Identification of the first transcriptional activator of an archaellum operon in a euryarchaeon. Issue 1 (11th July 2016)

• Record Type: Journal Article
• Authors: Ding, Yan; Nash, John; Berezuk, Alison; Khursigara, Cezar M.; Langelaan, David N.; Smith, Steven P.; Jarrell, Ken F.
• Is Part Of: Molecular microbiology. Volume 102:Issue 1(2016)
• Page Start: 54
• Page End: 70
• Publication Date: 2016-07-11
• Subjects: Molecular microbiology -- Periodicals; 572.829
• DOI: 10.1111/mmi.13444
• Languages: English
• ISSNs: 0950-382X

Abstract

Summary: The archaellum is the swimming organelle of the third domain, the Archaea. In the euryarchaeon Methanococcus maripaludis, genes involved in archaella formation, including the three archaellins flaB1, flaB2 and flaB3, are mainly located in the fla operon. Previous studies have shown that transcription of fla genes and expression of Fla proteins are regulated under different growth conditions. In this study, we identify MMP1718 as the first transcriptional activator that directly regulates the fla operon in M. maripaludis . Mutants carrying an in‐frame deletion in mmp1718 did not express FlaB2 detected by western blotting. Quantitative reverse transcription PCR analysis of purified RNA from the Δmmp1718 mutant showed that transcription of flaB2 was negligible compared to wildtype cells. In addition, no archaella were observed on the cell surface of the Δmmp1718 mutant. FlaB2 expression and archaellation were restored when the Δmmp1718 mutant was complemented with mmp1718 in trans . Electrophoretic motility shift assay and isothermal titration calorimetry results demonstrated the specific binding of purified MMP1718 to DNA fragments upstream of the fla promoter. Four 6 bp consensus sequences were found immediately upstream of the fla promoter and are considered the putative MMP1718‐binding sites. Herein, we designate MMP1718 as EarA, the firste uryarchaeala rchaellumr egulator. Abstract : Analysis of the complete genome sequence of a mutant of the archaeon Methanococcus maripaludis that had stopped production of components of archaella led to the identification of the first transcriptional activator, EarA, for the archaella operon in this major branch of Archaea. Cells deleted for earA do not transcribe the archaella operon and are nonarchaellated. Both in vitro and in vivo experiments demonstrate that EarA binds directly upstream of the promoter for the archaella operon.