Nonsense suppression in archaea

Abstract

Bacterial strains carrying nonsense suppressor tRNA genes played a crucial role in early work on bacterial and bacterial viral genetics. In eukaryotes as well, suppressor tRNAs have played important roles in the genetic analysis of yeast and worms. Surprisingly, little is known about genetic suppression in archaea, and there has been no characterization of suppressor tRNAs or identification of nonsense mutations in any of the archaeal genes. Here, we show, using the β-gal gene as a reporter, that amber, ochre, and opal suppressors derived from the serine and tyrosine tRNAs of the archaeon Haloferax volcanii are active in suppression of their corresponding stop codons. Using a promoter for tRNA expression regulated by tryptophan, we also show inducible and regulatable suppression of all three stop codons in H. volcanii. Additionally, transformation of a ΔpyrE2 H. volcanii strain with plasmids carrying the genes for a pyrE2 amber mutant and the serine amber suppressor tRNA yielded transformants that grow on agar plates lacking uracil. Thus, an auxotrophic amber mutation in the pyrE2 gene can be complemented by expression of the amber suppressor tRNA. These results pave the way for generating archaeal strains carrying inducible suppressor tRNA genes on the chromosome and their use in archaeal and archaeviral genetics. We also provide possible explanations for why suppressor tRNAs have not been identified in archaea.nonsense suppression | suppressor tRNA | inducible tRNA expression | haloarchaea | Haloferax volcanii

Why Is the UAG (Amber) Stop Codon Almost Absent in Highly Expressed Bacterial Genes?

“…UGA (opal) non-sense mutants and their suppressors were identified later [ 12 , 13 ]. A few years ago, non-sense suppressors were identified in Haloferax volcanii [ 14 ], thus this is a common mechanism in all domains of life (archaea, bacteria, and eukaryotes). Readthrough of stop codons, whether as an adaptive mechanism or not [ 15 ], has been largely described in eukaryotes and bacteria [ 16 ].…”

Monitoring Physiological Changes in Haloarchaeal Cell during Virus Release

“…The recent isolation of more archaeal viruses [ 5 , 6 ] has offered the possibility of new archaeal virus-host model systems. Nevertheless, the limited set—or complete lack—of genetic tools to manipulate archaeal viruses, e.g., suppression [ 61 , 62 , 63 ], and the extreme conditions required for their propagation pose significant challenges. Consequently, our knowledge of archaeal virus life cycles is nearly non-existent.…”

Transfer RNAs: diversity in form and function

“…Some tRNA variants significantly increase the frequency of mistranslation. Many of these were first identified as suppressor mutations that compensate for nonsense or missense mutations in E. coli, yeast [reviewed in [361][362][363], mammalian cells [364] and archaea [365]. Intergenic suppressor mutations that had broad cellular consequences as a result of altering information flow from DNA to protein were termed informational suppressors [366].…”