SGEF, a RhoG guanine nucleotide exchange factor that stimulates macropinocytosis

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 -