Intramolecular activation mechanism of the Dictyostelium LRRK2 homolog Roco protein GbpC.
GbpC is a large multidomain protein involved in cGMP-mediated chemotaxis in the cellular slime mold Dictyostelium discoideum. GbpC belongs to the Roco family of proteins that often share a central core region, consisting of leucine-rich repeats, a Ras domain (Roc), a Cor domain, and a MAPKKKinase domain. In addition to this core, GbpC contains a RasGEF domain and two cGMP-binding domains. Here, we report on an intramolecular signaling cascade of GbpC. In vitro, the RasGEF domain of GbpC specifically accelerates the GDP/GTP exchange of the Roc domain. Moreover, cGMP binding to GbpC strongly stimulates the binding of GbpC to GTP-agarose, suggesting cGMP-stimulated GDP/GTP exchange at the Roc domain. The function of the protein in vivo was investigated by rescue analysis of the chemotactic defect of gbpC null cells. Mutants that lack a functional guanine exchange factor (GEF), Roc, or kinase domain are inactive in vivo. Together, the results suggest a four-step intramolecular activation mechanism of the Roco protein GbpC: cGMP binding to the cyclic nucleotide-binding domains, activation of the GEF domain, GDP/GTP exchange of Roc, and activation of the MAPKKK domain.
van Egmond
Wouter N
WN
Department of Cell Biochemistry, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands.
Kortholt
Arjan
A
Plak
Katarzyna
K
Bosgraaf
Leonard
L
Bosgraaf
Sylvia
S
Keizer-Gunnink
Ineke
I
van Haastert
Peter J M
PJ
D000595:Amino Acid Sequence; D000818:Animals; D002633:Chemotaxis; D006152:Cyclic GMP / Q000378:metabolism; D004023:Dictyostelium / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000737:chemistry; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry / Q000378:metabolism*; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2; D008954:Models, Biological; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction
D047908:Intracellular Signaling Peptides and Proteins; D006160:Guanosine Triphosphate; C495280:LRRK2 protein, human; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2; D017346:Protein Serine-Threonine Kinases; D006152:Cyclic GMP