Entamoeba histolytica: inhibition of cellular functions by overexpression of EhGEF1, a novel Rho/Rac guanine nucleotide exchange factor.
The molecular, biochemical, and cellular characterization of EhGEF1 protein is described. Complete cDNA sequence of 1890 bp revealed an open reading frame that encodes a protein of 69 kDa. EhGEF1 is constituted of Dbl homology domain, pleckstrin homology domain, and several putative regulation sites. Studies of guanine nucleotide exchange activity of EhGEF1 on several GTPases from Entamoeba histolytica and Homo sapiens showed preferential activation on EhRacG, suggesting that EhGEF1 protein could be involved in mechanisms related to actin cytoskeleton activation, cytokinesis, capping, and uroid formation in trophozoite. Confocal microscopy studies of pExEhNeo/HSV-tagged-EhGEF1-transfected cells showed that trophozoites stimulated with ConA, EhGEF1, and EhRacG were localized at plasma membrane. Cellular studies showed that F-actin content of pExEhNeo/HSV-tagged-EhGEF1-transfected trophozoites as well as cellular migration and cell damage capacity were significantly altered. The observations suggest that EhRacG was the principal target of EhGEF1 and that EhGEF1 may provide a link between F-actin dynamics and EhRacG signaling.
Aguilar-Rojas
Arturo
A
Departamento de Biomedicina Molecular, Centro de Investigación y de Estudios, Avanzados del IPN, Mexico.
Almaraz-Barrera
Ma de Jesús
Mde J
Krzeminski
Mickaël
M
Robles-Flores
Martha
M
Hernández-Rivas
Rosaura
R
Guillén
Nancy
N
Maroun
Rachid C
RC
Vargas
Miguel
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D003208:Concanavalin A / Q000494:pharmacology; D017124:Conserved Sequence; D018076:DNA, Complementary / Q000737:chemistry / Q000302:isolation & purification; D016054:DNA, Protozoan / Q000737:chemistry / Q000302:isolation & purification; D004748:Entamoeba histolytica / Q000235:genetics / Q000502:physiology*; D015870:Gene Expression / Q000502:physiology; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D016366:Open Reading Frames / Q000235:genetics; D016133:Polymerase Chain Reaction; D014162:Transfection; D020744:rac GTP-Binding Proteins / Q000235:genetics / Q000502:physiology; D020741:rho GTP-Binding Proteins / Q000235:genetics / Q000502:physiology
D018076:DNA, Complementary; D016054:DNA, Protozoan; D020662:Guanine Nucleotide Exchange Factors; D003208:Concanavalin A; D020744:rac GTP-Binding Proteins; D020741:rho GTP-Binding Proteins