Kinetics of recruitment and allosteric activation of ARHGEF25 isoforms by the heterotrimeric G-protein Gαq.
Rho GTPases are master regulators of the eukaryotic cytoskeleton. The activation of Rho GTPases is governed by Rho guanine nucleotide exchange factors (GEFs). Three RhoGEF isoforms are produced by the gene ARHGEF25; p63RhoGEF580, GEFT and a recently discovered longer isoform of 619 amino acids (p63RhoGEF619). The subcellular distribution of p63RhoGEF580and p63RhoGEF619is strikingly different in unstimulated cells, p63RhoGEF580is located at the plasma membrane and p63RhoGEF619is confined to the cytoplasm. Interestingly, we find that both P63RhoGEF580and p63RhoGEF619activate RhoGTPases to a similar extent after stimulation of Gαq coupled GPCRs. Furthermore, we show that p63RhoGEF619relocates to the plasma membrane upon activation of Gαq coupled GPCRs, resembling the well-known activation mechanism of RhoGEFs activated by Gα12/13. Synthetic recruitment of p63RhoGEF619to the plasma membrane increases RhoGEF activity towards RhoA, but full activation requires allosteric activation via Gαq. Together, these findings reveal a dual role for Gαq in RhoGEF activation, as it both recruits and allosterically activates cytosolic ARHGEF25 isoforms.
Swammerdam Institute for Life Sciences, Section of Molecular Cytology, van Leeuwenhoek Centre for Advanced Microscopy, University of Amsterdam, P.O. Box 94215, NL-1090 GE Amsterdam, The Netherlands.
Jakobus
J
van Unen
Center for Cell Analysis and Modeling, University of Connecticut Health Center, 400 Farmington Avenue, Farmington, CT 06032-6406, USA.
Taofei
T
Yin
Center for Cell Analysis and Modeling, University of Connecticut Health Center, 400 Farmington Avenue, Farmington, CT 06032-6406, USA.
Yi I
YI
Wu
Swammerdam Institute for Life Sciences, Section of Molecular Cytology, van Leeuwenhoek Centre for Advanced Microscopy, University of Amsterdam, P.O. Box 94215, NL-1090 GE Amsterdam, The Netherlands.
Marieke
M
Mastop
Swammerdam Institute for Life Sciences, Section of Molecular Cytology, van Leeuwenhoek Centre for Advanced Microscopy, University of Amsterdam, P.O. Box 94215, NL-1090 GE Amsterdam, The Netherlands.
Theodorus W J
TW
Gadella
Swammerdam Institute for Life Sciences, Section of Molecular Cytology, van Leeuwenhoek Centre for Advanced Microscopy, University of Amsterdam, P.O. Box 94215, NL-1090 GE Amsterdam, The Netherlands.
Joachim
J
Goedhart
D000494:Allosteric Regulation; D000595:Amino Acid Sequence; D002462:Cell Membrane / Q000378:metabolism; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D020033:Protein Isoforms / Q000378:metabolism; D021381:Protein Transport; D064067:Rho Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D015398:Signal Transduction
C475063:ARHGEF25 protein, human; D020033:Protein Isoforms; D064067:Rho Guanine Nucleotide Exchange Factors; D043802:GTP-Binding Protein alpha Subunits, Gq-G11