Phospholipid Binding Switch of Prohibitin Determines Aging and Mitochondrial Dysfunction in the RPE

Abstract

Purpose: Previously, our proteomics study suggested that the coordinated translocalization of prohibitin may determine the cell viability and the apoptotic cell population in the retina and RPE. The current study tested whether or not there is a positive correlation between prohibitin depletion in the retinal pigment epithelium(RPE) and age-related macular degeneration(AMD). The lipid binding switch mechanism of prohibitin with phosphatidylinositoltriphosphate(PIP3) and cardiolipin may suggest the origin of prohibitin’s effect on energy metabolism(PIP3) and aging process(cardiolipin).

Methods: We used in vivo human models and interaction assays that include lipids, nucleotides and proteins to determine the molecular mechanisms involved in AMD. Immunoprecipitation and immunocytochemistry were performed to identify and visualize protein localization and expressions from RPE cells. Two-dimensional SDS-PAGE and Western blotting were used to identify proteins in AMD eyes compared to age-matching control. Selected spots from 2D electrophoresis were excised and analyzed by MALDI-TOF/ESI MS/MS. Lipid-prohibitin interaction assay was performed to identify prohibitin binding molecules, inlcuding cardiolipin, cholesterol and phosphatidyl inositides.

Results: Our current data reveal that prohibitin in the peripheral RPE is down-regulated in AMD. Our study implies that prohibitin may function as an anti-apoptotic molecule by binding with mitochondrial DNA in the beginning of retinal degeneration, yet it may become a transcriptional regulator under aging conditions. Prohibitin showed strong affinity with PIP3 compared to PI(4,5)P2. Prohibitin-PIP3 interaction may facilitate the forward reaction of the PI3K pathway in normal conditions. Under stress conditions, prohibitin interacts with cardiolipin as a retrograde response to activate mitochondrial proliferation. The lipid-binding switch mechanism of prohibitin with PIP3 and cardiolipin may suggest the role of prohibitin effects on energy metabolism and age-related diseases.

Conclusions: The current study aimed to show that prohibitin downregulation in the RPE leads to AMD. Decreased prohibitin may modulate lipid metabolism, transcriptional activation, and RPE apoptosis to accelerate the progression of AMD. Preventing prohibitin depletion in the RPE may provide a critical step for the development of more effective therapeutic strategies for the treatment of AMD.