During evolution mitochondrial DNA (mtDNA) fragments migrated from the mitochondrial precursor organelle to the nucleus and inserted into nuclear DNA forming the eukaryotic nuclear genome. The migration process is still ongoing in many different species, leading to the disruption of nuclear genes and development of human diseases. We found in yeast the rate of mtDNA fragments migrating to the nucleus increased during the chronological life span (CLS). The CLS is determined by the survival of non-dividing cell populations. We demonstrated that yme mutants (yme;yeast mtDNA escape) with high mtDNA migration rates to the nucleus age faster, whereas mutants with decreased transfer rates live longer (ynt100-1). These phenotypes are dependent on mtDNA in mitochondria. Further, linear mtDNA fragments accumulate with likely nuclear localization in the yme1-1 mutant. The accumulation of the linear mtDNA fragments and the short CLS of the yme1-1 mutant are rescued in the absence of the non-essential DNA ligase Lig4 (Dnl4). Dnl4 is part of the non-homologous-end-joining (NHEJ) pathway, which is required for the capture of mtDNA fragments during the repair of double stranded DNA breaks in yeast. We hypothesize that linear, fragmented mtDNA enters the nucleus and accelerates aging by potentially increasing the rate of gross-chromosomal-rearrangements (GCR) and chromosome loss. The proposed project is using fibroblasts as a model system to investigate whether the findings in yeast are conserved in mammals. Several mouse and rat cell lines were created just before Hurricane Sandy struck New Jersey including the entire UMDNJ-Newark campus. Because of a 23 hours loss in electrical power supply, freezers were thawing and cell culture incubators stopped working. These conditions resulted in the loss of important mammalian cell lines. The primary purpose of this application is to regenerate these cell lines. The project should allow generating essential data which are needed to submit an R01 application, which was prevented by the caused damage of Hurricane Sandy.
In aim 1 we will reconstruct mouse and rat cell lines with reduced expression of Yme1 and Lig4. This experiment has the aim to determine whether the flow of mtDNA fragments to the nucleus exists in mammals and whether it affects aging. In addition we will investigate whether mtDNA fragments insert into chromosomal DNA of fibroblasts during aging and whether that affects aging. This project should be a starting point to understand the impact of mtDNA fragments translocating to the nucleus on nuclear genomic instability and the aging process.
An important function of mitochondria is to provide cellular energy. For this purpose, mitochondria carry out the respiration process that is linked to the generation of cellular energy. Although the majority of the proteins taking part in the respiration process are encoded by nuclear DNA, a few proteins are encoded by its own DNA, mitochondrial DNA (mtDNA). Surprisingly, fragments of mtDNA were found to migrate to the nucleus and get inserted into the nuclear genome. In fact, several mtDNA sequences have been identified throughout the human nuclear genome. The process of mtDNA migration to the nuclear genome is thought to be a mechanism of evolutionary change in eukaryotes. The possibility that these migrations may occur in real- time and cause the disruption of nuclear genes has not received much attention. This proposal should be a starting point to understand how fragmentation of mtDNA, migration of mtDNA to the nucleus, chromosomal abnormality, and the aging process are linked to each other. The ultimate downstream goal is to determine if the transfer of mtDNA to the nucleus correlates with specific diseases, and if this process can be interfered in order to minimize or prevent these diseases.
|Ivessa, Andreas S (2017) Aging: Fewer Obstacles to DNA Replication? Trends Cancer 3:387-390|
|Cabral, Marleny; Cheng, Xin; Singh, Sukhwinder et al. (2016) Absence of Non-histone Protein Complexes at Natural Chromosomal Pause Sites Results in Reduced Replication Pausing in Aging Yeast Cells. Cell Rep 17:1747-1754|
|Goto, Greicy H; Zencir, Sevil; Hirano, Yukinori et al. (2015) Binding of Multiple Rap1 Proteins Stimulates Chromosome Breakage Induction during DNA Replication. PLoS Genet 11:e1005283|