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Si us plau utilitzeu sempre aquest identificador per citar o enllaçar aquest document: https://hdl.handle.net/2445/212821
CERKL as a modulator gene of mitochondrial dynamics and stress response in the retina
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[eng] Inherited Retinal Dystrophies (IRDs) are a clinically and genetically highly heterogeneous group of genetic pathologies characterized by progressive attrition of photoreceptor cells and other retinal neurons, which eventually leads to vision loss. The retina, the most affected tissue in IRDs, is the specialized region of the central nervous system capable of transducing light into neural signals. This neurosensory tissue is particularly susceptible to genetic and environmental alterations due to its highly active metabolism, external location, and daily light irradiation. Therefore, the disturbance of the balance between retinal resilience systems and endogenous and exogenous stress factors eventually leads to several alterations that underlie the pathogenesis of many IRDs, such as mitochondrial dysfunction, misregulated autophagy, and activation of cell death pathways. CERKL (CERamide Kinase-Like) mutations cause two different IRDs in humans: Retinitis Pigmentosa and Cone-Rod Dystrophy. While the precise role of CERKL remains unclear, numerous studies have proposed CERKL as a resilience gene against oxidative stress, by participating in the formation of stress granules, regulation of the antioxidant protein TRX2 and inhibition of oxidative stress-induced apoptosis, among other functions. Previous work from our research group led to the generation of the CerklKD/KO mouse model, characterised by a strong deficiency in the expression of Cerkl. This model mimics the disease progression of CERKL-associated Retinitis Pigmentosa-affected patients, showing a slow and progressive loss of photoreceptors and, ultimately, vision impairment. Taking advantage of the CerklKD/KO mouse model, we aimed to dissect CERKL function in mitochondrial metabolism and dynamics. Our findings describe a pool of CERKL isoforms colocalizing with mitochondria. In addition, we observed accumulation of fragmented mitochondria and mitochondrial bioenergetics dysfunction in CerklKD/KO retinas. Moreover, mitochondrial distribution and trafficking were reduced in retinal and hippocampal neurons upon Cerkl depletion, reflecting the important role of CERKL in the regulation of mitochondrial network morphology and energy production. Furthermore, we sought to analyse the impact of CERKL downregulation on stress response and activation of photoreceptor death mechanisms upon light/oxidative stress. Data collected from CERKL silencing and overexpression experiments in ARPE-19 cells (derived from retinal pigment epithelium) revealed that CERKL exerts a protective role maintaining the mitochondrial network morphology against oxidative damage. Additionally, using CerklKD/KO albino mouse models, we assessed immediate (early) or after two weeks (late) retinal stress response to light injury, using data from transcriptomics, metabolomics, and immunohistochemistry images. Our results showed that Cerkl depletion causes an exacerbated response to stress in basal conditions, through alterations in glutathione metabolism and stress granule production. Consequently, upon light-stress exposure, CerklKD/KO retinas cannot cope with additional stress factors, resulting in increased ROS levels and the subsequent activation of several cell death mechanisms. To sum up, our studies indicate that Cerkl gene is a novel player in regulating mitochondrial organization and metabolism, together with light-challenged retinal homeostasis, thus suggesting that CERKL mutations cause blindness by impairing the mitochondrial homeostasis and oxidative stress response in the retina. These findings contributed to determine early phenotypic biomarkers of the Cerkl-depleted mouse retina, which will be compared and confirmed with those observed in retinal organoids derived from CERKLR257X patient-derived hiPSCs (currently under differentiation). Altogether, these studies will allow us to test the feasibility of genetic rescue using a proof-of-principle AAV-based gene augmentation therapy for CERKL-associated IRDs, as well as novel gene delivery systems using gold nanoparticles as vectors. The principal area of cooperation of this Thesis in the Sustainable Development Goals (SDGs) is related to the SDG 3: “Good health and well-being” according to the point 3.8: “Achieve universal health coverage, including financial risk protection, access to quality essential health-care services and access to safe, effective, quality and affordable essential medicines and vaccines for all”. Moreover, an important part of our findings was result of fruitful collaborations with groups with high expertise in other areas. Therefore, the objectives of this work are also related to SDG 17: “Partnerships for the goals” according to the point 17.6: “Knowledge sharing and cooperation for access to science, technology and innovation”. Finally, this PhD Thesis has been directed, supervised and tutorized by women, Dr Gemma Marfany and Dr Serena Mirra, and our research group has also been mostly composed of female scientists during many years, contributing to SDG 5: “Gender equity and empowering of girls and women” according to the point 5.5: “Assure effective and full participation of women and equity in leadership opportunities at all deciding levels in politic, economic and public life”.
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GARCÍA ARROYO, Rocío. CERKL as a modulator gene of mitochondrial dynamics and stress response in the retina. [consulta: 26 de novembre de 2025]. [Disponible a: https://hdl.handle.net/2445/212821]