Retinal degeneration-3 protein attenuates photoreceptor degeneration in transgenic mice expressing dominant mutation of human retinal guanylyl cyclase

Peshenko, Igor V.; Dizhoor, Alexander M.

doi:10.1016/j.jbc.2021.101201

Cited by 6 publications

(6 citation statements)

References 49 publications

(111 reference statements)

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“…GC1 activity assays in the presence of RD3 (Figure 6, left panel) or RD3ppt (Figure 4A) suggest that the enzymatic activity is partially inhibited when GCAP1 variants are present. The incomplete shut down of GC1 by RD3/RD3ppt reported here or by others [26] could be explained by a combination of: (i) the interaction occurring between RD3 and GCAP1, proven to be essential for the proper trafficking of the GC1-GCAP1-RD3 assembly to the photoreceptor outer segments (POS) [27]; (ii) the competitive regulation of GC1 by RD3 and GCAP1 [19]. Here the interaction between either WT-or E111V-GCAP1 and RD3/RD3ppt was investigated in GC1-activating and inhibiting conditions by CD spectroscopy experiments which underpinned major structural variations upon the RD3(ppt)-GCAP1 complex formation potentially affecting GC1 regulation and preventing rigid-body docking simulations from determining a unique binding pose.…”

Section: Introductionsupporting

confidence: 68%

Supramolecular complexes of GCAP1: towards the development of effective biologics for inherited retinal dystrophies

Biasi,

Marino,

Dal Cortivo

et al. 2024

Preprint

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Guanylate Cyclase Activating Protein 1 (GCAP1) is a neuronal Ca2+-sensor protein expressed in photoreceptors where it regulates the enzymatic activity of retinal Guanylate Cyclase 1 (GC1) in a Ca2+-dependent manner. Recently, over 20 missense mutations in GUCA1A (encoding for GCAP1) have been associated with inherited autosomal dominant retinal diseases, namely cone dystrophy (COD) and cone-rod dystrophy (CORD). Since GCAP1 is known to be a functional dimer, COD/CORD patients present a heterogeneous pool of GCAP1 assemblies constituted by wild-type and mutated homodimers and heterodimers. Here, we present an integrated in silico and biochemical investigation on the effects of the E111V substitution, associated with a severe form of CORD, on GCAP1 homo- and hetero-dimerization. Despite inducing a constitutive activation of GC1 due to impaired Ca2+-binding in the high-affinity EF-hand 3 motif, the E111V substitution did not affect either the homo- or the hetero dimerization process as clearly highlighted by aSEC and molecular docking experiments. Indeed, both variants exhibited micromolar monomer-dimer equilibrium constants in the presence of both Mg2+ and Ca2++, indicating that at physiological cellular concentrations both variants are predominantly monomers under Ca2+-loaded and, to a lesser extent, Mg2+-loaded conditions. Molecular docking and dynamics simulations confirmed chromatographic results highlighting slight alterations in free energy of binding involving the pathogenic E111V variant in the Ca2+-bound state and increased mobility over time affecting the Ca2+-coordinating EF3 motif. In addition, to evaluate possible therapeutic approaches, the regulation of the catalytic activity of GC1 by WT and E111V-GCAP1 was studied in the presence of retinal degeneration protein 3 (RD3), an α-helical protein that strongly inhibits GC1, and a RD3-derived peptide (RD3ppt) which encompasses a region of RD3 that is essential for its inhibitory activity. GC1 activity assays in the presence of RD3ppt suggest that the enzymatic activity is partially inhibited by the peptide at low micromolar concentrations when GCAP1 variants are present. The incomplete shut down of GC1 by RD3 could be explained by the interaction occurring between RD3 and GCAP1, known to form a complex with GC1 in the endoplasmic reticulum. This fundamental interaction was here investigated spectroscopically and in silico, unveiling major structural rearrangements upon complex formation. Interestingly, the full RD3 protein was able to better modulate GC1 activity and restore the abnormal cGMP production induced by the pathogenic E111V-GCAP1 variant to a physiological level.

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Section: Introductionsupporting

confidence: 68%

Supramolecular complexes of GCAP1: towards the development of effective biologics for inherited retinal dystrophies

Biasi,

Marino,

Dal Cortivo

et al. 2024

Preprint

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“… 7 – 11 However, beyond phototransduction, it remains largely unclear whether the OS compartment plays unidentified functions. 34 – 38 In our present study, we used untargeted metabolomics and Cep250 KO mice to determine that cilia can transport the metabolic-related protein ARG1 to the OS. Through experiments using an AAV-based knockdown of Arg1, we demonstrated that normal expression of Arg1 is essential for the survival of photoreceptors.…”

Section: Discussionmentioning

confidence: 99%

Dysregulated Arginine Metabolism Is Linked to Retinal Degeneration in Cep250 Knockout Mice

Xiang,

Yang,

Xie

et al. 2023

Invest. Ophthalmol. Vis. Sci.

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Purpose Degeneration of retinal photoreceptors is frequently observed in diverse ciliopathy disorders, and photoreceptor cilium gates the molecular trafficking between the inner and the outer segment (OS). This study aims to generate a homozygous global Cep250 knockout (KO) mouse and study the resulting phenotype. Methods We used Cep250 KO mice and untargeted metabolomics to uncover potential mechanisms underlying retinal degeneration. Long-term follow-up studies using optical coherence tomography (OCT) and electroretinography (ERG) were performed. Results OCT and ERG results demonstrated gradual thinning of the outer nuclear layer (ONL) and progressive attenuation of the scotopic ERG responses in Cep250 −/− mice. More TUNEL signal was observed in the ONL of these mice. Immunostaining of selected OS proteins revealed mislocalization of these proteins in the ONL of Cep250 −/− mice. Interestingly, untargeted metabolomics analysis revealed arginine-related metabolic pathways were altered and enriched in Cep250 −/− mice. Mis-localization of a key protein in the arginine metabolism pathway, arginase 1 (ARG1), in the ONL of KO mice further supports this model. Moreover, adeno-associated virus (AAV)-based retinal knockdown of Arg1 led to similar architectural and functional alterations in wild-type retinas. Conclusions Altogether, these results suggest that dysregulated arginine metabolism contributes to retinal degeneration in Cep250 −/− mice. Our findings provide novel insights that increase understanding of retinal degeneration in ciliopathy disorders.

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“…MMF treatment significantly delays the onset of retinal degeneration, cGMP-dependent photoreceptor cytotoxicity, retinal/visual function, and microglial activation in rd10 and rd1 mice [ 124 ]. In addition, the overexpression of RD3 attenuates photoreceptor degeneration in transgenic mice expressing the human R838S RetGC1 dominant mutant [ 125 ]. The R838S RetGC1 mutation increases the enzyme’s affinity for Mg 2+ -GCAP (the activated form of GCAP) or makes it more difficult for RD3 to prevent the aberrant activation of RetGC by GCAP in the inner segment, leading to the increased synthesis of cGMP.…”

Section: Experimental Evidence Supporting the Contribution Of Elevate...mentioning

confidence: 99%

cGMP Signaling in Photoreceptor Degeneration

Yang

et al. 2023

IJMS

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Photoreceptors in the retina are highly specialized neurons with photosensitive molecules in the outer segment that transform light into chemical and electrical signals, and these signals are ultimately relayed to the visual cortex in the brain to form vision. Photoreceptors are composed of rods and cones. Rods are responsible for dim light vision, whereas cones are responsible for bright light, color vision, and visual acuity. Photoreceptors undergo progressive degeneration over time in many hereditary and age-related retinal diseases. Despite the remarkable heterogeneity of disease-causing genes, environmental factors, and pathogenesis, the progressive death of rod and cone photoreceptors ultimately leads to loss of vision/blindness. There are currently no treatments available for retinal degeneration. Cyclic guanosine 3′, 5′-monophosphate (cGMP) plays a pivotal role in phototransduction. cGMP governs the cyclic nucleotide-gated (CNG) channels on the plasma membrane of the photoreceptor outer segments, thereby regulating membrane potential and signal transmission. By gating the CNG channels, cGMP regulates cellular Ca2+ homeostasis and signal transduction. As a second messenger, cGMP activates the cGMP-dependent protein kinase G (PKG), which regulates numerous targets/cellular events. The dysregulation of cGMP signaling is observed in varieties of photoreceptor/retinal degenerative diseases. Abnormally elevated cGMP signaling interferes with various cellular events, which ultimately leads to photoreceptor degeneration. In line with this, strategies to reduce cellular cGMP signaling result in photoreceptor protection in mouse models of retinal degeneration. The potential mechanisms underlying cGMP signaling-induced photoreceptor degeneration involve the activation of PKG and impaired Ca2+ homeostasis/Ca2+ overload, resulting from overactivation of the CNG channels, as well as the subsequent activation of the downstream cellular stress/death pathways. Thus, targeting the cellular cGMP/PKG signaling and the Ca2+-regulating pathways represents a significant strategy for photoreceptor protection in retinal degenerative diseases.

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Retinal degeneration-3 protein attenuates photoreceptor degeneration in transgenic mice expressing dominant mutation of human retinal guanylyl cyclase

Cited by 6 publications

References 49 publications

Supramolecular complexes of GCAP1: towards the development of effective biologics for inherited retinal dystrophies

Supramolecular complexes of GCAP1: towards the development of effective biologics for inherited retinal dystrophies

Dysregulated Arginine Metabolism Is Linked to Retinal Degeneration in Cep250 Knockout Mice

cGMP Signaling in Photoreceptor Degeneration

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