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Prohibitin as the Molecular Binding Switch in the Retinal Pigment Epithelium

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Abstract

Previously, our molecular binding study showed that prohibitin interacts with phospholipids, including phosphatidylinositide and cardiolipin. Under stress conditions, prohibitin interacts with cardiolipin as a retrograde response to activate mitochondrial proliferation. The lipid-binding switch mechanism of prohibitin with phosphatidylinositol-3,4,5-triphosphate and cardiolipin may suggest the role of prohibitin effects on energy metabolism and age-related diseases. The current study examined the region-specific expressions of prohibitin with respect to the retina and retinal pigment epithelium (RPE) in age-related macular degeneration (AMD). A detailed understanding of prohibitin binding with lipids, nucleotides, and proteins shown in the current study may suggest how molecular interactions control apoptosis and how we can intervene against the apoptotic pathway in AMD. Our data imply that decreased prohibitin in the peripheral RPE is a significant step leading to mitochondrial dysfunction that may promote AMD progression.

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Abbreviations

DAPI:

4′,6-Diamidino-2-phenylindole

DMEM:

Dulbecco’s modified eagle medium

IP:

Immunoprecipitation

ROS:

Reactive oxygen species

RPE:

Retinal pigment epithelium

siRNA:

Small interfering RNA

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Acknowledgments

The authors thank Jeremy Goldman and Mike Gibson for insightful discussions and sharing equipment. Matthew Durocher and Dr. Tristan Purvis are acknowledged for their suggestions and critical reading. This study was supported by the Century II Equipment fund and the Research Excellence Fund from Michigan Technological University, Research Assistantship and Teaching Assistantship from American University of Nigeria.

Authors’ Contributions

SRS and WJJ designed the hypothesis, aim, and experiments. SRS, ODS, FL, WH, TM, JYU conducted the experiments. WR, PSB, MB, WJJ provided materials, equipment, and participated in discussion. All authors wrote the paper.

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Authors and Affiliations

  1. Department of Biological Sciences, Michigan Technological University, Houghton, MI, 49931, USA

    Srinivas R. Sripathi, Weilue He, Trevor Moser, Ji-Yeon Um & Wusirika Ramakrishna

  2. Department of Ophthalmology, Wilmer Eye Institute, The Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA

    Srinivas R. Sripathi

  3. Retina Proteomics Laboratory, Department of Petroleum Chemistry, American University of Nigeria, Yola, Nigeria

    O’Donnell Sylvester & Wan Jin Jahng

  4. Department of Ophthalmology, Georgia Regents University, Augusta, GA, 30912, USA

    Folami Lamoke & Manuela Bartoli

  5. Department of Ophthalmology and Visual Sciences, Moran Eye Center, University of Utah School of Medicine, Salt Lake City, UT, 84132, USA

    Paul S. Bernstein

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  1. Srinivas R. Sripathi
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  2. O’Donnell Sylvester
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Correspondence to Wan Jin Jahng.

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10930_2015_9641_MOESM1_ESM.pdf

Supplementary material 1 Supplement Figure S1. A. Immunoprecipitation of PHB interacting proteins. Coomassie blue stained gel shows immunoprecipitated proteins from control ARPE-19 cells (lysate) by anti-PHB antibody binding. Cells were harvested, lysed and proteins were analyzed by immunoprecipitation and SDS-PAGE. Proteins in W1 to W3 represents wash fractions; E1 to E3 represents elution fractions; Eb represents the protein elution by immunoprecipitated boiling protein-A beads with SDS-sample buffer. Elution fractions showed the potential binding partners of prohibitin and were analyzed further using mass spectrometry. Coomassie blue stained gel shows immunoprecipitated proteins from oxidative stress (200 μM H2O2 or tBUO2H) treated ARPE-19 cells (lysate) with anti-prohibitin antibody. W1 to W3 represents wash fractions; E1 to E3 represents elution fractions; Eb represents the protein elution by immunoprecipitated boiling protein-A beads with sample buffer (Eb). Protein standard marker represents the size between 10 to 120 kDa. Elution fractions showed the potential binding partners of prohibitin and were further processed for protein identification. B. Prohibitin interacting proteins in the retina. Coomassie blue stained gel shows immunoprecipitated proteins from control human retinal progenitor (HRP) cells using anti-prohibitin antibody. Cells were harvested and lysed, followed by protein analysis using SDS-PAGE and mass spectrometry. Proteins in W1-W3 lanes represents washing fractions; E1 to E3 represents elution fractions; Eb represents the protein elution by protein-A beads boiling with sample buffer. Elution fractions showed the potential binding partners of prohibitin and were further analyzed using mass spectrometry. (PDF 119 kb)

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Sripathi, S.R., Sylvester, O., He, W. et al. Prohibitin as the Molecular Binding Switch in the Retinal Pigment Epithelium. Protein J 35, 1–16 (2016). https://doi.org/10.1007/s10930-015-9641-y

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