Construction of Zn2+/Cd(2+)-tolerant cyanobacteria with a modified metallothionein divergon: further analysis of the function and regulation of smt.

Abstract

This paper reports the (de novo) construction of mutants of Synechococcus PCC 7942 lacking the repressor (SmtB) of the metallothionein gene, smtA. These smtA+/B- cells are more tolerant to elevated [Zn2+] and [Cd2+] than cells containing an intact metallothionein divergon (smt). Previously selected (by step-wise adaptation) Cd(2+)-tolerant mutants contain additional copies of smtA and possibly other undetected mutations. It is now confirmed that these cells also contain a deletion within 'all' copies of smtB and hence fail to revert to wild type following subculture in medium which has not been supplemented with Cd2+ or Zn2+. Northern analysis showed enhanced accumulation of smtA transcripts, even in the absence of added metal ions in these mutants. An increase in the accumulation of Zn2+ is reported in cells containing an intact metallothionein divergon compared to cells deficient in both smtA and smtB. This supports the assumption that SmtA binds Zn2+ within cyanobacterial cells. We also describe the use of the above mentioned mutants to identify additional factors involved in the regulation of transcription from the smtA operator-promoter.