GTPase Regulators and Photoresponses in Cones of the Eastern Chipmunk

Zhang, Xue; Wensel, Theodore G.; Kraft, Timothy W.

doi:10.1523/jneurosci.23-04-01287.2003

Cited by 86 publications

(103 citation statements)

References 62 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Because of higher temperatures and lower concentrations of chloride ion (an inhibitor of GTP hydrolysis), GTP hydrolysis kinetics likely are more rapid than those observed under our conditions. However, if GTP hydrolysis in rods were 10-fold faster than our results suggest, it would be hard to understand why mammalian cones, which recover less than 10-fold more rapidly than rods, require 10 times higher levels of the RGS9-1⅐G␤ 5 complex (24,40,41). Thus, our estimate of maximal RGS9-1 GAP activity, when appropriately corrected for differences in conditions, seems likely to reflect the value attained in intact rod cells.…”

Section: Resultsmentioning

confidence: 72%

Activation of RGS9-1GTPase Acceleration by Its Membrane Anchor, R9AP

Zhang

Wensel

2003

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

The GTPase-accelerating protein (GAP) complex RGS9-1⅐G␤ 5 plays an important role in the kinetics of light responses by accelerating the GTP hydrolysis of G␣ t in vertebrate photoreceptors. Much, but not all, of this complex is tethered to disk membranes by the transmembrane protein R9AP. To determine the effect of the R9AP membrane complex on GAP activity, we purified recombinant R9AP and reconstituted it into lipid vesicles along with the photon receptor rhodopsin. Full-length RGS9-1⅐G␤ 5 bound to R9AP-containing vesicles with high affinity (K d < 10 nM), but constructs lacking the DEP (dishevelled/EGL-10/pleckstrin) domain bound with much lower affinity, and binding of those lacking the entire N-terminal domain (i.e. the dishevelled/EGL-10/pleckstrin domain plus intervening domain) was not detectable. Formation of the membranebound complex with R9AP increased RGS9-1 GAP activity by a factor of 4. Vesicle titrations revealed that on the time scale of phototransduction, the entire reaction sequence from GTP uptake to GAP-catalyzed hydrolysis is a membrane-delimited process, and exchange of G␣ t between membrane surfaces is much slower than hydrolysis. Because in rod cells different pools exist of RGS9-1⅐G␤ 5 that are either associated with R9AP or not, regulation of the association between R9AP and RGS9-1⅐G␤ 5 represents a potential mechanism for the regulation of recovery kinetics.Timely deactivation of G protein ␣ subunits is a key element of responses to the stimulation of G protein-coupled receptors. It plays an especially important role in fast cellular responses such as those of vertebrate photoreceptors. In the rod and cone cell outer segments, the recovery phase of light responses depends on the presence of a GTPase-accelerating protein (GAP) 1 complex RGS9-1⅐G␤ 5 (1-4). Whether and how the GAP activity of this complex is regulated is unknown.RGS9-1 contains multiple functional domains, including an RGS domain that is responsible for its GAP activity (3), a G protein ␥ subunit-like domain for G␤ 5L binding (4,5), an Nterminal domain that includes a DEP (dishevelled/EGL-10/ pleckstrin) domain (6) and an intermediate domain (7), and a C-terminal domain that is unique to RGS9-1 among all of the RGS proteins (8). All of these domains have been found to participate in the regulation of GAP activity and substrate specificity (9 -11). The inhibitory PDE␥ subunit of the effector regulated by G␣ t , cGMP phosphodiesterase (PDE6), interacts with both G␣ t and the catalytic core of RGS9-1 and enhances RGS9-1 GAP activity (12-14) in vitro, but it is not clear how this enhancement is accomplished in a physiological setting in which tight PDE␥ binding to PDE6 catalytic subunits blocks GAP enhancement (15-18).Additional possible mechanisms for regulation include a light-and calcium-regulated phosphorylation (19, 20) of RGS9-1 and interactions with the recently discovered membrane anchor protein, R9AP (7, 21). R9AP is a 25-kDa protein that is selectively expressed in photoreceptor outer segments. Homologues are apparen...

show abstract

Section: Resultsmentioning

confidence: 72%

Activation of RGS9-1GTPase Acceleration by Its Membrane Anchor, R9AP

Zhang

Wensel

2003

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

show abstract

“…An interesting candidate is the cGMP phosphodiesterase, which may span the interdiscal space (Fig. 4 C and D) and is present at 1:310 relative to rhodopsin (52), so that it has a surface density on pairs of discs of 250 μm −2 , intriguingly close to that of the as-yet unidentified "spacers" (49).…”

Section: Estimating the Spring Constant Of An Elementary Rod Os Elasticmentioning

confidence: 99%

In vivo optophysiology reveals that G-protein activation triggers osmotic swelling and increased light scattering of rod photoreceptors

Zhang

Zawadzki

Goswami

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

111

164

View full text Add to dashboard Cite

The light responses of rod and cone photoreceptors have been studied electrophysiologically for decades, largely with ex vivo approaches that disrupt the photoreceptors' subretinal microenvironment. Here we report the use of optical coherence tomography (OCT) to measure light-driven signals of rod photoreceptors in vivo. Visible light stimulation over a 200-fold intensity range caused correlated rod outer segment (OS) elongation and increased light scattering in wildtype mice, but not in mice lacking the rod G-protein alpha subunit, transducin (Gα t ), revealing these responses to be triggered by phototransduction. For stimuli that photoactivated one rhodopsin per Gα t the rod OS swelling response reached a saturated elongation of 10.0 ± 2.1%, at a maximum rate of 0.11% s −1. Analyzing swelling as osmotically driven water influx, we find the H 2 O membrane permeability of the rod OS to be (2.6 ± 0.4) × 10 −5 cm·s, comparable to that of other cells lacking aquaporin expression. Application of Van't Hoff's law reveals that complete activation of phototransduction generates a potentially harmful 20% increase in OS osmotic pressure. The increased backscattering from the base of the OS is explained by a model combining cytoplasmic swelling, translocation of dissociated G-protein subunits from the disc membranes into the cytoplasm, and a relatively higher H 2 O permeability of nascent discs in the basal rod OS. Translocation of phototransduction components out of the OS may protect rods from osmotic stress, which could be especially harmful in disease conditions that affect rod OS structural integrity.osmotic stress | phototransduction | optical coherence tomography | intrinsic optical signals | photoreceptor waveguiding

show abstract

“…One argument is that rods have ϳ60-fold less RGS9 than transducin (Zhang et al, 2003). However, the strongest evidence comes from electrophysiological studies.…”

Section: Transducin Translocation Takes Place In Three Major Stepsmentioning

confidence: 99%

Transducin Translocation in Rods Is Triggered by Saturation of the GTPase-Activating Complex

Lobanova

Finkelstein²,

Song³

et al. 2007

J. Neurosci.

121

View full text Add to dashboard Cite

Light causes massive translocation of G-protein transducin from the light-sensitive outer segment compartment of the rod photoreceptor cell. Remarkably, significant translocation is observed only when the light intensity exceeds a critical threshold level. We addressed the nature of this threshold using a series of mutant mice and found that the threshold can be shifted to either a lower or higher light intensity, dependent on whether the ability of the GTPase-activating complex to inactivate GTP-bound transducin is decreased or increased. We also demonstrated that the threshold is not dependent on cellular signaling downstream from transducin. Finally, we showed that the extent of transducin ␣ subunit translocation is affected by the hydrophobicity of its acyl modification. This implies that interactions with membranes impose a limitation on transducin translocation. Our data suggest that transducin translocation is triggered when the cell exhausts its capacity to activate transducin GTPase, and a portion of transducin remains active for a sufficient time to dissociate from membranes and to escape from the outer segment. Overall, the threshold marks the switch of the rod from the highly light-sensitive mode of operation required under limited lighting conditions to the less-sensitive energy-saving mode beneficial in bright light, when vision is dominated by cones.

show abstract

GTPase Regulators and Photoresponses in Cones of the Eastern Chipmunk

Cited by 86 publications

References 62 publications

Activation of RGS9-1GTPase Acceleration by Its Membrane Anchor, R9AP

Activation of RGS9-1GTPase Acceleration by Its Membrane Anchor, R9AP

In vivo optophysiology reveals that G-protein activation triggers osmotic swelling and increased light scattering of rod photoreceptors

Transducin Translocation in Rods Is Triggered by Saturation of the GTPase-Activating Complex

Contact Info

Product

Resources

About