Proteomic and genetic analysis of S. cerevisiae response to soluble copper leads to improvement of antimicrobial function of cellulosic copper nanoparticles

摘要

Copper (Cu) was used in antiquity to prevent waterborne and food diseases because, as a broad-spectrum antimicrobial agent, it generates reactive oxygen species, ROS. New technologies incorporating Cu into low-cost biodegradable nanomaterials built on cellulose, known as cellulosic cupric nanoparticles or c-CuNPs, present novel approaches to deliver Cu in a controlled manner to control microbial growth. We challenged strains of Saccharomyces cerevisiae to soluble Cu and c-CuNPs to evaluate the potential of c-CuNPs as antifungal agents. Cells exposed to c-CuNPs demonstrated greater sensitivity to Cu than cells exposed to soluble Cu, although Cu-resistant strains were more tolerant than Cu-sensitive strains of c-CuNP exposure. At the same level of growth inhibition, 157 μM c-CuNP led to the same internal Cu levels as did 400 CuSO4, offering evidence for alternative mechanisms of toxicity, perhaps through β-arrestin dependent endocytosis, which was supported by flow cytometry and fluorescence microscopy of c-CuNPs distributed both on the cell surface and within the cytoplasm. Genes responsible for genetic variation to copper were mapped to the ZRT2 and the CUP1 loci. Through proteomic analyses, we found that the expression of other zinc (Zn) transporters increased in Cu-tolerant yeast compared to Cu-sensitive strains. Further, the addition of Zn at low levels increased the potency of c-CuNP to inhibit even the most Cu-tolerant yeast. Through unbiased systems biological approaches, we identified Zn as a critical component of yeast response to Cu and the addition of Zn increased potency of the c-CuNPs.