beta gamma subunits of heterotrimeric G proteins can exist in two distinct conformations. In the R (relaxed) state, corresponding to the structure of the free beta gamma or the structure of beta gamma in the alpha beta gamma heterotrimer, the hydrophobic farnesyl moiety of the gamma subunit is exposed, thereby mediating membrane association. In the T (tense) state, as observed in the phosducin-Gt beta gamma structure, the farnesyl moiety of the gamma subunit is effectively buried in the cavity formed between blades 6 and 7 of the beta subunit. Binding of phosducin to Gt beta gamma induces the formation of this cavity, resulting in a switch from the R to the T conformation. This sequesters beta gamma from the membrane to the cytosol and turns off the signal-transduction cascade. Regulation of this membrane association/dissociation switch of Gt beta gamma by phosducin may be a general mechanism for attenuation of G protein coupled signal transduction cascades.

Abstract. The stimulation of meiotic maturation of starfish oocytes by the hormone 1-methyladenine is mimicked by injection of 8)' subunits of G-proteins from either retina or brain. Conversely, the hormone response is inhibited by injection of the GDP-bound forms of O/il or O/t subunits, or by injection of phosducin; all of these proteins should bind free/33'. t~-subunit forms with reduced affinity for 87 (O/il or O/t bound to hydrolysis-resistant GTP analogs, or t~irGMPPCP treated with trypsin to remove the amino terminus of the protein) are less effective inhibitors of 1-methyladenine action. These results indicate that the /33, subunit of a G-protein mediates 1-methyladenine stimulation of oocyte maturation.

Phototransduction in retinal rods involves a G protein-coupled signaling cascade that leads to cGMP hydrolysis and the closure of cGMP-gated cation channels that are open in darkness, producing a membrane hyperpolarization as the light response. For many years there have also been reports of the presence of a phosphoinositide pathway in the rod outer segment, though its functions and the molecular identities of its components are still unclear. Using immunocytochemistry with antibodies against various phosphoinositide-specific phospholipase C (PLC) isozymes (␤1-4, ␥1-2, and ␦1-2), we have found PLC␤4-like immunoreactivity in rod outer segments. Similar experiments with antibodies against the ␣-subunits of the G q family of G proteins, which are known to activate PLC␤4, have also demonstrated G ␣11 -like immunoreactivity in this location. Immunoblots of total proteins from whole retina or partially purified rod outer segments with anti-PLC␤4 and anti-G ␣11 antibodies gave, respectively, a single protein band of the expected molecular mass, suggesting specific labelings. The retinal locations of the two proteins were also supported by in situ hybridization experiments on mouse retina with probes specific for the corresponding mouse genes. These two proteins, or immunologically identical isoforms, therefore likely mediate the phosphoinositide signaling pathway in the rod outer segment. At present, G ␣11 or a G ␣11 -like protein represents the only G protein besides transducin (which mediates phototransduction) identified so far in the rod outer segment. Although absent in the outer segment layer, other PLC isoforms as well as G ␣q (another G q family member), are present elsewhere in the retina.Visual transduction in the retina takes place in the outer segments of rod and cone photoreceptors and is known to involve a cGMP signaling cascade (see ref. 1 for a recent review). In this process, light isomerizes the visual pigment into an active form, which, via the G protein transducin, stimulates a cGMP-phosphodiesterase to lead to cGMP hydrolysis. In darkness, cytoplasmic cGMP binds to and opens cGMP-activated cation channels on the plasma membrane of the outer segment. These open channels sustain an inward dark current to keep the cell partially depolarized and maintain a steady release of glutamate from the synaptic terminal of the photoreceptor. In the light, the hydrolysis of cGMP leads to the closure of these channels, producing a membrane hyperpolarization as the light response and reducing the glutamate release from the cell.Over the years, however, there have also been reports of a phosphoinositide signaling pathway in the rod outer segment. In this pathway, the membrane phospholipid phosphatidylinositol-4,5-bisphosphate is hydrolyzed by phospholipase C (PLC) enzymes to release the second messengers inositol-1,4,5-trisphosphate and diacylglycerol, with the former leading to intracellular Ca 2ϩ release and the latter activating the enzyme protein kinase C (PKC). In rod outer segment preparations, phosph...