OTHER FUN DISCOVERIES
In addition to mPOS, we contributed to the understanding of mtDNA recombination/repair/replication, the biology of mitochondrial carrier proteins, the development of molecular biology tools in Kluyveromyces lactis
1. mtDNA recombination/replication/repair and maintenance - We discovered the MGM101 (Mitochondrial Genome Maintenance 101) gene by forward genetic screen for mtDNA maintenance genes in 1993. We then found that Mgm101 is a Rad52-family protein of bacteriophage origin that is essential for mtDNA recombination and replication. We also found that the Krebs-cycle enzyme, aconitase, is essential for maintaining mtDNA integrity in yeast.
REFERENCES:
Chen, X.J., Guan, M.X. and Clark-Walker, G,D. (1993) MGM101, a nuclear gene involved in maintenance of the mitochondrial genome in Saccharomyces cerevisiae. Nucleic Acid Res. 21:3473-3477.
Zuo, X.M., Clark-Walker, G.D. and Chen, X.J. (2002). The mitochondrial nucleoid protein, Mgm101p, of is involved in the maintenance of rho+ and ori/rep-devoid petite genomes but is not required for hypersuppressive rho- mtDNA. Genetics 160:1389-1400.
Chen, X.J. and Butow, R.A. (2005) Organization and inheritance of mitochondrial nucleoids. Nature Review/Genetics 6:815-825.
Chen, X.J., Wang, X.W., Kaufman, B.A. and Butow, R.A. (2005) Aconitase couples metabolic regulation to mitochondrial DNA maintenance. Science 307:714-717. (for comments, see: http://www.jcb.org/cgi/content/full/168/5/675-a; Shadel (2005) Trend in Biochemical Sciences 30:294-296)
Chen, X.J. (co-corresponding author), Wang, X.W. and Butow R.A. (2007) Yeast aconitase binds and provides metabolically coupled protection to mitochondrial DNA. PNAS 104:13738-13743.
Mbantenkhu, M.*, Wang, X.* (* equal contribution), Nardozzi, J.D., Wilkens, S., Hoffman, E., Patel, A., Cosgrove, M.S. and Chen, X.J. (2011) Mgm101 is a Rad52-related protein required for mitochondrial DNA recombination. J. Biol. Chem. 286:42360-42370.
Nardozzi, J.D. *, Wang, X.* (* equal contribution), Mbantenkhu, M., Wilkens, S. and Chen, X.J. (2012) A properly configured ring structure is critical for the function of the mitochondrial DNA recombination protein Mgm101. J. Biol. Chem. 287:37259-37268.
Mbantenkhu, M., Wierzbicki, S., Wang, X., Guo, S., Wilkens, S. and Chen, X.J. (2013) A short carboxyl-terminal tail is reauired for ssDNA-binding, higher order structural organization and stability of the mitochondrial single strand annealing protein Mgm101. Molecular Biology of the Cell 24:1507-18.
Wang, X., Mbantenkhu, M., Wierzbicki, S. and Chen, X.J. (2013) Preparation of the Mgm101 recombination protein by MBP-based tagging strategy. Journal of Visualized Experiments. J. Vis. Exp. Jun 25; (76). doi: 10.3791/50448.
Chen, X.J. (2013) Mechanism of homologous recombination and aging-related deletions in mitochondrial DNA. Microbiol. Mol. Biol. Reviews 77:476-96.
2. Sal1 - coupling cytosolic calcium signaling with mitochondrial biogenesis - We discovered a novel pathway that couples cytosolic calcium-signaling with mitochondrial protein synthesis and biogenesis. We found that the yeast Sal1 (for Suppressor of AAC2-lethality 1), homologous to the human SLC25A25 transporter on the mitochondrial inner membrane, is required for mitochondrial protein synthesis and biogenesis in a calcium-dependent manner. The Sal1 activity overlaps with a novel function (the “V function”) associated with the adenine nucleotide translocase (Ant), which is distinct from the ATP/ADP exchange (and therefore respiratory) function (the R function). Sal1/SLC25A25 is believed to play a role in the net import of adenine nucleotides into mitochondria, as suggested by biochemical studies from other groups. Then, does Aac2/Ant also promote net accumulation of adenine nucleotide inside mitochondria? Who knows.
REFERENCES:
Chen, X.J. (2004) Sal1p, a calcium-dependent carrier protein that suppresses an essential cellular function associated with the Aac2p isoform of ADP/ATP translocase in Saccharomyces cerevisiae. Genetics 167:607-617
Kucejova, B., Li, L., Wang, X.W., Giannatasio, S. and Chen, X.J. (2008) Pleiotropic effects of the yeast Sal1 and Aac2 carriers on mitochondrial function via an activity distinct from adenine nucleotide transport. Molecular Genetics and Genomics 280:25-39.
3. Characterization of pKD1 and discovery of pKW1 - In the Fukuhara lab, I contributed to the characterization of the newly discovered yeast plasmid pKD1, by sequencing half of the plasmid (2.4 Kb) using the Maxam-Gilbert method. This was completed within 8 months! I then constructed a series of pKD1-based shuttle vectors that are now widely used in the Kluyveromyces research community. These molecular tools also facilitated our discovery of the gain-of-function F1-ATPase mutants that suppress rho-zero lethality in K. lactis.
After my Ph.D work, I serendipitously discovered a novel yeast plasmid during my time as a post-doc at Institut Curie, when running K. waltii DNA on an agarose gel on the 1988 Bastille day. The plasmid was named pKW1. For the first time, I appreciated that discovering and naming something new in the nature is REALLY fun.
REPRESENTATIVE PUBLICATIONS:
Chen, X.J., Saliola, M., Falcone, C., Bianchi, M.M. and Fukuhara, H. (1986) Sequence organization of the circular plasmid pKD1 from the yeast Kluyveromyces drosophilarum. Nucl. Acids Res. 14:4471-4481.
Chen, X.J., Cong, Y.S., Li, Y.Y. and Fukuhara, H. (1992) Characterization of a circular plasmid from the yeast Kluyveromyces waltii. J. Gen. Microbiol. 138:337-145.
