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We fully annotated two large plasmids, pMOL28 (164 open reading frames [ORFs]; 171,459 bp) and pMOL30 (247 ORFs; 233,720 bp), in the genome of Cupriavidus metallidurans CH34. pMOL28 contains a backbone of maintenance and transfer genes resembling those found in plasmid pSym of C. taiwanensis and plasmid pHG1 of C. eutrophus, suggesting that they belong to a new class of plasmids. Genes involved in resistance to the heavy metals Co(II), Cr(VI), Hg(II), and Ni(II) are concentrated in a 34-kb region on pMOL28, and genes involved in resistance to Ag(I), Cd(II), Co(II), Cu(II), Hg(II), Pb(II), and Zn(II) occur in a 132-kb region on pMOL30. We identified three putative genomic islands containing metal resistance operons flanked by mobile genetic elements, one on pMOL28 and two on pMOL30. Transcriptomic analysis using quantitative PCR and microarrays revealed metal-mediated up-regulation of 83 genes on pMOL28 and 143 genes on pMOL30 that coded for all known heavy metal resistance proteins, some new heavy metal resistance proteins (czcJ, mmrQ, and pbrU), membrane proteins, truncated transposases, conjugative transfer proteins, and many unknown proteins. Five genes on each plasmid were down-regulated; for one of them, chrI localized on pMOL28, the down-regulation occurred in the presence of five cations. We observed multiple cross-responses (induction of specific metal resistance by other metals), suggesting that the cellular defense of C. metallidurans against heavy metal stress involves various regulons and probably has multiple stages, including a more general response and a more metal-specific response.

Plasmids pMOL28 and pMOL30 of Cupriavidus metallidurans Are Specialized in the Maximal Viable Response to Heavy Metals