A temperature sensitive mutant screen reveals translational stress-induced cell cycle regulation in a thermophilic archaeon

Abstract

The homology of the archaeal and eukaryotic ribosome provides one of the key pieces of evidence that underpins the idea that eukaryotes acquired their core information processing machinery from archaea. Since this discovery, reverse genetics has been used to study the functions of many archaeal proteins with eukaryotic homologues. Yet, our general understanding of archaeal growth and division remains unclear, in part because of difficulties of carrying out unbiased genetic screens in archaea. Here, by overcoming several technical hurdles we have used a screen of temperature sensitive mutants in Sulfolobus acidocaldarius to identify core regulators of cell growth and division. First, flow cytometry was used to define DNA content, identifying a set of mutants defective in cell cycle progression at elevated growth temperatures. Using genome sequencing and plasmid rescue, we then identified a point mutation in the large ribosomal subunit that inhibits translation and prevents entry into division following a shift to the restrictive temperature. This study reveals a link between translation and cell cycle control, and opens up the future possibility of using forward genetic screens in archaea to further our understanding of the similarities and differences in the cell biology of archaea, bacteria and eukaryotes. Significance statementO_LICurrent knowledge of archaeal cell biology is limited by the lack of forward genetics. C_LIO_LIWhole genome sequencing and plasmid rescue identifies causative mutation in a temperature sensitive mutant strain. C_LIO_LIA mutation in a ribosomal subunit blocks translation to prevent entry into division. C_LI