Dr. Matthew J. Young received his PhD from the University of Manitoba in 2008 under the supervision of Dr. Deborah A. Court. His PhD work included determining the function of a unique domain found in the yeast mitochondrial DNA (mtDNA) polymerase gamma (γ) and determining the roles of genetic background on yeast mtDNA maintenance. MtDNA polymerase γ is the enzyme that replicates mtDNA. Dr. Young completed his postdoctoral work in 2015 under the supervision of Dr. William C. Copeland at the National Institute of Environmental Health Sciences (NIEHS). During his post-doctoral studies, Dr. Young investigated the molecular effects of various disease mutations in a subunit of the human mtDNA polymerase γ (pol γ). Also, during this time Dr. Young developed cell line models of mitochondrial disease. Dr. Matthew Young joined the faculty in the Department of Biochemistry and Molecular Biology at Southern Illinois University School of Medicine in 2015.
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Surprisingly, mtDNA mutations are common in the population and it has been estimated that one in 200 healthy humans harbor a pathogenic mtDNA mutation that potentially causes disease in the offspring of female carriers (Elliott et al. AJHG 2008). Other researchers have estimated the incidence of mitochondrial disease to be about one in 5000 (Wallace and Chalkia. CSH Perspect Biol 2013) and mitochondrial dysfunction is at the root of genetic disorders that affect both older adults and young children. Other diseases associated with mitochondrial dysfunction include Parkinson's, Huntington's, Alzheimer's, ALS, epilepsy, and cancer. For these reasons the Young laboratory is energized to determine the mechanisms of respiratory dysfunction in cell line models of mitochondrial disease.
Human mtDNA is maternally inherited and occurs as a double stranded negatively supercoiled circular genome of 16,569 base pairs (bp) that encodes 37 genes required for energy production. The remaining mitochondrial genome includes 1158 nuclear genes, and mutations in a class of these genes are associated with Mendelian disorders. When cells in the body lose their mtDNA, or mtDNA becomes mutated, mitochondria become dysfunctional and mitochondrial disease ensues. Human mtDNA is a multicopied genome occurring at about 1000 copies per cell and these molecules are replicated and repaired by pol γ. Pol γ is a trimer composed of subunits encoded by two nuclear genes:
1) The POLG gene encodes the monomeric catalytic subunit, p140 and
2) The POLG2 gene encodes the homodimeric processivity subunit, p55.
Mitochondrial diseases, including those linked to POLG and POLG2 mutations, are devastating disorders that comprise a continuum of overlapping phenotypes and the age of onset of these diseases range from early childhood to late adulthood. Currently, there are no cures for these disorders. One barrier to the field of mitochondrial disease research is the absence of cell line models harboring disease mutations. The Young laboratory aims to develop and characterize cell line models of mitochondrial disease to determine and characterize cellular mechanisms of respiratory dysfunction.
Knockdown and over-expression studies in human cell lines have demonstrated that the human p55 pol γ subunit regulates the content of mtDNA within mitochondrial nucleoids (Di Re et al. NAR 2009). Mitochondrial nucleoids are discrete regions of protein-mtDNA interactions that occur along the matrix side of the mitochondrial inner membrane. Also, the p55 dimer confers processivity onto the Pol γ holoenzyme. Processivity is a measurement of the extent of DNA synthesis during a single enzyme-binding event. Pol γ is the replicative mtDNA polymerase that also participates in mtDNA repair. Pol γ functions in conjunction with a number of additional components in the mtDNA replication machinery or replisome (Figure 1). Nuclear mutations associated with mtDNA maintenance disorders have also been discovered in genes encoding other components of the mtDNA replication machinery that include: the Twinkle mtDNA helicase, the helicase/nuclease DNA2, the 5'-3' MGME1 exonuclease, and the enzyme required to remove RNA primers, RNaseH1 (Figure 1).
Figure 1. The human mtDNA replication fork. The cartoon illustrates the mtDNA replication machinery or replisome. Black lines represent template mtDNA while green lines represent nascent mtDNA. Main factors highlighted at the replication fork include: 1) the 5’-3’ mtDNA polymerase, Pol γ 2) the Top2β topoisomerase or gyrase (Topo) required for mtDNA unwinding ahead of the replication fork. The phosphodiester backbones of both mtDNA strands are enzymatically broken and rejoined allowing relaxation of positive supercoils introduced ahead of the replisome during replication fork elongation, 3) the hexameric replicative Twinkle mtDNA helicase, which is required for ATP-dependent disruption of the hydrogen bonds that hold the two DNA strands together thereby denaturing the mtDNA duplex (strand separation), 4) mitochondrial RNA polymerase (mtRNAP) required for mitochondrial transcription as well as for RNA primer formation to initiate DNA replication, 5) RNaseH1 required for RNA primer removal, 6) mitochondrial single-stranded DNA (ssDNA) binding protein (mtSSB) required for ssDNA stabilization during mtDNA replication, 7) DNA ligase III (mtLigIII) required for mtDNA break (nick) sealing, 8) mitochondrial genome maintenance 5’-3’ exonuclease 1 (MGME1), 9) flap endonuclease (FEN1), and 10) the helicase/nuclease, DNA2.
The mission of the Young laboratory is to identify toxicants that disrupt mtDNA maintenance and to identify therapies for mitochondrial dysfunction. The aims of this research include:
1) To design and exploit cell line models of mitochondrial disease to determine mechanisms of respiratory defects.
2) To determine how disease variant subunits of the replisome impact nucleoid organization.3) To investigate mitochondrion-environment interactions and the implications on mtDNA replication.
Young MJ and Copeland WC. Human mitochondrial DNA replication machinery and disease. Curr Opin Genet Dev. Apr 8;38:52-62. 2016. PMID: 27065468.
Young MJ, Humble MM, DeBalsi KL, Sun KY, and Copeland WC. POLG2 disease variants: analyses reveal a dominant negative heterodimer, altered mitochondrial localization and impaired respiratory capacity. Hum Mol Genet. Sept15;24(18), 5184-5197. 2015. PMID: 26123486.
Craig K+, Young MJ+, Blakely EL, Longley MJ, Turnbull DM, Copeland WC, and Taylor RW. A p.R369G POLG2 mutation associated with adPEO and multiple mtDNA deletions causes decreased affinity between polymerase γ subunits. Mitochondrion. Mar:12(2):313-9. 2012. PMID: 22155748. +MJY and CK contributed equally to this work.
Bay DC, Hafez M, Young MJ, Court DA. Phylogenetic and coevolutionary analysis of the β-barrel protein family comprised of mitochondrial porin (VDAC) and Tom40. Biochim Biophys Acta. Jun; 1818(6):1502-19. Dec 4. 2012. PMID: 22178864.
Young MJ, Longley MJ, Li FY, Kasiviswanathan R, Wong LJ, and Copeland WC. Biochemical analysis of human POLG2 variants associated with mitochondrial disease. Hum Mol Genet. Aug 1:20(15):3052-66. Epub May 9. 2011. PMID: 21555342. *Article highlighted on the cover.
Kasiviswanathan R, Longley MJ, Young MJ, Copeland WC. Purification and functional characterization of human mitochondrial DNA polymerase gamma harboring disease mutations. Methods. Aug;51(4):379-84. 2010. PMID: 20176107.
Young MJ and Court DA. Effects of the S288c genetic background and common auxotrophic markers on mitochondrial DNA function in Saccharomyces cerevisiae. Yeast. Dec; 25(12): 903-12. 2008. PMID: 19160453.
Young MJ+, Bay DC+, Hausner G, and Court DA. The evolutionary history of mitochondrial porins. BMC Evol Biol Feb 28;7:31. 2007. PMID: 17328803. +MJY and DCB contributed equally to this work.
Young MJ, Theriault SS, Li M, and Court DA. The carboxyl-terminal extension on fungal mitochondrial DNA polymerases: identification of a critical region of the enzyme from Saccharomyces cerevisiae. Yeast. Jan 30; 23(2):101-16. 2006. PMID: 16491467.
Young MJ and Court DA. Quick Measurement of Glucose Concentration in Saccharomyces cerevisiae Cultures. The Genetics Society of Canada Bulletin. 35(4): 109-110. 2004. Bulletin.
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