title: Structural determinants of RGS-RhoGEF signaling critical to Entamoeba histolytica pathogenesis.
abstract: G protein signaling pathways, as key components of physiologic responsiveness and timing, are frequent targets for pharmacologic intervention. Here, we identify an effector for heterotrimeric G protein α subunit (EhGα1) signaling from Entamoeba histolytica, the causative agent of amoebic colitis. EhGα1 interacts with this effector and guanosine triphosphatase-accelerating protein, EhRGS-RhoGEF, in a nucleotide state-selective fashion. Coexpression of EhRGS-RhoGEF with constitutively active EhGα1 and EhRacC leads to Rac-dependent spreading in Drosophila S2 cells. EhRGS-RhoGEF overexpression in E. histolytica trophozoites leads to reduced migration toward serum and lower cysteine protease activity, as well as reduced attachment to, and killing of, host cells. A 2.3 Å crystal structure of the full-length EhRGS-RhoGEF reveals a putative inhibitory helix engaging the Dbl homology domain Rho-binding surface and the pleckstrin homology domain. Mutational analysis of the EhGα1/EhRGS-RhoGEF interface confirms a canonical "regulator of G protein signaling" domain rather than a RhoGEF-RGS ("rgRGS") domain, suggesting a convergent evolution toward heterotrimeric and small G protein cross-talk.
authors.affiliation: Department of Pharmacology, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
authors.forename: Dustin E
authors.initials: DE
authors.lastname: Bosch
authors.affiliation: Department of Pharmacology, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
authors.forename: Adam J
authors.initials: AJ
authors.lastname: Kimple
authors.affiliation: Department of Biology, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
authors.forename: Alyssa J
authors.initials: AJ
authors.lastname: Manning
authors.affiliation: Department of Pharmacology, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
authors.forename: Robin E
authors.initials: RE
authors.lastname: Muller
authors.affiliation: Department of Pharmacology, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
authors.forename: Francis S
authors.initials: FS
authors.lastname: Willard
authors.affiliation: Department of Pharmacology, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
authors.forename: Mischa
authors.initials: M
authors.lastname: Machius
authors.affiliation: Department of Biology, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
authors.forename: Stephen L
authors.initials: SL
authors.lastname: Rogers
authors.affiliation: Department of Physiology and Pharmacology, West Virginia University School of Medicine, Robert C. Byrd Health Sciences Center, Morgantown, WV 26506, USA. Electronic address: dpsiderovski@hsc.wvu.edu.
authors.forename: David P
authors.initials: DP
authors.lastname: Siderovski
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D002448:Cell Adhesion; D002460:Cell Line; D048430:Cell Shape; D002470:Cell Survival; D002633:Chemotaxis; D018360:Crystallography, X-Ray; D004331:Drosophila melanogaster; D004748:Entamoeba histolytica / Q000378:metabolism / Q000502:physiology*; D044385:GTP-Binding Protein alpha Subunits / Q000737:chemistry; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000737:chemistry; D006790:Host-Parasite Interactions; D006868:Hydrolysis; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D017433:Protein Structure, Secondary; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D064067:Rho Guanine Nucleotide Exchange Factors; D015398:Signal Transduction; D053058:Trophozoites / Q000378:metabolism / Q000502:physiology*
chemical_list: D044385:GTP-Binding Protein alpha Subunits; D020662:Guanine Nucleotide Exchange Factors; D015800:Protozoan Proteins; D064067:Rho Guanine Nucleotide Exchange Factors; D006160:Guanosine Triphosphate