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Exploring the role of lipin1 in mitophagy process using lipin1 deficient-EGFP tagged LC3 transgenic mice

Alshudukhi, Abdullah Ali
http://rave.ohiolink.edu/etdc/view?acc_num=wright151377463726015

Abstract Details

2017, Master of Science (MS), Wright State University, Biochemistry and Molecular Biology.
Lipin1 (phosphatidic acid phosphatase) is a key molecule in the cells with two functions: first, it converts phosphatidic acid into diacylglycerol in the cytosol which in turn makes triglycerides. Second, in nucleus lipin1 acts as a transcriptional factor which regulates the expression of genes involved in the fatty acid oxidation and lipid metabolism. Clinically, Lpin1 gene mutations have been detected in patients with severe rhabdomyolysis accompanied with aggregated and dysfunctional mitochondria in their type II muscle fiber. Previously, we have observed that mice with lipin1 deficiency had aggregated mitochondria and abnormal autophagosomes formations by electron microscopy. The mechanism underlying the mitochondrial alterations observed in rhabdomyolysis patients is not revealed yet. Therefore, we think there is a link between impaired mitophagy and lipin1 deficiency in skeletal muscle. Mitochondrial quality control is responsible for regulating the turnover of the mitochondria within the cells. Mitochondrial autophagy (mitophagy) is a part of the mitochondrial quality control and considered as a key process within the eukaryotes. Dysregulated mitophagy has been shown to be involved in Parkinson's disease, cardiovascular diseases and neurodegenerative disorders. In this project we want to examine the role of lipin1 deficiency on the impaired mitophagy process in the muscular tissue. We generated a unique mice model called lipin1-/--EGFP-LC3 transgenic mice to study the mitophagy process. This unique model expresses GFPs tagged to autophagosomes in mitophagy. After analyzing the tissue specimens from the mice we observed increased autophagosomes vacuoles in the lipin1-/- -EGFP-LC3 transgenic mice in the glycolytic muscles fibers compared to the wild type (WT). These autophagosomes represent mitochondrial autophagy. The mitophagy in this context is mediated by Bnip3 (a mitochondrial damage marker) as we found it to be significantly upregulated in the lipin1-/- -EGFP-LC3 transgenic mice. We further treated the mice animals, WT and lipin1-/- -EGFP-LC3 transgenic mice, with chloroquine diphosphate (CQ) which act as autophagy inhibitor and cyclosporine A (CsA), which has been commonly used to improve mitochondrial membrane potential. CQ treatment inhibited the mitophagy and resulted in increased accumulation of LC3 (an autophagy marker) and Bnip3 in the both WT and lipin1-/- -EGFP-LC3 transgenic mice. CsA treatment was effective in increasing the mitochondrial membrane potential which led to decrease expression of the LC3 and Bnip3 in the cytosol of both WT and lipin1-/- -EGFP-LC3 transgenic mice. Lipin1 deficiency upregulated AMP-activated protein kinase (AMPK) in fasting status through a stress produced by nutrients imbalance. AMPK activates FOXO3 (a transcription factor belongs to the forkhead box O-3 family) which has been shown to increase transcription of LC3 and Bnip3 genes. We found that the gene expressions of the LC3 and Bnip3 of lipin1-/- -EGFP-LC3 transgenic mice were slightly increased compared to WT suggesting that the accumulated proteins in the cytoplasm was partially due to increased transcription activity of these molecules. Also, we do not rule out that the increased protein expression in the cytosol is caused by blockage in the mitophagy process as well. Based on these findings, lipin1 deficiency may contribute to the impaired mitophagy in the lipin1-/- -EGFP-LC3 transgenic mice which leads to muscle weakness and atrophy.
Hongmei Ren, Ph.D. (Advisor)
Nicholas Reo, Ph.D. (Committee Member)
Markey Michael, Ph.D. (Committee Member)
69 p.
Biochemistry; Molecular Biology
Mitophagy; GFP-LC3; Bnip3; Mitochondria

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Citations

  • Alshudukhi, A. A. (2017). Exploring the role of lipin1 in mitophagy process using lipin1 deficient-EGFP tagged LC3 transgenic mice [Master's thesis, Wright State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=wright151377463726015

    APA Style (7th edition)

  • Alshudukhi, Abdullah. Exploring the role of lipin1 in mitophagy process using lipin1 deficient-EGFP tagged LC3 transgenic mice. 2017. Wright State University, Master's thesis. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=wright151377463726015.

    MLA Style (8th edition)

  • Alshudukhi, Abdullah. "Exploring the role of lipin1 in mitophagy process using lipin1 deficient-EGFP tagged LC3 transgenic mice." Master's thesis, Wright State University, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=wright151377463726015

    Chicago Manual of Style (17th edition)

wright151377463726015
515
© 2017, some rights reserved.Creative Commons License Exploring the role of lipin1 in mitophagy process using lipin1 deficient-EGFP tagged LC3 transgenic mice by Abdullah Ali Alshudukhi is licensed under a Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License. Based on a work at etd.ohiolink.edu.
This open access ETD is published by Wright State University and OhioLINK.