Jill M
AU - Bowman, Dan R
AU - DeMali, Kris A
AU - Der, Channing
AU - Burridge, Keith
PY - 2004/7
Y1 - 2004/7
N2 - SGEF (SH3-containing Guanine Nucleotide Exchange Factor) is a RhoGEF of unknown function. We found the SGEF protein to be expressed in many established cell lines and highly expressed in human liver tissue. SGEF stimulated the formation of large interconnected membrane ruffles across dorsal surfaces when expressed in fibroblasts. SGEF required its proline-rich amino-terminus to generate dorsal, but not lateral, membrane ruffles and a functional SH3 domain to colocalize with filamentous actin at sites of membrane protrusion. Full-length SGEF activated RhoG, but not Rac, when expressed in fibroblasts. Further, recombinant SGEF DH/PH protein exchanged nucleotide on RhoG, but not on Rac1 or Rac3, in vitro. Scanning electron microscopy of fibroblasts demonstrated that SGEF induced dorsal ruffles that were morphologically similar to those generated by constitutively active RhoG, but not constitutively active Rac1. Transient expression of SGEF stimulated fibroblast uptake of 10-kDa dextran, a marker of macropinocytosis. This required the full-length protein and a catalytically active DH domain. Finally, activated RhoG was found to be more effective than activated Rac, and comparable to SGEF, in its ability to trigger dextran uptake. Together, this work establishes SGEF as a RhoG exchange factor and provides evidence that both SGEF and RhoG regulate membrane dynamics in promotion of macropinocytosis.
AB - SGEF (SH3-containing Guanine Nucleotide Exchange Factor) is a RhoGEF of unknown function. We found the SGEF protein to be expressed in many established cell lines and highly expressed in human liver tissue. SGEF stimulated the formation of large interconnected membrane ruffles across dorsal surfaces when expressed in fibroblasts. SGEF required its proline-rich amino-terminus to generate dorsal, but not lateral, membrane ruffles and a functional SH3 domain to colocalize with filamentous actin at sites of membrane protrusion. Full-length SGEF activated RhoG, but not Rac, when expressed in fibroblasts. Further, recombinant SGEF DH/PH protein exchanged nucleotide on RhoG, but not on Rac1 or Rac3, in vitro. Scanning electron microscopy of fibroblasts demonstrated that SGEF induced dorsal ruffles that were morphologically similar to those generated by constitutively active RhoG, but not constitutively active Rac1. Transient expression of SGEF stimulated fibroblast uptake of 10-kDa dextran, a marker of macropinocytosis. This required the full-length protein and a catalytically active DH domain. Finally, activated RhoG was found to be more effective than activated Rac, and comparable to SGEF, in its ability to trigger dextran uptake. Together, this work establishes SGEF as a RhoG exchange factor and provides evidence that both SGEF and RhoG regulate membrane dynamics in promotion of macropinocytosis.
KW - Actins/analysis
KW - Animals
KW - Cell Line
KW - Fibroblasts/metabolism
KW - GTP Phosphohydrolases/analysis
KW - Guanine Nucleotide Exchange Factors/analysis
KW - Humans
KW - Pinocytosis/genetics
KW - Point Mutation/genetics
KW - rho GTP-Binding Proteins
U2 - 10.1091/mbc.e04-02-0146
DO - 10.1091/mbc.e04-02-0146
M3 - Journal article
C2 - 15133129
SN - 1059-1524
VL - 15
SP - 3309
EP - 3319
JO - Molecular Biology of the Cell
JF - Molecular Biology of the Cell
IS - 7
ER -