SPACR Encoded by IMPG1 Is Essential for Photoreceptor Survival by Interplaying between the Interphotoreceptor Matrix and the Retinal Pigment Epithelium.

Several pathogenic variants have been reported in the IMPG1 gene associated with the inherited retinal disorders vitelliform macular dystrophy (VMD) and retinitis pigmentosa (RP). IMPG1 and its paralog IMPG2 encode for two proteoglycans, SPACR and SPACRCAN, respectively, which are the main components of the interphotoreceptor matrix (IPM), the extracellular matrix surrounding the photoreceptor cells. To determine the role of SPACR in the pathological mechanisms leading to RP and VMD, we generated a knockout mouse model lacking Impg1, the mouse ortholog. Impg1-deficient mice show abnormal accumulation of autofluorescent deposits visible by fundus imaging and spectral-domain optical coherence tomography (SD-OCT) and attenuated electroretinogram responses from 9 months of age. Furthermore, SD-OCT of Impg1-/- mice shows a degeneration of the photoreceptor layer, and transmission electron microscopy shows a disruption of the IPM and the retinal pigment epithelial cells. The decrease in the concentration of the chromophore 11-cis-retinal supports this loss of photoreceptors. In conclusion, our results demonstrate the essential role of SPACR in maintaining photoreceptors. Impg1-/- mice provide a novel model for mechanistic investigations and the development of therapies for VMD and RP caused by IMPG1 pathogenic variants.

Dominant mutations in mtDNA maintenance gene SSBP1 cause optic atrophy and foveopathy.

Mutations in genes encoding components of the mitochondrial DNA (mtDNA) replication machinery cause mtDNA depletion syndromes (MDSs), which associate ocular features with severe neurological syndromes. Here, we identified heterozygous missense mutations in single-strand binding protein 1 (SSBP1) in 5 unrelated families, leading to the R38Q and R107Q amino acid changes in the mitochondrial single-stranded DNA-binding protein, a crucial protein involved in mtDNA replication. All affected individuals presented optic atrophy, associated with foveopathy in half of the cases. To uncover the structural features underlying SSBP1 mutations, we determined a revised SSBP1 crystal structure. Structural analysis suggested that both mutations affect dimer interactions and presumably distort the DNA-binding region. Using patient fibroblasts, we validated that the R38Q variant destabilizes SSBP1 dimer/tetramer formation, affects mtDNA replication, and induces mtDNA depletion. Our study showing that mutations in SSBP1 cause a form of dominant optic atrophy frequently accompanied with foveopathy brings insights into mtDNA maintenance disorders.

ER-mitochondria cross-talk is regulated by the Ca2+ sensor NCS1 and is impaired in Wolfram syndrome.

Communication between the endoplasmic reticulum (ER) and mitochondria plays a pivotal role in Ca2+ signaling, energy metabolism, and cell survival. Dysfunction in this cross-talk leads to metabolic and neurodegenerative diseases. Wolfram syndrome is a fatal neurodegenerative disease caused by mutations in the ER-resident protein WFS1. Here, we showed that WFS1 formed a complex with neuronal calcium sensor 1 (NCS1) and inositol 1,4,5-trisphosphate receptor (IP3R) to promote Ca2+ transfer between the ER and mitochondria. In addition, we found that NCS1 abundance was reduced in WFS1-null patient fibroblasts, which showed reduced ER-mitochondria interactions and Ca2+ exchange. Moreover, in WFS1-deficient cells, NCS1 overexpression not only restored ER-mitochondria interactions and Ca2+ transfer but also rescued mitochondrial dysfunction. Our results describe a key role of NCS1 in ER-mitochondria cross-talk, uncover a pathogenic mechanism for Wolfram syndrome, and potentially reveal insights into the pathogenesis of other neurodegenerative diseases.

OPA1 gene therapy prevents retinal ganglion cell loss in a Dominant Optic Atrophy mouse model.

Dominant optic atrophy (DOA) is a rare progressive and irreversible blinding disease which is one of the most frequent forms of hereditary optic neuropathy. DOA is mainly caused by dominant mutation in the OPA1 gene encoding a large mitochondrial GTPase with crucial roles in membrane dynamics and cell survival. Hereditary optic neuropathies are commonly characterized by the degeneration of retinal ganglion cells, leading to the optic nerve atrophy and the progressive loss of visual acuity. Up to now, despite increasing advances in the understanding of the pathological mechanisms, DOA remains intractable. Here, we tested the efficiency of gene therapy on a genetically-modified mouse model reproducing DOA vision loss. We performed intravitreal injections of an Adeno-Associated Virus carrying the human OPA1 cDNA under the control of the cytomegalovirus promotor. Our results provide the first evidence that gene therapy is efficient on a mouse model of DOA as the wild-type OPA1 expression is able to alleviate the OPA1-induced retinal ganglion cell degeneration, the hallmark of the disease. These results displayed encouraging effects of gene therapy for Dominant Optic Atrophy, fostering future investigations aiming at clinical trials in patients.

Mutations in DNM1L, as in OPA1, result in dominant optic atrophy despite opposite effects on mitochondrial fusion and fission.

Dominant optic atrophy is a blinding disease due to the degeneration of the retinal ganglion cells, the axons of which form the optic nerves. In most cases, the disease is caused by mutations in OPA1, a gene encoding a mitochondrial large GTPase involved in cristae structure and mitochondrial network fusion. Using exome sequencing, we identified dominant mutations in DNM1L on chromosome 12p11.21 in three large families with isolated optic atrophy, including the two families that defined the OPA5 locus on chromosome 19q12.1-13.1, the existence of which is denied by the present study. Analyses of patient fibroblasts revealed physiological abundance and homo-polymerization of DNM1L, forming aggregates in the cytoplasm and on highly tubulated mitochondrial network, whereas neither structural difference of the peroxisome network, nor alteration of the respiratory machinery was noticed. Fluorescence microscopy of wild-type mouse retina disclosed a strong DNM1L expression in the ganglion cell layer and axons, and comparison between 3-month-old wild-type and Dnm1l+/- mice revealed increased mitochondrial length in retinal ganglion cell soma and axon, but no degeneration. Thus, our results disclose that in addition to OPA1, OPA3, MFN2, AFG3L2 and SPG7, dominant mutations in DNM1L jeopardize the integrity of the optic nerve, suggesting that alterations of the opposing forces governing mitochondrial fusion and fission, similarly affect retinal ganglion cell survival.

Increased steroidogenesis promotes early-onset and severe vision loss in females with OPA1 dominant optic atrophy.

OPA1 mutations are responsible for autosomal dominant optic atrophy (ADOA), a progressive blinding disease characterized by retinal ganglion cell (RGC) degeneration and large phenotypic variations, the underlying mechanisms of which are poorly understood. OPA1 encodes a mitochondrial protein with essential biological functions, its main roles residing in the control of mitochondrial membrane dynamics as a pro-fusion protein and prevention of apoptosis. Considering recent findings showing the importance of the mitochondrial fusion process and the involvement of OPA1 in controlling steroidogenesis, we tested the hypothesis of deregulated steroid production in retina due to a disease-causing OPA1 mutation and its contribution to the visual phenotypic variations. Using the mouse model carrying the human recurrent OPA1 mutation, we disclosed that Opa1 haploinsufficiency leads to very high circulating levels of steroid precursor pregnenolone in females, causing an early-onset vision loss, abolished by ovariectomy. In addition, steroid production in retina is also increased which, in conjunction with high circulating levels, impairs estrogen receptor expression and mitochondrial respiratory complex IV activity, promoting RGC apoptosis in females. We further demonstrate the involvement of Muller glial cells as increased pregnenolone production in female cells is noxious and compromises their role in supporting RGC survival. In parallel, we analyzed ophthalmological data of a multicentre OPA1 patient cohort and found that women undergo more severe visual loss at adolescence and greater progressive thinning of the retinal nerve fibres than males. Thus, we disclosed a gender-dependent effect on ADOA severity, involving for the first time steroids and Müller glial cells, responsible for RGC degeneration.

A dominant mutation in MAPKAPK3, an actor of p38 signaling pathway, causes a new retinal dystrophy involving Bruch's membrane and retinal pigment epithelium.

Inherited retinal dystrophies are clinically and genetically heterogeneous with significant number of cases remaining genetically unresolved. We studied a large family from the West Indies islands with a peculiar retinal disease, the Martinique crinkled retinal pigment epitheliopathy that begins around the age of 30 with retinal pigment epithelium (RPE) and Bruch's membrane changes resembling a dry desert land and ends with a retinitis pigmentosa. Whole-exome sequencing identified a heterozygous c.518T>C (p.Leu173Pro) mutation in MAPKAPK3 that segregates with the disease in 14 affected and 28 unaffected siblings from three generations. This unknown variant is predicted to be damaging by bioinformatic predictive tools and the mutated protein to be non-functional by crystal structure analysis. MAPKAPK3 is a serine/threonine protein kinase of the p38 signaling pathway that is activated by a variety of stress stimuli and is implicated in cellular responses and gene regulation. In contrast to other tissues, MAPKAPK3 is highly expressed in the RPE, suggesting a crucial role for retinal physiology. Expression of the mutated allele in HEK cells revealed a mislocalization of the protein in the cytoplasm, leading to cytoskeleton alteration and cytodieresis inhibition. In Mapkapk3-/- mice, Bruch's membrane is irregular with both abnormal thickened and thinned portions. In conclusion, we identified the first pathogenic mutation in MAPKAPK3 associated with a retinal disease. These findings shed new lights on Bruch's membrane/RPE pathophysiology and will open studies of this signaling pathway in diseases with RPE and Bruch's membrane alterations, such as age-related macular degeneration.

Recessive Mutations in RTN4IP1 Cause Isolated and Syndromic Optic Neuropathies.

Autosomal-recessive optic neuropathies are rare blinding conditions related to retinal ganglion cell (RGC) and optic-nerve degeneration, for which only mutations in TMEM126A and ACO2 are known. In four families with early-onset recessive optic neuropathy, we identified mutations in RTN4IP1, which encodes a mitochondrial ubiquinol oxydo-reductase. RTN4IP1 is a partner of RTN4 (also known as NOGO), and its ortholog Rad8 in C. elegans is involved in UV light response. Analysis of fibroblasts from affected individuals with a RTN4IP1 mutation showed loss of the altered protein, a deficit of mitochondrial respiratory complex I and IV activities, and increased susceptibility to UV light. Silencing of RTN4IP1 altered the number and morphogenesis of mouse RGC dendrites in vitro and the eye size, neuro-retinal development, and swimming behavior in zebrafish in vivo. Altogether, these data point to a pathophysiological mechanism responsible for RGC early degeneration and optic neuropathy and linking RTN4IP1 functions to mitochondrial physiology, response to UV light, and dendrite growth during eye maturation.