Dataset Information

Investigating Autophagy Disruption Induces Neurodegeneration in Glaucoma OPTN(E50K) Retinal Ganglion Cells

ABSTRACT: Retinal ganglion cells (RGCs) are the sole projection neurons that connect the eye with the brain, and the degeneration of these cells in diseases such as glaucoma results in vision loss or blindness. Similar to other neurons throughout the central nervous system, RGCs are postmitotic and therefore highly dependent upon autophagy to remove damaged proteins or organelles to maintain proper cellular homeostasis. Autophagy deficits have been implicated in multiple neurodegenerative diseases including glaucoma. In addition, a subpopulation of glaucoma patients possesses mutations in the autophagy receptor Optineurin (OPTN) resulting glaucoma within a normal range of intraocular pressure, with the OPTN(E50K) mutation known to induce a severe degeneration. Despite this, our knowledge of how autophagy impairment promotes neurodegeneration within RGCs remains limited. We advanced a human pluripotent stem cell (hPSC) model of RGC neurodegeneration with an underlying OPTN(E50K) mutation to study how autophagy disruption contributes to RGC neurodegeneration. We identified OPTN protein was reduced but accumulated within the somas in RGCs with the OPTN(E50K) mutation, and this accumulation was associated with a decrease in autophagic flux. To rule out the possibilities that the specificity of the antibody incapable to recognize E50K region resulting the reduction of OPTN protein, we performed proteomics analysis at week 2 hPSC-RGCs of post purification and identified 154 downregulated proteins as well as 178 upregulated proteins associated with OPTN(E50K) RGCs compared with isogenic controls. Taken together, we identified proteins that were altered by OPTN(E50K) mutation in RGCs, and which may provide potential mechanisms that contribute to RGC neurodegeneration.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Retinal Ganglion Cell, Stem Cell

SUBMITTER: Emma Doud

LAB HEAD: Amber Mosley

PROVIDER: PXD033173 | Pride | 2025-05-06

REPOSITORIES: Pride

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Dataset's files

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			Action	DRS
	2022_02_34_Meyer_Eclipse_FAIMS_aurora_Global_MS2_F.mzML	Mzml
	2022_02_34_Meyer_Eclipse_FAIMS_aurora_Global_MS2_F.mzid.gz	Mzid
	2022_02_34_Meyer_Eclipse_FAIMS_aurora_Global_MS2_F.pdResult	Other
	2022_02_34_Meyer_Eclipse_FAIMS_aurora_Global_MS2_F1.raw	Raw
	2022_02_34_Meyer_Eclipse_FAIMS_aurora_Global_MS2_F2.raw	Raw

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Publications

Acquisition of neurodegenerative features in isogenic OPTN(E50K) human stem cell-derived retinal ganglion cells associated with autophagy disruption and mTORC1 signaling reduction.

Huang Kang-Chieh KC Gomes Cátia C Shiga Yukihiro Y Belforte Nicolas N VanderWall Kirstin B KB Lavekar Sailee S SS Fligor Clarisse M CM Harkin Jade J Hetzer Shelby M SM Patil Shruti V SV Di Polo Adriana A Meyer Jason S JS

Acta neuropathologica communications 20241018 1

The ability to derive retinal ganglion cells (RGCs) from human pluripotent stem cells (hPSCs) has led to numerous advances in the field of retinal research, with great potential for the use of hPSC-derived RGCs for studies of human retinal development, in vitro disease modeling, drug discovery, as well as their potential use for cell replacement therapeutics. Of all these possibilities, the use of hPSC-derived RGCs as a human-relevant platform for in vitro disease modeling has received the great ...[more]

PMID: 39425218

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Project description:Glaucoma is a group of optic neuropathies characterized by the progressive degeneration of retinal ganglion cells (RGCs) as well as their axons leading to irreversible loss of sight. Medical management of the intraocular pressure (IOP) still represents the gold standard in glaucoma therapy, which only manages a risk factor and does not directly address the neurodegenerative component of this eye disease. Recently, our group showed that antibody derived immunopeptides (encoding complementarity-determining regions, CDRs) provide attractive glaucoma medication candidates and directly interfere its pathogenic mechanisms by different modes of action. In accordance with these findings, the present study showed the synthetic CDR2 peptide (INSDGSSTSYADSVK) significantly increased the RGCs viability in vitro in a concentration-dependent manner (p < 0.05 using a CDR2 concentration of 50 µg/mL). Employing state-of the-art immunoprecipitation experiments, we confirmed that synthetic CDR2 exhibited a high affinity towards the retinal target protein histone H3.1 (HIST1H3A) (p < 0.001 and log2 fold change > 3). Furthermore, virtual docking analyses predicted potential CDR2-specific binding regions of HIST1H3A, which might represent essential PTM sites for epigenetic regulations. Quantitative MS analysis revealed that 39 proteins were significantly changed between CDR2-treated and untreated control retinae (p < 0.05). An up-regulation of proteins involved in the energy production (e.g., ATP5F1B and MT-CO2) as well as the regulatory ubiquitin proteasome system (e.g., PSMC5) was induced by the synthetic CDR2 peptide. On the other hand, metabolic key enzymes (e.g., DDAH1 and MAOB) as well as ER stress-related proteins (e.g., SEC22B and VCP) were found with low abundancy in CDR2-treated retinae and were also partially confirmed by microarray technology. Based on these findings, it can be hypothesized that the specific protein-peptide interaction influences the regulatory epigenetic function of HIST1H3A promoting the neuroprotective mechanism on RGCs in vitro. It also might serve as basis for a synergistic immunotherapy of glaucoma in the future.