Characterization of Cu,Zn superoxide dismutase (SOD1) gene knock‐out mutant of Cryptococcus neoformans var. gattii: role in biology and virulence

Abstract

SummaryThe pathogenic yeast Cryptococcus neoformans ( Cn ) var. gattii causes meningoencephalitis in healthy individuals, unlike the better known Cn varieties grubii and neoformans , which are common in immunocompromised individuals. The virulence determinants and mechanisms of host predilection are poorly defined for var. gattii . The present study focused on the characterization of a Cu,Zn superoxide dismutase ( SOD1 ) gene knock-out mutant constructed by developing a DNA transformation system. The sod1 mutant was highly sensitive to the redox cycling agent menadione, and showed fragmentation of the large vacuole in the cytoplasm, but no other defects were seen in growth, capsule synthesis, mating, sporulation, stationary phase survival or auxotrophies for sulphurcontaining amino acids. The sod1 mutant was markedly attenuated in virulence in a mouse model, and it was significantly susceptible to in vitro killing by human neutrophils (PMNs). The deletion of SOD1 also resulted in defects in the expression of a number of virulence factors, i.e. laccase, urease and phospholipase. Complementation of the sod1 mutant with SOD1 resulted in recovery of virulence factor expression and menadione resistance, and in restoration of virulence. Overall, these results suggest that the antioxidant function of Cu,Zn SOD is critical for the pathogenesis of the fungus, but is dispensable in its saprobic life. This report constitutes the first instance in which superoxide dismutase has been directly implicated in the virulence of a fungal pathogen.

Extracellular Superoxide Dismutase in Insects

“…Production of ROS is part of this response, as evidenced for D. melanogaster (7)(8)(9), and the detoxification of ROS is known to be part of virulence mechanisms. SODs, for instance, are used as immune-suppressive toxins by a large number of pathogens, including eukaryotic protozoan parasites (21)(22)(23)(24)(25)(26).…”

“…This is quite surprising as a well known virulence strategy employed by numerous bacterial and fungal pathogens, including pathogens of insects as well as protozoan parasites, is the detoxification of reactive species by antioxidant enzymes, mainly superoxide dismutases (SODs) (21)(22)(23)(24)(25)(26). SODs are one of the most important cellular enzymatic defenses against the detrimental ROS generated by the aerobic metabolism.…”

The superoxide dismutases of Candida glabrata protect against oxidative damage and are required for lysine biosynthesis, DNA integrity and chronological life survival

“…8). This is surprising since previous observations have proposed SODs as virulence factors in bacteria and pathogenic yeasts (Hwang et al, 2002;Langford et al, 2002;Narasipura et al, 2003;Youseff et al, 2012). This evidence indicates that there are additional factors that compensate for the lack of SODs in vivo.…”

“…gattii (Hwang et al, 2002;Narasipura et al, 2003;Youseff et al, 2012); however, the role of the SODs in C. glabrata has been only partially analysed (Cuéllar-Cruz et al, 2009;Roetzer et al, 2011). C. glabrata can withstand twice the concentration of menadione [MD, an oxidant that generates O 2 $2 through redox cycling (Hampsey, 1997)] compared with Sac.…”

Downregulation of fungal cytochrome c peroxidase expression by antifungal quinonemethide triterpenoids

“…Because terpenoids are known to induce an increase in intracellular ROS, we selected genes encoding antioxidant proteins, sod1 and sod2, for the superoxide dismutases that convert superoxide radical to hydrogen peroxide, and ccp1, for the cytochrome c peroxidase that detoxifies hydrogen peroxide (Cox et al 2003;Narasipura et al 2003;Giles et al 2005;Narasipura et al 2005). In the gene expression experiment, the tendency of the anti-oxidant gene expression decreased when the cells were exposed to quinonemethide.…”