vated transducin a subunits bind GTP and stimulate 
a cGMP phosphodiesterase. The ensuing cGMP hy- 
drolysis shuts down cGMP-gated cation channels 
within the photoreceptor outer-segment plasma 
membrane, blocking Na^ and Ca^"^ influx and hyper- 
polarizing the cell. Recovery occurs as transducin 
deactivates itself by hydrolyzing its bound GTP. De- 
pletion of intracellular Ca^^ by a Na/Ca exchanger 
also contributes to recovery by stimulating a Ca^^- 
sensitive guanylate cyclase to resynthesize cGMP. 
Dr. Hurley and his colleagues are unraveling the 
molecular mechanisms of photoreceptor recovery 
and adaptation. In collaboration with Drs. Melvin 
Simon and Denis Baylor, transgenic mice that ex- 
press a GTPase-deficient form of transducin a sub- 
unit in their rod photoreceptors were produced and 
analyzed. Mutant transducin accumulates in the 
outer segments of the transgenic rod cells at con- 
centrations up to sixfold higher than endogenous 
transducin. To compensate for the presence of the 
mutant transducin, the rods respond by specifically 
and almost completely eliminating catalytic sub- 
units of the transducin target, cGMP phospho- 
diesterase. Other phototransduction enzymes are 
unaffected, including rhodopsin, transducin /3, 
phosphodiesterase 7, recoverin, arrestin, and phos- 
ducin. The altered rod cells show no signs of degen- 
erating, but their electrical responses are slow and 
severely desensitized. These results show that the 
presence of persistently activated transducin stimu- 
lates a powerful adaptive feedback mechanism that 
specifically down-regulates its target enzyme. 
During recovery cGMP is resynthesized by guany- 
late cyclase as the concentration of intracellular 
Ca^^ is lowered within the submicromolar range fol- 
lowing photoexcitation. Dr. Hurley and his col- 
leagues showed that recoverin, a 23-kDa Ca'^^-bind- 
ing protein, stimulates a photoreceptor membrane 
guanylate cyclase only when the concentration of 
free Ca'^^ is <200 nM. Recoverin was purified and 
examined for post-translational modifications by 
electrospray mass spectrometry. Its amino terminus 
was found to be heterogeneously acylated by one of 
four different types of short-chain fatty acid, CI 4:0, 
C14:1, Cl4:2, and CI 2:0. Similar heterogeneous 
acylation was also detected at the amino terminus of 
the transducin a subunit. 
Dr. Hurley and his colleagues are investigating the 
functional role of heterogeneous amino-terminal ac- 
ylation of recoverin and transducin. They found that 
acylated recoverin binds to phospholipid mem- 
branes in the presence of >1 ixM Ca^^, but nonacyl- 
ated recombinant recoverin does not interact with 
membranes. The recoverin-membrane interaction 
relies on the binding of Ca^^, which exposes the 
acylated amino terminus of recoverin and allows the 
fatty acid to serve as a membrane anchor. 
Recoverin activates membrane guanylate cyclase 
at Ca^^ concentrations <200 nM but dissociates 
from membranes under these same conditions. 
These two observations suggest that recoverin does 
not activate the cyclase directly. Dr. Hurley and his 
colleagues are investigating the possibility that ad- 
ditional factors participate in recoverin-mediated 
guanylate cyclase activation. 
Drosophila Phototransduction 
The mechanisms of phototransduction are quite 
different for invertebrates and vertebrates. In Dro- 
sophila photoreceptors, light stimulates a GTP- 
sensitive phospholipase C, which is essential for 
generating a photoresponse. Dr. Hurley and his col- 
leagues have identified a unique Drosophila G pro- 
tein /3 subunit, GBE, that may participate in Dro- 
sophila phototransduction. In situ hybridization 
and immunocytochemical analyses reveal that GBE 
mRNA and protein are photoreceptor specific. 
The role of G protein /? subunits in signal trans- 
duction is not well understood. In addition to pre- 
senting G protein a subunits to their receptors, 
subunits may also regulate other effector enzymes. 
Dr. Hurley and his colleagues are investigating the 
role of GBE in Drosophila phototransduction. In 
collaboration with Dr. Charles Zuker (HHMI, Uni- 
versity of California, San Diego), the laboratory has 
produced mutant Drosophila that are deficient in 
GBE. In situ biochemical analyses of these mutants 
reveal that GBE is essential for light-stimulated bind- 
ing of GTP to a protein in Drosophila photorecep- 
tors. Other enzymatic assays and electrophysiologi- 
cal analyses that may reveal the role of the G protein 
/3 subunit in Drosophila phototransduction are in 
progress. 
Dr. Hurley is also Associate Professor of Bio- 
chemistry at the University of Washington School 
of Medicine, Seattle. 
Articles 
Curcio, C.A., Allen, K.A., Sloan, K.R., Lerea, C.L., 
Hurley, J.B., Klock, I.B., and Milam, A.H. 1991. 
Distribution and morphology of human cone pho- 
toreceptors stained with anti-blue opsin. / Comp 
Neurol 512:610-624. 
Dizhoor, A.M., Ericsson, L.H., Johnson, R.S., Ku- 
mar, S., Olshevskaya, E., Zozulya, S., Neubert, 
T.A., Stryer, L., Hurley, J.B., and Walsh, K.A. 
1992. The NH2 terminus of retinal recoverin is 
acylated by a small family of fatty acids. / Biol 
Chem 267:16033-16036. 
NEUROSCIENCE 407 
