ACTIVATION OF THE RETINAL CGMP-SPECIFIC PHOSPHODIESTERASE BY THE GDP-LOADED ALPHA-SUBUNIT OF TRANSDUCIN

The interaction of the GDP-bound form of the alpha-subunit of transduc in (T alpha(GDP)) with the cGMP-specific phosphodiesterase, the effect or enzyme in the visual system, has been studied. T alpha(GDP) is demo nstrated to be able to activate the phosphodiesterase: (a) the basal a ctivity in suspensions of dark-adapted retinal rod outer segments, exa mined in the absence of GTP, was found to be inhibited by binding of t ransducin to activated rhodopsin (Rh) and by the complex of the beta- and gamma-subunits of transducin (T beta gamma); (b) purified T alpha( GDP) is able to activate phosphodiesterase in the presence of membrane s; (c) no activation is obtained either with holotransducin (T alpha(G DP)T beta gamma) or with T alpha(GDP) in the presence of excess T beta gamma to prevent dissociation of T-GDP. The maximal level of phosphod iesterase activation reached with T alpha(GDP) (about 1500 mol cGMP/mo l phosphodiesterase(-1) s(-1)) is similar to that obtained through the 'classical' activation by T alpha(GTP), whereas the apparent affinity of T alpha(GDP) for phosphodiesterase (K-d about 50 mu M) is much low er than that of T alpha(GTP). Our data suggest that GTP hydrolysis its elf does not inactivate T alpha. The role of T beta gamma to sequester T alpha is therefore of critical importance for phosphodiesterase ina ctivation. Our results support observations on the regulation of adeny lyl cyclase by G-proteins, which suggested the ability of the free alp ha-subunits loaded with GDP to activate their effecters.