Phosphorylation of IQGAP1 modulates its binding to Cdc42, revealing a new type of rho-GTPase regulator.
The Rho-GTPase Cdc42 is important for the establishment and maintenance of epithelial polarity. Signaling from Cdc42 is propagated via its effector molecules that specifically bind to Cdc42 in the GTP-bound form. The cell-cell contact regulator and actin-binding protein IQGAP1 is described as effector of Cdc42 and Rac. Unexpectedly, we show in this study that IQGAP1 bound also directly nucleotide-depleted Cdc42 (Cdc42-ND). This interaction was enhanced in the presence of phosphatase inhibitors and in epithelial cells without cell-cell contacts. Tandem mass spectrometry analysis and immunoprecipitation experiments revealed that IQGAP1 was Ser1443-phosphorylated in vivo, potentially by protein kinase Cepsilon and upon loss of cell-cell contacts. In addition, we identified two independent domains of the IQGAP1 C terminus that bound exclusively Cdc42-ND. These domains interacted with each other, favoring the binding to Cdc42-GTP. Moreover, phosphorylation on Ser1443 strongly inhibited this intramolecular interaction. Thus, we unraveled a molecular mechanism that reveals a novel type of Rho-GTPase regulator. We propose that, depending on its phosphorylation state, IQGAP1 might serve as an effector or sequester nucleotide-free Cdc42 to prevent signaling.
Grohmanova
Katarina
K
ETH-Zürich, Institute of Biochemistry, Schafmattstrasse 18, Zürich 8093, Switzerland and Friedrich Miescher Institute, Basel 4002, Switzerland.
Schlaepfer
Dominik
D
Hess
Daniel
D
Gutierrez
Peter
P
Beck
Matthias
M
Kroschewski
Ruth
R
D000465:Algorithms; D000595:Amino Acid Sequence; D001665:Binding Sites; D015153:Blotting, Western; D002021:Buffers; D002450:Cell Communication; D002460:Cell Line; D045744:Cell Line, Tumor; D005786:Gene Expression Regulation; D005982:Glutathione Transferase / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry; D006153:Guanosine Diphosphate / Q000737:chemistry; D006160:Guanosine Triphosphate / Q000737:chemistry; D006801:Humans; D047468:Immunoprecipitation; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009711:Nucleotides / Q000737:chemistry; D009841:Oligonucleotides / Q000737:chemistry; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D011493:Protein Kinase C / Q000737:chemistry; D051744:Protein Kinase C-epsilon; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000737:chemistry; D015398:Signal Transduction; D012984:Software; D013997:Time Factors; D020764:cdc42 GTP-Binding Protein / Q000737:chemistry* / Q000378:metabolism; D020830:rac1 GTP-Binding Protein / Q000378:metabolism; D020703:ras GTPase-Activating Proteins / Q000378:metabolism / Q000502:physiology*; D020741:rho GTP-Binding Proteins / Q000378:metabolism*
D002021:Buffers; D020662:Guanine Nucleotide Exchange Factors; C101203:IQ motif containing GTPase activating protein 1; D009711:Nucleotides; D009841:Oligonucleotides; D011994:Recombinant Proteins; D020703:ras GTPase-Activating Proteins; D006153:Guanosine Diphosphate; D012694:Serine; D006160:Guanosine Triphosphate; D005982:Glutathione Transferase; C497255:PRKCE protein, human; D011493:Protein Kinase C; D051744:Protein Kinase C-epsilon; D020764:cdc42 GTP-Binding Protein; D020830:rac1 GTP-Binding Protein; D020741:rho GTP-Binding Proteins