Functional redundancy of R7 RGS proteins in ON-bipolar cell dendrites

Abstract

PURPOSE. In the G␤5 Ϫ/Ϫ mouse, the electroretinogram (ERG) b-wave is absent, and the R7 subfamily of regulators of G protein signaling (RGS), which includes RGS6, -7, -9, and -11, is downregulated. Mutant mouse strains deficient in RGS7 or -11 were characterized, and the SG711 strain which is deficient in both proteins was examined, to learn whether the loss of some of these RGS proteins causes the absence of the ERG b-wave. METHODS. Antibodies to RGS7 and -11 were generated to determine their expression levels and localizations in retinas with various genetic backgrounds by Western blot analysis and immunohistochemistry, respectively. The implicit times and amplitudes of ERG a-and b-waves were analyzed to examine photoreceptor and bipolar cell functions. RESULTS. RGS7 and -11 co-localized to the dendritic tips of the ON-bipolar cells. In the RGS11 Ϫ/Ϫ mouse, the level of RGS7 protein increased. However, the level of RGS11 protein remained unchanged in the RGS7 mutant mouse, where a truncated RGS7 protein was expressed due to the deletion of exon 10. In the SG711 mouse retina, the G␤5-S protein level was reduced. The ERG b-wave of SG711 mice was markedly delayed. In contrast, RGS11 Ϫ/Ϫ mice showed a moderately delayed b-wave, whereas the RGS7 mutant mice showed normal ERG responses. CONCLUSIONS. The data demonstrate the presence of a delayed ERG b-wave in SG711 mice and a functionally redundant role for RGS11 and -7 at the tips of ON-bipolar cell dendrites. These results suggest that RGS11 or -7 works as the major physiological GAP (GTPase acceleration protein) for G␣o1 in ON-bipolar cells. (Invest Ophthalmol Vis Sci. 2010;51:686 -693) DOI: 10.1167/iovs.09-4084 V ision begins at retinal photoreceptors, and encoded information is relayed to bipolar cells at the retinal outer plexiform layer (OPL). The phototransduction cascade responsible for transducing light into neural signals in photoreceptors is G-protein mediated, 1 as is the metabotropic glutamate receptor 6 (mGluR6) signaling pathway responsible for relaying visual information in the depolarizing bipolar cells (DBCs). 6 Akin to the knockout mouse deficient in mGluR6 or G␣o, 4,7 the TRPM1-knockout mouse lacks the ERG b-wave. Recently, several RGS proteins, including RGS7, RGS11, and Ret-RGS1, were postulated to participate in the mGluR6 signaling pathway. 8 -10 RGS functions inside a cell as a negative regulator of a subset of heterotrimeric G-proteins. 11 Members of the RGS protein family contain a conserved RGS domain roughly 120 amino acids in length and can be further classified into subgroups by their sizes as well as sequences outside the RGS domain. RGS9-1 was the first mammalian RGS protein identified to have a physiological function. It is essential for timely deactivation of transducin in photoreceptors in mice and humans. 12,13 RGS9-1 exists in complex with two other proteins: G␤5-L (long-splice form of the fifth member of the G-protein ␤ subunit) and RGS9 anchoring protein (R9AP). 12,17 Three additional RGS proteins contain the hallmark GGL domain: RGS6, -7, and -11. Together with RGS9, they constitute the R7 subfamily of RGS proteins. 11 Unlike RGS9-1, however, these three R7 RGS proteins interact with the short-splice form of G␤5 (G␤5-S), which is more broadly expressed in the nervous system. From th