mTOR Inhibition via Rapamycin Treatment Partially Reverts the Deficit in Energy Metabolism Caused by FH Loss in RPE Cells
Age-related macular degeneration (AMD) is a complex degenerative disease of the retina with multiple risk-modifying factors, including aging, genetics, and lifestyle choices. The combination of these factors leads to oxidative stress, inflammation, and metabolic failure in the retinal pigment epithe...
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MDPI AG
2021-12-01
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| Series: | Antioxidants |
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| Online Access: | https://www.mdpi.com/2076-3921/10/12/1944 |
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| author | David A. Merle Francesca Provenzano Mohamed Ali Jarboui Ellen Kilger Simon J. Clark Michela Deleidi Angela Armento Marius Ueffing |
| author_facet | David A. Merle Francesca Provenzano Mohamed Ali Jarboui Ellen Kilger Simon J. Clark Michela Deleidi Angela Armento Marius Ueffing |
| author_sort | David A. Merle |
| collection | DOAJ |
| description | Age-related macular degeneration (AMD) is a complex degenerative disease of the retina with multiple risk-modifying factors, including aging, genetics, and lifestyle choices. The combination of these factors leads to oxidative stress, inflammation, and metabolic failure in the retinal pigment epithelium (RPE) with subsequent degeneration of photoreceptors in the retina. The alternative complement pathway is tightly linked to AMD. In particular, the genetic variant in the complement factor H gene (<i>CFH</i>), which leads to the Y402H polymorphism in the factor H protein (FH), confers the second highest risk for the development and progression of AMD. Although the association between the FH Y402H variant and increased complement system activation is known, recent studies have uncovered novel FH functions not tied to this activity and highlighted functional relevance for intracellular FH. In our previous studies, we show that loss of <i>CFH</i> expression in RPE cells causes profound disturbances in cellular metabolism, increases the vulnerability towards oxidative stress, and modulates the activation of pro-inflammatory signaling pathways, most importantly the NF-kB pathway. Here, we silenced <i>CFH</i> in hTERT-RPE1 cells to investigate the mechanism by which intracellular FH regulates RPE cell homeostasis. We found that silencing of <i>CFH</i> results in hyperactivation of mTOR signaling along with decreased mitochondrial respiration and that mTOR inhibition via rapamycin can partially rescue these metabolic defects. To obtain mechanistic insight into the function of intracellular FH in hTERT-RPE1 cells, we analyzed the interactome of FH via immunoprecipitation followed by mass spectrometry-based analysis. We found that FH interacts with essential components of the ubiquitin-proteasomal pathway (UPS) as well as with factors associated with RB1/E2F signalling in a complement-pathway independent manner. Moreover, we found that FH silencing affects mRNA levels of the E3 Ubiquitin-Protein Ligase Parkin and PTEN induced putative kinase (Pink1), both of which are associated with UPS. As inhibition of mTORC1 was previously shown to result in increased overall protein degradation via UPS and as FH mRNA and protein levels were shown to be affected by inhibition of UPS, our data stress a potential regulatory link between endogenous FH activity and the UPS. |
| first_indexed | 2024-03-10T04:38:50Z |
| format | Article |
| id | doaj.art-a96bde4bc24e4a1698fe33f4427a320d |
| institution | Directory Open Access Journal |
| issn | 2076-3921 |
| language | English |
| last_indexed | 2024-03-10T04:38:50Z |
| publishDate | 2021-12-01 |
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| record_format | Article |
| series | Antioxidants |
| spelling | doaj.art-a96bde4bc24e4a1698fe33f4427a320d2023-11-23T03:33:18ZengMDPI AGAntioxidants2076-39212021-12-011012194410.3390/antiox10121944mTOR Inhibition via Rapamycin Treatment Partially Reverts the Deficit in Energy Metabolism Caused by FH Loss in RPE CellsDavid A. Merle0Francesca Provenzano1Mohamed Ali Jarboui2Ellen Kilger3Simon J. Clark4Michela Deleidi5Angela Armento6Marius Ueffing7Institute for Ophthalmic Research, Department for Ophthalmology, Eberhard Karls University of Tübingen, 72076 Tübingen, GermanyGerman Center for Neurodegenerative Diseases (DZNE), 72076 Tübingen, GermanyInstitute for Ophthalmic Research, Department for Ophthalmology, Eberhard Karls University of Tübingen, 72076 Tübingen, GermanyInstitute for Ophthalmic Research, Department for Ophthalmology, Eberhard Karls University of Tübingen, 72076 Tübingen, GermanyInstitute for Ophthalmic Research, Department for Ophthalmology, Eberhard Karls University of Tübingen, 72076 Tübingen, GermanyGerman Center for Neurodegenerative Diseases (DZNE), 72076 Tübingen, GermanyInstitute for Ophthalmic Research, Department for Ophthalmology, Eberhard Karls University of Tübingen, 72076 Tübingen, GermanyInstitute for Ophthalmic Research, Department for Ophthalmology, Eberhard Karls University of Tübingen, 72076 Tübingen, GermanyAge-related macular degeneration (AMD) is a complex degenerative disease of the retina with multiple risk-modifying factors, including aging, genetics, and lifestyle choices. The combination of these factors leads to oxidative stress, inflammation, and metabolic failure in the retinal pigment epithelium (RPE) with subsequent degeneration of photoreceptors in the retina. The alternative complement pathway is tightly linked to AMD. In particular, the genetic variant in the complement factor H gene (<i>CFH</i>), which leads to the Y402H polymorphism in the factor H protein (FH), confers the second highest risk for the development and progression of AMD. Although the association between the FH Y402H variant and increased complement system activation is known, recent studies have uncovered novel FH functions not tied to this activity and highlighted functional relevance for intracellular FH. In our previous studies, we show that loss of <i>CFH</i> expression in RPE cells causes profound disturbances in cellular metabolism, increases the vulnerability towards oxidative stress, and modulates the activation of pro-inflammatory signaling pathways, most importantly the NF-kB pathway. Here, we silenced <i>CFH</i> in hTERT-RPE1 cells to investigate the mechanism by which intracellular FH regulates RPE cell homeostasis. We found that silencing of <i>CFH</i> results in hyperactivation of mTOR signaling along with decreased mitochondrial respiration and that mTOR inhibition via rapamycin can partially rescue these metabolic defects. To obtain mechanistic insight into the function of intracellular FH in hTERT-RPE1 cells, we analyzed the interactome of FH via immunoprecipitation followed by mass spectrometry-based analysis. We found that FH interacts with essential components of the ubiquitin-proteasomal pathway (UPS) as well as with factors associated with RB1/E2F signalling in a complement-pathway independent manner. Moreover, we found that FH silencing affects mRNA levels of the E3 Ubiquitin-Protein Ligase Parkin and PTEN induced putative kinase (Pink1), both of which are associated with UPS. As inhibition of mTORC1 was previously shown to result in increased overall protein degradation via UPS and as FH mRNA and protein levels were shown to be affected by inhibition of UPS, our data stress a potential regulatory link between endogenous FH activity and the UPS.https://www.mdpi.com/2076-3921/10/12/1944retinal pigment epithelium (RPE) cellsage-related macular degeneration (AMD)complement factor H (<i>CFH</i>/FH)mammalian target of rapamycin (mTOR)mitochondrial respirationinteractome |
| spellingShingle | David A. Merle Francesca Provenzano Mohamed Ali Jarboui Ellen Kilger Simon J. Clark Michela Deleidi Angela Armento Marius Ueffing mTOR Inhibition via Rapamycin Treatment Partially Reverts the Deficit in Energy Metabolism Caused by FH Loss in RPE Cells Antioxidants retinal pigment epithelium (RPE) cells age-related macular degeneration (AMD) complement factor H (<i>CFH</i>/FH) mammalian target of rapamycin (mTOR) mitochondrial respiration interactome |
| title | mTOR Inhibition via Rapamycin Treatment Partially Reverts the Deficit in Energy Metabolism Caused by FH Loss in RPE Cells |
| title_full | mTOR Inhibition via Rapamycin Treatment Partially Reverts the Deficit in Energy Metabolism Caused by FH Loss in RPE Cells |
| title_fullStr | mTOR Inhibition via Rapamycin Treatment Partially Reverts the Deficit in Energy Metabolism Caused by FH Loss in RPE Cells |
| title_full_unstemmed | mTOR Inhibition via Rapamycin Treatment Partially Reverts the Deficit in Energy Metabolism Caused by FH Loss in RPE Cells |
| title_short | mTOR Inhibition via Rapamycin Treatment Partially Reverts the Deficit in Energy Metabolism Caused by FH Loss in RPE Cells |
| title_sort | mtor inhibition via rapamycin treatment partially reverts the deficit in energy metabolism caused by fh loss in rpe cells |
| topic | retinal pigment epithelium (RPE) cells age-related macular degeneration (AMD) complement factor H (<i>CFH</i>/FH) mammalian target of rapamycin (mTOR) mitochondrial respiration interactome |
| url | https://www.mdpi.com/2076-3921/10/12/1944 |
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