mRNAs in the methanogenic archaeon Methanococcus vannielii: numbers, half‐lives and processing

Abstract

Cells from the early exponential growth phase of cultures of the methanogenic archaeon Methanococcus vannielii have been shown to contain c. 180 transcripts of the mcrBDCGA (mcr) operon, c. 100 transcripts of the MvaL1,L10,L12 (Mva) operon, c. 8 transcripts of the argG gene and c. 1 transcript of the secY gene. These values decreased to c. 50 mcr transcripts, c. 30 Mva transcripts, c. 3 argG transcripts and < 1 secY transcript per cell as the cultures entered the stationary phase of growth. Addition of bromo-ethanesulphonate (BES) or removal of H2 inhibited growth and RNA synthesis in vivo and, at 37 degrees C in the presence of BES, the half-lives of the mcr, Mva, argG and secY transcripts were found to be 15 min, 30 min, 57 min and 7 min, respectively. Addition of puromycin, pseudomonic acid or virginiamycin also inhibited growth but did not inhibit transcription. In the presence of puromycin the half-lives of the mcr and Mva transcripts increased c. 4.6-fold and c. 3.5-fold, respectively, and there was a net accumulation of the Mva transcript. Addition of pseudomonic acid or virginiamycin also increased the half-life of the Mva transcript and also resulted in the accumulation of a second, shorter Mva transcript but did not increase the half-life of the mcr transcript. Transcription of the mcr operon was not stimulated by partial inhibition of methanogenesis.

Growth phase-dependent transcription of the genes that encode the two methyl coenzyme M reductase isoenzymes and N5-methyltetrahydromethanopterin:coenzyme M methyltransferase in Methanobacterium thermoautotrophicum delta H

“…Transcription of the genes that encode MRI (the mcr operon) and MRII (designated the mrt operon, for methyl reductase two [31,32]) might therefore be differentially regulated by growth conditions in M. thermoautotrophicum strains. This is in contrast to the results recently reported for Methanococcus vannielii, which apparently contains only one MR-encoding region, which is transcribed under all growth conditions (14). To address this directly, the sequences of both the mcr and mrt operons were needed from the same M. thermoautotrophicum strain to identify sequences that could be used as transcript-specific probes.…”

Mechanism for Stabilizing mRNAs Involved in Methanol-Dependent Methanogenesis of Cold-Adaptive Methanosarcina mazei zm-15

“…2, mtaA1, mtaC1 plus mtaB1, and pta plus ackA constituted three separate operons. Next, using RT-qPCR, the in vivo halflives of mtaA1, mtaC1B1, and pta-ackA transcripts were determined in the 30°C and 15°C cultures after inhibiting transcription with 100 g/ml actinomycin D according to the method of Hennigan and Reeve (30). The results showed that mtaA1 and mtaC1B1 were very stable in the cultures grown at both temperatures, with half-lives of about 1 h. In contrast, the half-life of ptaackA was relatively short (25 min) at 30°C and even shorter (15.5 min) at 15°C (Fig.…”

Sequence and transcript analysis of a novel Methanosarcina barkeri methyltransferase II homolog and its associated corrinoid protein homologous to methionine synthase

“…Degenerate oligonucleotide probes were constructed with the N-terminal sequences of both subunits of the 480-kDa CoM methylase (55) RNA isolation and Northern hybridizations. RNA was prepared and Northern blotting was performed by modifications of the techniques described by Hennigan and Reeve (27) and Pihl et al (45). M. barkeri was grown to the mid-log phase and harvested by filtering through Whatman no.…”