MET5 同源基因為隱球菌生長、有性生殖及致病性所必須

Abstract

Sulfur-containing amino acids such as cysteine and methionine are important for cellular physiology. Sulfate assimilation pathway (SAP) is a reduction sequence involved in the biosynthesis of these amino acids from the inorganic oxidized sulfur source. In Fungi, sequential steps of SAP are similar, and enzymes involved in this pathway are also highly conserved. In Saccharomyces cerevisiae, exogenous sulfate is transported from the environment into yeast cells by sulfate transporter, Sul1p and Sul2p, and catalyzed by ATP sulfurylase, Met3p, to form 5’-adenylylsulfate (APS). APS is subsequently phosphorylated by Met14p, APS kinase, to produce 3’-phospho- 5’-adenylylsulfate (PAPS), and then reduced by Met16p (PAPS reductase) to generate sulfite. Finally, the sulfite reductase enzyme complex consisted of α subunit Met10p and β subunit Met5p further reduces sulfite to sulfide. The reduced sulfur ion can then be incorporated into sulfur-containing amino acids or compounds. In this report, we study the roles of the MET5 homologue in the human fungal pathogen Cryptococcus neoformans. The gene disruption construct was created by in vitro transposition and delivered into the wild-type strain by biolistic transformation. Gene disrupted mutants were verified and characterized. C. neoformans met5 mutants were auxotrophic for cysteine, reduced the growth rate, severely attenuated for mating differentiation, failed to produce melanin in vitro, and lost virulence in the alternative insect host model. All the defects were reverted to the wild-type by reintroduction of the intact copy MET5 gene. Consistent with previous reports, our results showed that the components of SAP play important roles in the physiological processes of C. neoformans and maybe serve as potential targets for antifungal therapy.