Abstract
Guanylate cyclase-activating protein 1 (GCAP1) is involved in the shutdown of the phototransduction cascade by regulating the enzymatic activity of retinal guanylate cyclase via a Ca2+/cGMP negative feedback. While the phototransduction-associated role of GCAP1 in the photoreceptor outer segment is widely established, its implication in synaptic transmission to downstream neurons remains to be clarified. Here, we present clinical and biochemical data on a novel isolate GCAP1 variant leading to a double amino acid substitution (p.N104K and p.G105R) and associated with cone dystrophy (COD) with an unusual phenotype. Severe alterations of the electroretinogram were observed under both scotopic and photopic conditions, with a negative pattern and abnormally attenuated b-wave component. The biochemical and biophysical analysis of the heterologously expressed N104K-G105R variant corroborated by molecular dynamics simulations highlighted a severely compromised Ca2+-sensitivity, accompanied by minor structural and stability alterations. Such differences reflected on the dysregulation of both guanylate cyclase isoforms (RetGC1 and RetGC2), resulting in the constitutive activation of both enzymes at physiological levels of Ca2+. As observed with other GCAP1-associated COD, perturbation of the homeostasis of Ca2+ and cGMP may lead to the toxic accumulation of second messengers, ultimately triggering cell death. However, the abnormal electroretinogram recorded in this patient also suggested that the dysregulation of the GCAP1–cyclase complex further propagates to the synaptic terminal, thereby altering the ON-pathway related to the b-wave generation. In conclusion, the pathological phenotype may rise from a combination of second messengers’ accumulation and dysfunctional synaptic communication with bipolar cells, whose molecular mechanisms remain to be clarified.
Highlights
The signaling machinery underlying the phototransduction cascade is finely regulated by Ca2+ and cyclic guanosine monophosphate, which constitute strictly interconnected second messengers [1,2]
We propose that an alteration of the GC1–guanylate cyclase-activating protein 1 (GCAP1) transduction unit exists at the synaptic terminal as well as in the photoreceptor outer segment, and such alteration may be related with the abnormal synaptic transmission between photoreceptors and bipolar cells
The presence of ribbon synapses in photoreceptors distinguishes these cells from other neurons because it provides the above features by encoding graded changes of membrane potential induced by light into the modulation of continuous vesicle exocytosis, which permits the transmission of signals to the inner retina [41]
Summary
The signaling machinery underlying the phototransduction cascade is finely regulated by Ca2+ and cyclic guanosine monophosphate (cGMP), which constitute strictly interconnected second messengers [1,2]. The light-induced activation of phosphodiesterase 6 catalyzes the hydrolysis of cGMP, which causes cGMP-gated channels (CNG) to transiently close and lead to a hyperpolarization of the cell membrane which is sensed at the photoreceptor synaptic terminal. Mg2+ -GCAP1 acquires a conformation that stimulates the synthesis of cGMP by GC to rapidly restore dark-adapted cell conditions by reopening the CNG channels [6,7]. Two isoforms of retinal GC have been found in photoreceptors, namely, GC1 (or RetGC-1, GC-E) and GC2 (RetGC-2, GC-F), the latter produces less than 30% of cGMP in murine retina [8] and is 25-fold less abundant than GC1 in bovine rod outer segments [9]
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