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Dissertation Final for Ohio link.pdf (9.57 MB)

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OXIDATION OF SELECTIVE MRNAS CORRESPONDING TO MITOCHONDRIAL ETC COMPLEX SUBUNITS DYSREGULATE ENERGY PRODUCTION IN NEURODEGENERATIVE DISEASES

Mahendran, Thulasi
http://rave.ohiolink.edu/etdc/view?acc_num=kent1710246753270272

Abstract Details

2024, PHD, Kent State University, College of Arts and Sciences / Department of Chemistry and Biochemistry.
Mitochondria are the major sites of cellular energy production. Electrons from reduced metabolites flow through the ETC consisting of complexes I through IV that creates the mitochondrial membrane potential (MMP) which is harnessed by complex V, culminating in ATP synthesis. Any disruption in the activity of the ETC complexes hampers formation of MMP and consequently ATP synthesis. ETC in the inner mitochondrial membrane during oxidative phosphorylation also serves as the primary source of reactive oxygen species (ROS), which if not fully neutralized causes oxidation of major biomolecules. While the consequences of DNA, protein and lipid oxidation have been explored, the effect of RNA oxidation on cellular processes in general is rather poorly understood. It’s all the more important because the mechanisms for repair of oxidized RNA are still under debate. Various diseases including major neurological ailments and certain cancers are associated with RNA oxidation. Also, mitochondrial dysfunction is a hallmark of major neurodegenerative diseases including, Parkinson’s disease (PD), Alzheimer’s disease (AD), and Multiple Sclerosis (MS). However, the connection between the mitochondrial dysfunction and RNA oxidation is yet to be investigated. We hypothesize that “mitochondrial dysfunction can be a result of oxidation of the mRNAs encoding the subunits of the ETC complexes, which would impede subunit production leading to compromised ETC and decreased ATP production.” To systematically and comprehensively address the hypotheses, we focused on addressing three specific aims in this study: i) identify the oxidized mitochondrial mRNAs in neuronal cells, and determine if they lead to lowering of the cognate protein subunit levels and measure function of the ETC complexes and ATP level, ii) investigate if nuclear mRNAs encoding ETC complex subunits are oxidized and consequence of that on function of ETC complexes and ATP synthesis, and iii) detect oxidized mRNAs in EAE mice, post mortem MS and PD brain samples, determine if corresponding protein subunit production decreases and ramifications of that on mitochondrial ETC function and ATP level. Among the ETC complex subunits, 13 mRNAs are mitochondrially encoded while about 80 are nuclear encoded, and all of them were analyzed in this study to determine their level of oxidation under an oxidative environment. Utilizing differentiated human neuronal SH-SY5Y cells as a model system, we uncovered that four mRNAs namely ND2, ND4, ND5 and ND6 encoded by the mitochondrial genome get oxidized when cells were induced with oxidative stress. In addition, a set of nuclear encoded mRNAs (11) of ETC subunits were also identified as highly oxidized using RNA deep sequencing of mitochondrial oxidized mRNAs. We established that increased oxidation of both mitochondrial and nuclear encoded mRNAs impedes protein synthesis of ETC complex subunits using in vitro, in organello and in cellulo assays. We have shown that the oxidation of ETC complex subunit mRNAs downregulates the activities of ETC complexes I, III and IV as well as MMP and ATP synthesis in cultured neuronal cells. Reduced ATP level compromises neuronal cell viability. Apart from ETC mRNAs, we identified the oxidation of several other mRNAs related to neurodegenerative diseases including SNCA, HSPA and MAPK8. Furthermore, we observed that the oxidized mRNAs were stable as their non oxidized counterparts at least until 24 hrs inside neuronal cells as determined by RT-qPCR analysis of the transfected transcripts. We also tested our hypothesis in EAE mouse model for MS and postmortem MS and PD brain samples, where we were able to demonstrate the presence of ETC subunits’ mRNA oxidation in association with loss of complex activities and tissue ATP levels presumably contributing to disease progression. The comparison of mRNA oxidation data between SH-SY5Y cells and PD brain revealed 32 targets related to neurodegeneration is common between the two data sets. Knowledge gained from these studies will begin to mechanistically explain the biochemical consequences of oxidation of ETC complex subunit encoding mRNAs on its disruption and ATP production. A broader impact of this study is, while the focus is on MS and PD, the established strategy and framework could be extended to other neurodegenerative diseases associated with RNA oxidation and mitochondrial dysfunction.
Soumitra Basu (Committee Chair)
Sanjaya Abeysisrigunawardena (Committee Member)
Yaorong Zheng (Committee Member)
John Johnson (Committee Member)
Jennifer McDonough (Committee Member)
243 p.
Biochemistry; Molecular Biology
Neurodegeneration; oxidative stress; mitochondrial dysfunction; reactive oxygen species; RNA oxidation; ETC complexes; 8-oxoG; ATP synthesis

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Citations

  • Mahendran, T. (2024). OXIDATION OF SELECTIVE MRNAS CORRESPONDING TO MITOCHONDRIAL ETC COMPLEX SUBUNITS DYSREGULATE ENERGY PRODUCTION IN NEURODEGENERATIVE DISEASES [Doctoral dissertation, Kent State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=kent1710246753270272

    APA Style (7th edition)

  • Mahendran, Thulasi. OXIDATION OF SELECTIVE MRNAS CORRESPONDING TO MITOCHONDRIAL ETC COMPLEX SUBUNITS DYSREGULATE ENERGY PRODUCTION IN NEURODEGENERATIVE DISEASES. 2024. Kent State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=kent1710246753270272.

    MLA Style (8th edition)

  • Mahendran, Thulasi. "OXIDATION OF SELECTIVE MRNAS CORRESPONDING TO MITOCHONDRIAL ETC COMPLEX SUBUNITS DYSREGULATE ENERGY PRODUCTION IN NEURODEGENERATIVE DISEASES." Doctoral dissertation, Kent State University, 2024. http://rave.ohiolink.edu/etdc/view?acc_num=kent1710246753270272

    Chicago Manual of Style (17th edition)

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This open access ETD is published by Kent State University and OhioLINK.
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