TY - JOUR
T1 - XPLN, a guanine nucleotide exchange factor for RhoA and RhoB, but not RhoC
AU - Arthur, William T
AU - Ellerbroek, Shawn M
AU - Der, Channing J
AU - Burridge, Keith
AU - Wennerberg, Krister
PY - 2002/11/8
Y1 - 2002/11/8
N2 - Rho proteins cycle between an inactive, GDP-bound state and an active, GTP-bound state. Activation of these GTPases is mediated by guanine nucleotide exchange factors (GEFs), which promote GDP to GTP exchange. In this study we have characterized XPLN, a Rho family GEF. Like other Rho GEFs, XPLN contains a tandem Dbl homology and pleckstrin homology domain topography, but lacks homology with other known functional domains or motifs. XPLN protein is expressed in the brain, skeletal muscle, heart, kidney, platelets, and macrophage and neuronal cell lines. In vitro, XPLN stimulates guanine nucleotide exchange on RhoA and RhoB, but not RhoC, RhoG, Rac1, or Cdc42. Consistent with these data, XPLN preferentially associates with RhoA and RhoB. The specificity of XPLN for RhoA and RhoB, but not RhoC, is surprising given that they share over 85% sequence identity. We determined that the inability of XPLN to exchange RhoC is mediated by isoleucine 43 in RhoC, a position occupied by valine in RhoA and RhoB. When expressed in cells, XPLN activates RhoA and RhoB, but not RhoC, and stimulates the assembly of stress fibers and focal adhesions in a Rho kinase-dependent manner. We also found that XPLN possesses transforming activity, as determined by focus formation assays. In conclusion, here we describe a Rho family GEF that can discriminate between the closely related RhoA, RhoB, and RhoC, possibly giving insight to the divergent functions of these three proteins.
AB - Rho proteins cycle between an inactive, GDP-bound state and an active, GTP-bound state. Activation of these GTPases is mediated by guanine nucleotide exchange factors (GEFs), which promote GDP to GTP exchange. In this study we have characterized XPLN, a Rho family GEF. Like other Rho GEFs, XPLN contains a tandem Dbl homology and pleckstrin homology domain topography, but lacks homology with other known functional domains or motifs. XPLN protein is expressed in the brain, skeletal muscle, heart, kidney, platelets, and macrophage and neuronal cell lines. In vitro, XPLN stimulates guanine nucleotide exchange on RhoA and RhoB, but not RhoC, RhoG, Rac1, or Cdc42. Consistent with these data, XPLN preferentially associates with RhoA and RhoB. The specificity of XPLN for RhoA and RhoB, but not RhoC, is surprising given that they share over 85% sequence identity. We determined that the inability of XPLN to exchange RhoC is mediated by isoleucine 43 in RhoC, a position occupied by valine in RhoA and RhoB. When expressed in cells, XPLN activates RhoA and RhoB, but not RhoC, and stimulates the assembly of stress fibers and focal adhesions in a Rho kinase-dependent manner. We also found that XPLN possesses transforming activity, as determined by focus formation assays. In conclusion, here we describe a Rho family GEF that can discriminate between the closely related RhoA, RhoB, and RhoC, possibly giving insight to the divergent functions of these three proteins.
KW - 3T3 Cells
KW - Amino Acid Sequence
KW - Animals
KW - Brain/metabolism
KW - Cloning, Molecular
KW - Focal Adhesions/physiology
KW - Gene Library
KW - Guanine Nucleotide Exchange Factors/chemistry
KW - HeLa Cells
KW - Humans
KW - Kinetics
KW - Leukemia, Myeloid
KW - Mice
KW - Molecular Sequence Data
KW - Organ Specificity
KW - Rabbits
KW - Recombinant Proteins/chemistry
KW - Rho Guanine Nucleotide Exchange Factors
KW - Sequence Alignment
KW - Sequence Homology, Amino Acid
KW - Substrate Specificity
KW - Transfection
KW - ras Proteins
KW - rho GTP-Binding Proteins/metabolism
KW - rhoA GTP-Binding Protein/metabolism
KW - rhoB GTP-Binding Protein/metabolism
KW - rhoC GTP-Binding Protein
U2 - 10.1074/jbc.M207401200
DO - 10.1074/jbc.M207401200
M3 - Journal article
C2 - 12221096
VL - 277
SP - 42964
EP - 42972
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 45
ER -