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

Vision 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).2 In the outer segment of a rod photoreceptor, light activates rhodopsin, which in turn activates the photoreceptor-specific G-protein, transducin. Activated transducin sequesters the inhibitory subunit of a phosphodiesterase and in turn enables its catalytic subunits to hydrolyze cyclic guanosine monophosphate (cGMP), leading to a rapid decline in intracellular cGMP level and to the closure of cGMP-gated cation channels located in the plasma membranes of the outer segment. Channel closure leads to membrane hyperpolarization and decreases synaptic release of glutamate in the OPL. The reduction of glutamate concentration is sensed by dendrites of two types of bipolar cells. In hyperpolarizing bipolar cells (HBCs) ionotropic glutamate receptors are expressed and the cell becomes hyperpolarized in response to the light-induced decrease in OPL glutamate level. In contrast, mGluR6 is expressed in DBCs, and light exposure causes the cell to depolarize. An alternatively spliced form of Gαo, Gαo1, is necessary for DBC-derived ERG b-wave responses.3,4 It is known that activation of mGluR6 by glutamate leads to the closure of a cation channel,5 which has recently been suggested to be a transient receptor potential–like channel, TRPM1.6 Akin to the knockout mouse deficient in mGluR6 or Gαo,4,7 the TRPM1-knockout mouse lacks the ERG b-wave.6 Unlike the phototransduction cascade, many components involved in the mGluR6 signaling pathway remain unidentified. 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: Gβ5-L (long-splice form of the fifth member of the G-protein β subunit) and RGS9 anchoring protein (R9AP).14,15 Together, these three proteins form the transducin GTPase acceleration protein (GAP) complex, the level of which determines the duration of rhodopsin signaling in rods.16 The stability of the transducin GAP complex depends on a unique interaction between Gβ5-L and the G protein γ–like (GGL) domain of RGS9-1.12,17 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 Gβ5 (Gβ5-S), which is more broadly expressed in the nervous system.18 In Gβ5 knockout (Gβ5−/−) mice, all four R7 RGS proteins are destabilized, and the ERG b-wave is absent.8,19 In addition to the no-b-wave ERG phenotype, DBCs of Gβ5−/− mice exhibit unique dendritic defects as the result of an arrest in the formation of the triadic ribbon synapses during OPL development. Transgenic restoration of Gβ5-L in rods of adult Gβ5−/− mice rescued neither the OPL morphologic defects nor the absent ERG b-wave, indicating that both phenotypes come from the loss of Gβ5-S downstream of the photoreceptors.8 However, it remains uncertain whether a signaling defect in the mGluR6 signaling pathway or the reduced triadic ribbon synapses at the OPL, or both, causes the absence of ERG b-wave in Gβ5−/− mice. To gain further insight, we recorded ERG responses from mice carrying targeted mutations in both RGS7 and -11 genes. We found that RGS7 and -11 are co-localized at the tips of DBC dendrites and that both are involved in the generation of ERG b-waves in a functionally redundant manner. Because of the presence of a robust, though delayed ERG b-wave response when both RGS7 and -11 are mutated, our data suggest that OPL morphologic defects, rather than the prolonged mGluR6/Gαo1 signal transduction in DBCs, are the major contributing factor to the loss of ERG b-waves in Gβ5−/− mice.