Inactivation of homocitrate synthase causes lysine auxotrophy in copper/zinc-containing superoxide dismutase-deficient yeast Schizosaccharomyces pombe.
The fission yeast Schizosaccharomyces pombe lacking copper/zinc-containing superoxide dismutase (CuZn-SOD) is auxotrophic for lysine and sulfurous amino acids under aerobic growth conditions. A multicopy suppressor gene (phx1+) that restored the growth of CuZn-SOD-deficient cells on minimal medium was isolated. It encodes a putative DNA-binding protein with a conserved homeobox domain. Overproduction of Phx1 increased the amount of several proteins, and one of those turned out to be a putative homocitrate synthase (HCS) encoded by the lys4+ gene in S. pombe as judged by mass spectrometric analysis. Consistent with this observation, overexpression of the lys4+ gene increased HCS enzyme activity and was sufficient to suppress the lysine requirement of the CuZn-SOD-deficient cells. Enzyme activity and Western blot analyses revealed that the activity and protein level of HCS were dramatically reduced upon depletion of CuZn-SOD. Treatment of exponentially growing S. pombe cells with paraquat, a superoxide generator, caused a decrease in the amount of Lys4 protein as expected. These results led us to conclude that HCS, the first enzyme in the alpha-aminoadipate-mediated pathway for lysine synthesis common in fungi and some bacteria, is a labile target of oxidative stress caused by CuZn-SOD depletion and that its synthesis is positively regulated by the putative transcriptional regulator Phx1.
Kwon
Eun-Soo
ES
Laboratory of Molecular Microbiology, School of Biological Sciences, Seoul National University, Korea.
Jeong
Jae-Hoon
JH
Roe
Jung-Hye
JH
D015152:Blotting, Northern; D003001:Cloning, Molecular; D003470:Culture Media; D007700:Kinetics; D008239:Lysine / Q000235:genetics* / Q000378:metabolism*; D007652:Oxo-Acid-Lyases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012568:Schizosaccharomyces / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D029702:Schizosaccharomyces pombe Proteins / Q000037:antagonists & inhibitors / Q000235:genetics; D013482:Superoxide Dismutase / Q000172:deficiency / Q000235:genetics*
D003470:Culture Media; D029702:Schizosaccharomyces pombe Proteins; D013482:Superoxide Dismutase; C021365:homocitrate synthase; D007652:Oxo-Acid-Lyases; D008239:Lysine