Neurochemistry and Neuropharmacology
Author: Laura Camila Carrera Paez | email: Laurapaez_21315@hotmail.com
Laura C Carrera Paez 1°, Sabrina P Beltrame 1°, Yina M Carpintero Polanco 1°, Sergio R Auger 2°, Ahmad H Sabra 2°, Claudio R Bilder 3°, Claudia I Waldner 4°, Juan C Goin 1°
1° Laboratorio de Farmacología Molecular, Centro de Estudios Farmacológicos y Botánicos (CEFYBO-CONICET-UBA) and II Cátedra de Farmacología, Facultad de Medicina, Universidad de Buenos Aires. Paraguay 2155, Piso 16, (C1121ABG) Buenos Aires, Argentina.
2° Hospital D.F. Santojanni, Pilar 950, PB, (C1408INH) Buenos Aires, Argentina
3° Laboratorio de Neurogastroenterología, Fundación Favaloro-Hospital Universitario, Av. Belgrano 1746, 1er Piso, (C1093AAS) Buenos Aires, Argentina.
4° Laboratorio de Inmunidad Celular y Molecular, Centro de Estudios Farmacológicos y Botánicos (CEFYBO-CONICET-UBA), Facultad de Medicina, Universidad de Buenos Aires. Paraguay 2155, Piso 16, (C1121ABG) Buenos Aires, Argentina.
Serum levels of IgG autoantibodies against M2 muscarinic receptors (M2R AAb) correlate with the degree of cardiac vagal impairment in chronic Chagas disease (CCD). Recent data showed that the exposure of cardiac M2R to M2R AAb could attenuate agonist-induced Gi protein activation and arrestin-2 (Arr-2) recruitment in heterologous mammalian cells. In this study, we used Bioluminescent Resonance Energy Transfer to investigate the ability of M2R AAb from CCD patients with dysautonomia to modulate agonist-induced β-arrestin recruitment through an allosteric mechanism. Carbachol treatment of HEK 293T cells expressing M2R fused to Renilla luciferase (RM2-RLuc) and Arr-2 (or Arr-3) fused to yellow fluorescent protein (Arr-2-YFP or Arr-3-YFP) (BRET pair) stimulated the interaction between M2R and Arr-2 (or Arr-3) in the presence of GRK2. However, treatment of cells with serum IgG fractions from seropositive patients for M2R AAb (IgG Ch+) or IgG from uninfected individuals (control IgG) failed to promote Arr-2 or Arr-3 translocation, suggesting that these AAb are unable to desensitize M2R. Preincubation of cells coexpressing the BRET pair and GRK2 with IgG Ch+, followed by the addition of carbachol, resulted in a noncompetitive inhibition of Arr-2 recruitment (but not Arr-3), whereas control IgG was unable to modulate the translocation of either β-arrestin. This study suggests that M2R AAb act as Arr-2-biased allosteric modulators of agonist efficacy.