Abstract
Copper (Cu)-only superoxide dismutases (SOD) represent a newly characterized class of extracellular SODs important for virulence of several fungal pathogens. Previous studies of the Cu-only enzyme SOD5 from the opportunistic fungal pathogen Candida albicans have revealed that the active-site structure and Cu binding of SOD5 strongly deviate from those of Cu/Zn-SODs in its animal hosts, making Cu-only SODs a possible target for future antifungal drug design. C. albicans also expresses a Cu-only SOD4 that is highly similar in sequence to SOD5, but is poorly characterized. Here, we compared the biochemical, biophysical, and cell biological properties of C. albicans SOD4 and SOD5. Analyzing the recombinant proteins, we found that, similar to SOD5, Cu-only SOD4 can react with superoxide at rates approaching diffusion limits. Both SODs were monomeric and they exhibited similar binding affinities for their Cu cofactor. In C. albicans cultures, SOD4 and SOD5 were predominantly cell wall proteins. Despite these similarities, the SOD4 and SOD5 genes strongly differed in transcriptional regulation. SOD5 was predominantly induced during hyphal morphogenesis, together with a fungal burst in reactive oxygen species. Conversely, SOD4 expression was specifically up-regulated by iron (Fe) starvation and controlled by the Fe-responsive transcription factor SEF1. Interestingly, Candida tropicalis and the emerging fungal pathogen Candida auris contain a single SOD5-like SOD rather than a pair, and in both fungi, this SOD was induced by Fe starvation. This unexpected link between Fe homeostasis and extracellular Cu-SODs may help many fungi adapt to Fe-limited conditions of their hosts.
Highlights
Copper (Cu)-only superoxide dismutases (SOD) represent a newly characterized class of extracellular SODs important for virulence of several fungal pathogens
To compare the biochemical and biophysical properties of C. albicans SOD4 and SOD5, recombinant proteins missing the N-terminal signal peptide and C-terminal GPI anchor motif were produced in an Escherichia coli expression system
Based on the differential surface charges of SOD4 versus SOD5 (Fig. 1B) we tested whether SOD4 is monomeric or could form higher order complexes
Summary
The biochemical and biophysical properties of recombinant SOD4 closely parallel SOD5. SOD4, but not SOD5 protein levels were seen to increase in a mixed population of C. albicans yeast-form and hyphal cells under Fe-starvation conditions [36]. It is unclear whether SOD4 mRNA is induced by Fe starvation, as reports have been conflicting [37, 38]. We examined expression of CauSOD4 and CauSOD6 mRNA in laboratory cultures of C. auris These strains cannot form hyphae under laboratory conditions [64], a Festarvation state can be induced using BPS (Fig. 8B). Our studies are the first to report a connection between extracellular SODs and Fe limitation for any organism and may be more widespread among microbial pathogens
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