Data from: Lipid-driven membrane remodeling engages a lysosome-dependent adaptive repair program during retinal aging
Data files
Oct 06, 2026 version files 358.45 KB
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15mo_Elovl2_vs_18mo_WT_eyecup.csv
96.90 KB
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16mo_3x_suppl_lipidomics.csv
64.95 KB
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18mo_Elovl2_mouse_eyecup_lip.csv
68.96 KB
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18mo_WT_Elovl2_mouse_eyecup_tFA.csv
2.75 KB
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3mo_18mo_mouse_eyecup_total_FA.csv
3.85 KB
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ELOVL2_KD_lipidomics.csv
44.88 KB
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ELOVL2_KD_PM_lipidomics_exp1.csv
21.93 KB
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ELOVL2_KD_PM_lipidomics_exp2.csv
31.65 KB
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ELOVL2_KD_tFA.csv
2.98 KB
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iPSC_RPE_phagocytosis.csv
4.74 KB
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iPSC_RPE_tFA.csv
2.15 KB
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README.md
12.72 KB
Abstract
Age-associated remodeling of membrane lipid composition has been implicated in cellular dysfunction, yet the mechanisms linking lipid changes to membrane integrity and disease remain poorly defined. In the retinal pigment epithelium (RPE), lipid dysregulation is strongly associated with aging and age-related macular degeneration (AMD), a neurodegenerative disease of the central nervous system, but the causal pathways remain unclear. Here, we identify reduced activity of the lipid elongase ELOVL2 as a central driver of age-dependent membrane remodeling. Loss of ELOVL2-dependent polyunsaturated fatty acid (PUFA) elongation shifts plasma membrane lipid composition, leading to altered membrane biophysical properties and compromised membrane integrity. In response to this stress, RPE cells do not undergo apoptosis but instead activate a lysosome-dependent plasma membrane repair program that preserves barrier function under metabolic challenge. However, this adaptive response drives spatially polarized lysosomal exocytosis, promoting extracellular remodeling and accumulation of sub-RPE deposits associated with aging and AMD. Restoration of ELOVL2-derived lipid products reverses membrane abnormalities and suppresses lysosome-mediated remodeling phenotypes, demonstrating direct metabolic control of membrane homeostasis. Together, these findings define an ELOVL2-dependent lipid–lysosome axis that links PUFA elongation to plasma membrane integrity and reveals how compensatory repair mechanisms can contribute to tissue remodeling and disease progression in aging epithelia.
Dataset DOI: 10.5061/dryad.34tmpg51p
Files and variables
File: 16mo_3x_suppl_lipidomics.csv
Description: Lipidomics analysis from 16-month-old mouse eyecups that received intravitreal 24:5n-3 supplementation and contralateral vehicle control every month for 3 months
Variables
- LipidMolec: Names of the lipid molecules detected in the analysis.
- Class: Lipid class each molecule belongs to.
- Formula: Molecular formula of each lipid.
- BaseRt: Retention time at which the lipid elutes during chromatographic separation.
- MainGrade[c]: Quality and confidence classification assigned to lipid identification based on how completely the lipid class and constituent fatty acyl chains are resolved, ie. A: The lipid class and all constituent fatty acid chains/positions are completely and unambiguously identified (high structural confidence), B: The lipid class and some (but not all) fatty acid chains are identified, Grade C: Only the lipid class or a partial/ambiguous fatty acid component is identified
Sample Columns
- VEH-1: vehicle-injected eye, replicate 1
- VEH-2: vehicle-injected eye, replicate 2
- VEH-3: vehicle-injected eye, replicate 3
- VEH-4: vehicle-injected eye, replicate 4
- SUPPL-1: 24:5n-3-supplemented eye, replicate 1
- SUPPL-2: 24:5n-3-supplemented eye, replicate 2
- SUPPL-3: 24:5n-3-supplemented eye, replicate 3
- SUPPL-4: 24:5n-3-supplemented eye, replicate 4
File: iPSC_RPE_phagocytosis.csv
Description: Total fatty acids extracted from iPSC-RPE CTRL vs. ELOVL2 KO incubated with bovine photoreceptor outer segments (POS) at various concentrations (0, 10, 20 POS/cell) for 3 hours
Variables
- FAs: Names of the fatty acids detected in the analysis.
- Chemical formula: Molecular formula of each fatty acid.
- m/z: Mass-to-charge ratio of the detected fatty acid ion.
- tR/min: Retention time in minutes during chromatographic separation.
Sample Columns
- CTRL_0-1: CTRL, 0 POS/cell, replicate 1
- CTRL_0-2: CTRL, 0 POS/cell, replicate 2
- CTRL_10-1: CTRL, 10 POS/cell, replicate 1
- CTRL_10-2: CTRL, 10 POS/cell, replicate 2
- CTRL_10-3: CTRL, 10 POS/cell, replicate 3
- CTRL_20-1: CTRL, 20 POS/cell, replicate 1
- CTRL_20-2: CTRL, 20 POS/cell, replicate 2
- CTRL_20-3: CTRL, 20 POS/cell, replicate 3
- KO_0-1: ELOVL2 KO, 0 POS/cell, replicate 1
- KO_0-2: ELOVL2 KO, 0 POS/cell, replicate 2
- KO_10-1: ELOVL2 KO, 10 POS/cell, replicate 1
- KO_10-2: ELOVL2 KO, 10 POS/cell, replicate 2
- KO_10-3: ELOVL2 KO, 10 POS/cell, replicate 3
- KO_20-1: ELOVL2 KO, 20 POS/cell, replicate 1
- KO_20-2: ELOVL2 KO, 20 POS/cell, replicate 2
- KO_20-3: ELOVL2 KO, 20 POS/cell, replicate 3
File: iPSC_RPE_tFA.csv
Description: Total fatty acids extracted from iPSC-RPE CTRL vs. ELOVL2 KO
Variables
- FAs: Names of the fatty acids detected in the analysis.
- Chemical formula: Molecular formula of each fatty acid.
- m/z: Mass-to-charge ratio of the detected fatty acid ion.
- tR/min: Retention time in minutes during chromatographic separation.
Sample Columns
- C-1: CTRL, replicate 1
- C-2: CTRL, replicate 2
- KO_D-1: ELOVL2 KO, replicate 1
- KO_D-2: ELOVL2 KO, replicate 2
File: ELOVL2_KD_PM_lipidomics_exp1.csv
Description: Lipidomics analysis from plasma membrane (PM) fractions from CTRL vs. ELOVL2 KD ARPE-19 cells, experiment 1
Variables
- LipidMolec: Names of the lipid molecules detected in the analysis.
- Class: Lipid class each molecule belongs to.
- Formula: Molecular formula of each lipid.
- BaseRt: Retention time at which the lipid elutes during chromatographic separation.
- MainGrade[c]: Quality and confidence classification assigned to lipid identification based on how completely the lipid class and constituent fatty acyl chains are resolved, ie. A: The lipid class and all constituent fatty acid chains/positions are completely and unambiguously identified (high structural confidence), B: The lipid class and some (but not all) fatty acid chains are identified, Grade C: Only the lipid class or a partial/ambiguous fatty acid component is identified
Sample Columns
- C-1: CTRL PM, replicate 1, experiment 1
- C-2: CTRL PM, replicate 2, experiment 1
- C-3: CTRL PM, replicate 3, experiment 1
- KD-1: ELOVL2 KD PM, replicate 1, experiment 1
- KD-2: ELOVL2 KD PM, replicate 2, experiment 1
- KD-3: ELOVL2 KD PM, replicate 3, experiment 1
File: ELOVL2_KD_PM_lipidomics_exp2.csv
Description: Lipidomics analysis from plasma membrane (PM) fractions from CTRL vs. ELOVL2 KD ARPE-19 cells, experiment 2
Variables
- LipidMolec: Names of the lipid molecules detected in the analysis.
- Class: Lipid class each molecule belongs to.
- Formula: Molecular formula of each lipid.
- BaseRt: Retention time at which the lipid elutes during chromatographic separation.
- MainGrade[c]: Quality and confidence classification assigned to lipid identification based on how completely the lipid class and constituent fatty acyl chains are resolved, ie. A: The lipid class and all constituent fatty acid chains/positions are completely and unambiguously identified (high structural confidence), B: The lipid class and some (but not all) fatty acid chains are identified, Grade C: Only the lipid class or a partial/ambiguous fatty acid component is identified
Sample Columns
- C-1: CTRL PM, replicate 1, experiment 2
- C-2: CTRL PM, replicate 2, experiment 2
- C-3: CTRL PM, replicate 3, experiment 2
- KD-1: ELOVL2 KD PM, replicate 1, experiment 2
- KD-2: ELOVL2 KD PM, replicate 2, experiment 2
- KD-3: ELOVL2 KD PM, replicate 3, experiment 2
File: ELOVL2_KD_lipidomics.csv
Description: Lipidomics analysis from CTRL vs. ELOVL2 KD differentiated ARPE-19 cells
Variables
- LipidMolec: Names of the lipid molecules detected in the analysis.
- Class: Lipid class each molecule belongs to.
- Formula: Molecular formula of each lipid.
- BaseRt: Retention time at which the lipid elutes during chromatographic separation.
- MainGrade[c]: Quality and confidence classification assigned to lipid identification based on how completely the lipid class and constituent fatty acyl chains are resolved, ie. A: The lipid class and all constituent fatty acid chains/positions are completely and unambiguously identified (high structural confidence), B: The lipid class and some (but not all) fatty acid chains are identified, Grade C: Only the lipid class or a partial/ambiguous fatty acid component is identified
Sample Columns
- C1: CTRL, replicate 1
- C2: CTRL, replicate 2
- C3: CTRL, replicate 3
- C4: CTRL, replicate 4
- KD1: ELOVL2 KD, replicate 1
- KD2: ELOVL2 KD, replicate 2
- KD3: ELOVL2 KD, replicate 3
- KD4: ELOVL2 KD, replicate 4
File: ELOVL2_KD_tFA.csv
Description: Total fatty acids extracted from from CTRL vs. ELOVL2 KD differentiated ARPE-19 cells
Variables
- FAs: Names of the fatty acids detected in the analysis.
- Chemical formula: Molecular formula of each fatty acid.
- m/z: Mass-to-charge ratio of the detected fatty acid ion.
- tR/min: Retention time in minutes during chromatographic separation.
Sample Columns
- C1: CTRL, replicate 1
- C2: CTRL, replicate 2
- C3: CTRL, replicate 3
- C4: CTRL, replicate 4
- KD1: ELOVL2 KD, replicate 1
- KD2: ELOVL2 KD, replicate 2
- KD3: ELOVL2 KD, replicate 3
- KD4: ELOVL2 KD, replicate 4
File: 15mo_Elovl2_vs_18mo_WT_eyecup.csv
Description: Lipidomics analysis from 15-month-old Elovl2 mutant and 18-month-old WT mouse eyecups
Variables
- LipidMolec: Names of the lipid molecules detected in the analysis.
- Class: Lipid class each molecule belongs to.
- Formula: Molecular formula of each lipid.
- BaseRt: Retention time at which the lipid elutes during chromatographic separation.
- MainGrade[c]: Quality and confidence classification assigned to lipid identification based on how completely the lipid class and constituent fatty acyl chains are resolved, ie. A: The lipid class and all constituent fatty acid chains/positions are completely and unambiguously identified (high structural confidence), B: The lipid class and some (but not all) fatty acid chains are identified, Grade C: Only the lipid class or a partial/ambiguous fatty acid component is identified
Sample Columns
- 18moWT-1: 18-month-old, wildtype, replicate 1
- 18moWT-2: 18-month-old, wildtype, replicate 2
- 18moWT-3: 18-month-old, wildtype, replicate 3
- 15moELOVL2-1: 15-month-old, Elovl2 mutant, replicate 1
- 15moELOVL2-2: 15-month-old, Elovl2 mutant, replicate 2
- 15moELOVL2-3: 15-month-old, Elovl2 mutant, replicate 3
File: 18mo_WT_Elovl2_mouse_eyecup_tFA.csv
Description: Total fatty acids extracted from 18-month-old wildtype and Elovl2 mutant mouse eyecups
Variables
- FAs: Names of the fatty acids detected in the analysis.
- Chemical formula: Molecular formula of each fatty acid.
- m/z: Mass-to-charge ratio of the detected fatty acid ion.
- tR/min: Retention time in minutes during chromatographic separation.
Sample Columns
- WT-1: 18-month-old, wildtype, replicate 1
- WT-2: 18-month-old, wildtype, replicate 2
- WT-3: 18-month-old, wildtype, replicate 3
- C234W1: 18-month-old, Elovl2 mutant, replicate 1
- C234W-2: 18-month-old, Elovl2 mutant, replicate 2
- C234W-3: 18-month-old, Elovl2 mutant, replicate 3
File: 3mo_18mo_mouse_eyecup_total_FA.csv
Description: Total fatty acids extracted from 3-month-old and 18-month-old mouse eyeupcs
Variables
- FAs: Names of the fatty acids detected in the analysis.
- Chemical formula: Molecular formula of each fatty acid.
- m/z: Mass-to-charge ratio of the detected fatty acid ion.
- tR/min: Retention time in minutes during chromatographic separation.
Sample Columns
- 3mo-1: 3-month-old, replicate 1
- 3mo-2: 3-month-old, replicate 2
- 3mo-3: 3-month-old, replicate 3
- 3mo-4: 3-month-old, replicate 4
- 3mo-5: 3-month-old, replicate 5
- 18mo-1: 18-month-old, replicate 1
- 18mo-2: 18-month-old, replicate 2
- 18mo-3: 18-month-old, replicate 3
- 18mo-4: 18-month-old, replicate 4
- 18mo-5: 18-month-old, replicate 5
File: 18mo_Elovl2_mouse_eyecup_lip.csv
Description: Lipidomics analysis of 18-month-old wildtype and Elovl2 mutant mouse eyecups
Variables
- LipidMolec: Names of the lipid molecules detected in the analysis.
- Class: Lipid class each molecule belongs to.
- Formula: Molecular formula of each lipid.
- BaseRt: Retention time at which the lipid elutes during chromatographic separation.
- MainGrade[c]: Quality and confidence classification assigned to lipid identification based on how completely the lipid class and constituent fatty acyl chains are resolved, ie. A: The lipid class and all constituent fatty acid chains/positions are completely and unambiguously identified (high structural confidence), B: The lipid class and some (but not all) fatty acid chains are identified, Grade C: Only the lipid class or a partial/ambiguous fatty acid component is identified
Sample Columns
- WT-1: 18-month-old, wildtype, replicate 1
- WT-2: 18-month-old, wildtype, replicate 2
- C234W-1: 18-month-old, Elovl2 mutant, replicate 1
- C234W-2: 18-month-old, Elovl2 mutant, replicate 2
Code/software
Data analysis and post-processing
Data were analyzed with LipidSearch 4.2.21 software. Only peaks with molecular identification grade: A or B were accepted (A: lipid class and fatty acids completely identified or B: lipid class and some fatty acids identified). The relative abundance of each lipid species was obtained by normalization to the total lipid intensity. Significantly changed lipid species (FC > 1.5. P < 0.05) were submitted to Lipid Ontology (LION) for lipid ontology analysis. Data visualization was performed using Prism 7 software (GraphPad Software, Inc.).
FA analysis
Separation of PUFAs was achieved on an Acquity UPLC® BEH C18 column (1.7 μm, 2.1 mm × 100 mm, Waters Corporation). The Q Exactive MS was operated in a full MS scan mode (resolution 70,000 at m/z 200) in negative mode. For the compounds of interest, a scan range of m/z 250–800 was chosen. The identification of fatty acids was based on retention time and formula. Blank cells mean that the lipid or fatty acid species was not detected/below the limit of detection.
Lipidomic analysis
Lipid extraction
Lipid extractions were performed according to the methodology of Bligh and Dyer74. In brief, the tissue was homogenized in 200 μL water, transferred to a glass vial, and 750 μL 1:2 (v/v) CHCl3: MeOH was added and vortexed. Then 250 μL CHCl3 was added and vortexed. Finally, 250 μL ddH2O was added and vortexed. The samples were centrifuged at 3000 rpm for 5 min at 4 °C. The lower phase was transferred to a new glass vial, dried under nitrogen, and stored at -20 °C until subsequent lipid analysis.
LC-MS/MS
Separation of lipids was performed on an Accucore C30 column (2.6 μm, 2.1 mm × 150 mm, Thermo Scientific). The Q Exactive MS was operated in a full MS scan mode (resolution 70,000 at m/z 200) followed by ddMS2 (17,500 resolution) in both positive and negative modes. The AGC target value was set at 1E6 and 1E5 for the MS and MS/MS scans, respectively. The maximum injection time was 200 ms for MS and 50 ms for MS/MS. HCD was performed with a stepped collision energy of 30 ± 10% for negative and 25% and 30% for positive ion mode with an isolation window of 1.5 Da.
Data analysis and post-processing
Data were analyzed with LipidSearch 4.2.21 software. Only peaks with molecular identification grade: A or B were accepted (A: lipid class and fatty acids completely identified or B: lipid class and some fatty acids identified). The relative abundance of each lipid species was obtained by normalization to the total lipid intensity. Significantly changed lipid species (FC > 1.5. P < 0.05) were submitted to Lipid Ontology (LION) for lipid ontology analysis. Data visualization was performed using Prism 7 software (GraphPad Software, Inc.).
FA analysis
Separation of PUFAs was achieved on an Acquity UPLC® BEH C18 column (1.7 μm, 2.1 mm × 100 mm, Waters Corporation). The Q Exactive MS was operated in a full MS scan mode (resolution 70,000 at m/z 200) in negative mode. For the compounds of interest, a scan range of m/z 250–800 was chosen. The identification of fatty acids was based on retention time and formula.