Bioinformatic and Expression Analyses of Genes Mediating Zinc Homeostasis in Nostoc punctiforme

Zinc homeostasis was investigated in Nostoc punctiforme. Cell tolerance to Zn 2 over 14 days showed that ZnCl2 levels above 22 M significantly reduced cell viability. After 3 days in 22 M ZnCl2, ca. 12% of the Zn 2 was in an EDTA-resistant component, suggesting an intracellular localization. Zinquin fluorescence was detected within cells exposed to concentrations up to 37 M relative to 0 M treatment. Radiolabeled 65 Zn showed Zn 2 uptake increased over a 3-day period, while efflux occurred more rapidly within a 3-h time period. Four putative genes involved in Zn 2 uptake and efflux in N. punctiforme were identified: (i) the predicted Co/Zn/Cd cation transporter, putative CDF; (ii) the predicted divalent heavy-metal cation transporter, putative Zip; (iii) the ATPase component and Fe/Zn uptake regulation protein, putative Fur; and (iv) an ABC-type Mn/Zn transport system, putative zinc ZnuC, ZnuABC system component. Quantitative real-time PCR indicated the responsiveness of all four genes to 22 M ZnCl2 within 3 h, followed by a reduction to below basal levels after 24 h by putative ZIP, ZnuC, and Fur and a reduction to below basal level after 72 h by putative CDF efflux gene. These results demonstrate differential regulation of zinc transporters over time, indicating a role for them in zinc homeostasis in N. punctiforme. Cyanobacteria are photosynthetic prokaryotes, many of which accumulate heavy metals. Consequently, they have attracted interest as a tool for the removal of metals from wastewater. Cyanobacteria, including Anabaena nodosum (10), Nos