Ca 2+ ‐dependent Regulation of Phototransduction †

2؉switch helix changes solvent accessibility of Thr-171 and Leu-174 that affects the domain interface. Although the Ca 2؉ switch helix is not part of the RetGC1 binding site, insertion of an extra Gly residue between Ser-173 and Leu-174 as well as deletion of Arg-172, Ser-173, or Leu-174 all caused a decrease in Ca 2؉ binding affinity and abolished RetGC1 activation. We conclude that Ca 2؉ -dependent conformational changes in the Ca 2؉ switch helix are important for activating RetGC1 and provide further support for a Ca 2؉ -myristoyl tug mechanism.Guanylyl cyclase activating proteins (GCAPs) 2 belong to the neuronal calcium sensor (NCS) branch of the calmodulin superfamily (1-3) and regulate Ca 2ϩ -sensitive activity of retinal guanylyl cyclase (RetGC) in rod and cone cells (4 -6). Phototransduction in retinal rods and cones is modulated by intracellular Ca 2ϩ sensed by GCAPs (7,8), and defects in Ca 2ϩ signaling by GCAPs are linked to retinal diseases (9). GCAP proteins in the Ca 2ϩ -free/Mg 2ϩ -bound state activate RetGC (10), whereas Ca 2ϩ -bound GCAPs inhibit RetGC at high Ca 2ϩ levels maintained in the dark (11-13).The GCAPs (GCAP1 (6), GCAP2 (14), GCAP3 (15), and GCAP4 -8 (16) are all ϳ200-amino acid residue proteins containing a covalently attached N-terminal myristoyl group and four EF-hand motifs (EF1 through EF4; Fig. 1 RetGC (10,18,19). The x-ray crystal structure of Ca 2ϩ -bound GCAP1 (20) and NMR structure of GCAP2 (21) showed that the four EF-hands form two semiglobular domains (EF1 and EF2 in the N-domain and EF3 and EF4 in the C-domain); Ca 2ϩ is bound at EF2, EF3, and EF4, and the N-terminal myristoyl group in GCAP1 is buried inside the Ca 2ϩ -bound protein flanked by hydrophobic residues at the N and C termini (see the red residues in Fig. 1 Experimental ProceduresExpression and Purification of GCAP1 and Mutants-Mutations were introduced in a bovine GCAP1 coding plasmid using a "splice by overlap extension" approach as previously described (24). Myristoylated GCAP1 and its mutants were produced in Escherichia coli strain harboring yeast N-myristoyl

mentioning

confidence: 99%

Structure of Guanylyl Cyclase Activator Protein 1 (GCAP1) Mutant V77E in a Ca2+-free/Mg2+-bound Activator State

Lim

Peshenko

Olshevskaya

et al. 2016

“…This closes the cGMP-gated cation channels (CNG) in the plasma membrane (PM) and hyperpolarizes the membrane (16 -18). The role of GC1 in cGMP production is crucial for the recovery of phototransduction (19).…”

mentioning

confidence: 99%

“…The most likely explanation for this finding is that the structure of GCAP1 varies highly under different conditions. GCAP1's binding with its partners occurs transiently under certain fluctuating ionic conditions (19,30) known to occur during phototransduction. Mg 2ϩ and Ca 2ϩ levels significantly impact GC1's activation by GCAP1 (28).…”

mentioning

confidence: 99%

Impaired Association of Retinal Degeneration-3 with Guanylate Cyclase-1 and Guanylate Cyclase-activating Protein-1 Leads to Leber Congenital Amaurosis-1

Zulliger

Naash

Rajala

et al. 2015

Background: Defects in the function of guanylate cyclase 1 (GC1) cause Leber congenital amaurosis (LCA) type 1. Results: GC1, GCAP1, and RD3 form a complex in the endoplasmic reticulum that targets GC1 to outer segments. Conclusion: A subset of LCA1 is caused by impaired formation of the RD3-GC1-GCAP1 complex. Significance: Understanding the molecular interaction of RD3 with GC1 and GCAP1 has potential therapeutic benefits for LCA1.

“…This hydrolysis is catalyzed by a triumvirate complex of proteins consisting of RGS9-1, G␤5-L, and R9AP (the "RGS9 complex") (9). Finally, cGMP is restored through the action of guanylate cyclase (GC) (10).…”

Section: Phototransduction: Rhodopsin Activation Amplification and mentioning

confidence: 99%

“…The levels of Ca 2ϩ fall in light because its influx is reduced when cGMP-gated channels close, whereas Ca 2ϩ efflux via the Ca 2ϩ /K ϩ /Na ϩ exchanger continues. The reduction in intracellular Ca 2ϩ is sensed by several different Ca 2ϩ -binding proteins, including GC-activating proteins (GCAPs), which stimulate cGMP synthesis by GC when Ca 2ϩ falls (10). The Ca 2ϩ /GCAP-dependent regulation of GC activity forms a powerful feedback mechanism in which the rate of cGMP synthesis increases as Ca 2ϩ falls during the response to light, helping to restore cGMP levels rapidly and allowing the cGMP channels to reopen.…”

Section: Light Adaptation: Role Of Calciummentioning

confidence: 99%

Photoreceptor Signaling: Supporting Vision across a Wide Range of Light Intensities

Arshavsky

Burns

2012

178

169

For decades, photoreceptors have been an outstanding model system for elucidating basic principles in sensory transduction and biochemistry and for understanding many facets of neuronal cell biology. In recent years, new knowledge of the kinetics of signaling and the large-scale movements of proteins underlying signaling has led to a deeper appreciation of the photoreceptor's unique challenge in mediating the first steps in vision over a wide range of light intensities. First Steps in Vision Occur in Photoreceptor Outer SegmentsRetinal photoreceptors transduce information obtained in the form of absorbed photons into an electrical response that can be relayed across synapses to other neurons in the retina. In vertebrate photoreceptors, photon absorption and visual signaling take place in the outer segment (Fig. 1), a sensory cilium tightly packed with stacks of membranous discs containing extremely high densities of visual pigments and other signaling proteins. This morphological arrangement allows photons to be efficiently absorbed as they pass through the outer segment. The signal from activated visual pigment (rhodopsin in rods or cone opsins in cones) must then be sufficiently amplified to generate an electrical response that overcomes intrinsic noise. Transduction from the light-absorbing visual pigment into an electrical response utilizes a G protein signaling pathway termed the phototransduction cascade, which leads to a decrease in the second messenger cGMP and the closure of cGMP-sensitive cation channels. The resulting hyperpolarization transiently decreases the release of glutamate from the photoreceptor synaptic terminals, signaling the number of absorbed photons to the rest of the visual system. Remarkably, photoreceptors both detect low light levels (single photons in the case of rods) and continue to rapidly and reliably signal changes in light intensity as illuminance increases over 10 orders of magnitude during the course of a typical day. Phototransduction: Rhodopsin Activation, Amplification, and DeactivationPhototransduction has been the subject of many comprehensive reviews (1-4). Here, we provide a framework introduction and briefly summarize the latest findings that are shaping our understanding of this first step in vision.Phototransduction begins when a photon causes cis-transisomerization of the chromophore 11-cis-retinal, which induces a rapid conformational change to the protein's fully active form, R*. R* activates molecules of the G protein transducin by catalyzing GDP/GTP exchange on the transducin ␣-subunit, G␣ t (Fig. 1). G␣ t ⅐GTP separates from the transducin ␤␥-subunits and binds to the ␥-subunit of its effector, cGMP phosphodiesterase (PDE), 3 which releases this subunit's inhibitory constraint on the catalytic ␣-and ␤-subunits of PDE. Activated PDE rapidly hydrolyzes cGMP, thereby reducing its concentration in the cytoplasm and causing cGMP-sensitive cation channels in the plasma membrane to close. The closure of channels reduces the inward cation current, resulting in a tra...