Impairment of the Rod Outer Segment Membrane Guanylate Cyclase Dimerization in A Cone−Rod Dystrophy Results in Defective Calcium Signaling

Abstract

Rod outer segment membrane guanylate cyclase1 (ROS-GC1) is the original member of the membrane guanylate cyclase subfamily whose distinctive feature is that it transduces diverse intracellularly generated Ca 2+ signals in the sensory neurons. In the vertebrate retinal neurons, ROS-GC1 is pivotal for the operations of phototransduction and, most likely, of the synaptic activity. The phototransduction-and the synapse-linked domains are separate, and they are located in the intracellular region of ROS-GC1. These domains sense Ca 2+ signals via Ca 2+ -binding proteins. These proteins are ROS-GC activating proteins, GCAPs. GCAPs control ROS-GC1 activity through two opposing regulatory modes. In one mode, at nanomolar concentrations of Ca 2+ , the GCAPs activate the cyclase and as the Ca 2+ concentrations rise, the cyclase is progressively inhibited. This mode operates in phototransduction via two GCAPs: 1 and 2. The second mode occurs at micromolar concentrations of Ca 2+ via S100 . Here, the rise of Ca 2+ concentrations progressively stimulates the enzyme. This mode is linked with the retinal synaptic activity. In both modes, the final step in Ca 2+ signal transduction involves ROS-GC dimerization, which causes the cyclase activation. The identity of the dimerization domain is not known. A heterozygous, triple mutation -E786D, R787C, T788M-in ROS-GC1 has been connected with autosomal cone-rod dystrophy in a British family. The present study shows the biochemical consequences of this mutation on the phototransduction-and the synapse-linked components of the cyclase. (1) It severely damages the intrinsic cyclase activity. (2) It significantly raises the GCAP1-and GCAP2-dependent maximal velocity of the cyclase, but this compensation, however, is not sufficient to override the basal cyclase activity. (3) It converts the cyclase into a form that only marginally responds to S100 . The mutant produces insufficient amounts of the cyclic GMP needed to drive the machinery of phototransduction and of the retinal synapse at an optimum level. The underlying cause of the breakdown of both types of machinery is that, in contrast to the native ROS-GC1, the mutant cyclase is unable to change from its monomeric to the dimeric form, the form required for the functional integrity of the enzyme. The study defines the CORD in molecular terms, at a most basic level identifies a region that is critical in its dimer formation, and, thus, discloses a single unifying mechanistic theme underlying the complex pathology of the disease.Phototransduction is a biochemical process by which the vertebrate rods and cones generate electrical signals in response to captured photons (reviewed in refs 1 and 2). This process occurs in the rod (or cone) outer segments (ROS). 1The photon induces a decline in the level of cyclic GMP and closure of the cyclic GMP-gated cation channels. This results in a drop of intracellular Ca 2+ from ∼500 nM in darkadapted photoreceptor cell to below 100 nM after illumination. This drop activates ROS guanylate cyc...

Interactions within the Coiled-coil Domain of RetGC-1 Guanylyl Cyclase Are Optimized for Regulation Rather than for High Affinity

“…A (62,63), we initially found that the "ERT" mutation linked to ad-CORD causes reduced catalytic activity when the mutant polypeptide is expressed alone. But our subsequent analyses of this and other adCORD mutants showed that the effect on intrinsic activity is not sufficient to explain the disease phenotype.…”

Dysfunction of outer segment guanylate cyclase caused by retinal disease related mutations

“…However, disease causing mutations can also point to critical amino acid positions that determine structure-function relationships in natively folded protein structures and therefore might help to gain mechanistic insights of protein function and regulation (Duda et al, 1999a, 2000; Ramamurthy et al, 2001). For example, a biochemical study on mutations in the so-called dimerization domain of ROS-GC1 correlating with cone-rod dystrophy recently revealed that this dimerization or linker domain operates as a Ca 2 + -sensitive control switch module (Zägel et al, 2013).…”

Membrane guanylate cyclase, a multimodal transduction machine: history, present, and future directions

“…One example is ROS-GC1-E 786 D, R 787 C, T 788 M mutation. The mutation deranges the dimer formation of the guanylate cyclase and reduces the basal catalytic activity of the ROS-GC (Duda et al, 1999a, 2000; Tucker et al, 1999). This basic structural derangement in the guanylate cyclase, intriguingly, increases its sensitivity to GCAP1 and GCAP2.…”