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Abstract
The element Mercury (Hg) is toxic in all forms. Prokaryotes have evolved an efficient detoxification system widely disseminated by transposons and plasmids. This Hg resistance system (mer) encodes transporters that deliver Hg(II) to a specialized mercuric reductase, MerA, that uses NADPH to reduce reactive mercury, Hg(II), to volatile monoatomic Hg(0) vapor which diffuses away. With transcriptomics I examined the whole-cell response to Hg in a model prokaryote, Escherichia coli MG1655 (MG), carrying a 94-kb conjugative plasmid, NR1 (NR), bearing a classic mer operon. Aim 1 examined the burden of NR1 on chromosomal gene expression of MG and the benefit NR1 provided during Hg exposure. Without Hg exposure, NR1 did not evoke expression of any chromosomal genes and the plasmid itself expressed only its replication genes and several associated with transposons. Upon Hg(II) exposure, NR1’s mer operon genes expressed strongly but most other plasmid genes decreased or did not change. However, for many chromosomal genes Hg(II) provoked significant differences between MG and its un-exposed condition. These large transcriptional responses by MG diminished or disappeared in MG(NR). Thus, just hosting NR1 adds no expression in chromosomal genes and with mer proteins cells recovered growth sooner. Previous work suggested E. coli must increase NADPH production to support observed Hg(II) reduction by MerA. Aim 2 examined expression of chromosomal NADPH-producing enzymes. The only increase (30%) occurred in highly expressed (40,000/cell) TCA cycle enzyme, isocitrate dehydrogenase, ICD, which has a high kcat (72/sec). Those values and RNA-Seq data predicted a mere 30% increase in ICD could produce 18-fold more NADPH than MerA needs to reduce 3 million Hg(II)/min as observed in intact cells. Thus, at typical Hg exposures, MerA’s reductant needs are met by a slight tweak in normal metabolism. Aim 3 focused on 310 MG chromosomal genes of proteins we previously observed as highly vulnerable to binding mercurial compounds. Their transcriptional responses revealed distinct patterns consistent with their metabolic functions, notably respiration, sulfur metabolism, and amino acid biosynthesis. Many Hg-vulnerable proteins have homologs in human mitochondria and could be sensitive reporters of Hg exposure and of the efficacy of detoxification measures.