A 5′ Leader Sequence Regulates Expression of Methanosarcinal CO Dehydrogenase/Acetyl Coenzyme A Synthase

Abstract

In vivo expression of CO dehydrogenase/acetyl coenzyme A synthase in Methanosarcina spp. is coordinately regulated in response to substrate by at least two mechanisms: differential transcription initiation and early elongation termination near the 3 end of a 371-bp leader sequence. This is the first report of regulation of transcription elongation in the Archaea.Methanogenic species in the genus Methanosarcina are the most metabolically diverse among methanogens, with the ability to grow by CO 2 reduction with H 2 , methyl reduction with H 2 , aceticlastic fermentation of acetate, or methylotrophic catabolism of methanol, methylated amines, and dimethyl sulfide (34). Methanosarcina acetivorans is also reported to grow with carbon monoxide (23). These organisms preferentially use substrates with higher free energy, such as methanol and trimethylamine (TMA), before utilizing acetate, exhibiting diauxic growth in the presence of both types of substrates (4,10,16,21,26). Despite their preference for nonaceticlastic substrates and the relatively lower growth rates and yields of Methanosarcina spp. with acetate, approximately 70% of the biogenic methane produced globally is generated from the catabolism of acetate (19). As a result, regulation of genes involved in the aceticlastic pathway is likely to have a significant impact on anaerobic degradation processes and the global biogenesis of methane.Several enzymes associated with aceticlastic methanogenesis, including acetate kinase, phosphotransacetylase, carbonic anhydrase, and carbon monoxide dehydrogenase/acetyl coenzyme A (acetyl-CoA) synthase (CODH/ACS), are regulated in response to substrates (13,14,17). Expression of the CODH/ ACS operon (cdhABCDE), which catalyzes the catabolism of acetyl-CoA in the aceticlastic pathway, is highly regulated in response to growth on acetate compared with methanol and TMA, but the mechanism(s) of this regulation is not known (28, 31). Two CODH/ACS operons are detected in the annotated genome sequences of M. acetivorans (11), Methanosarcina barkeri (20), and Methanosarcina mazei (7). In contrast, only one cdhABCDE operon has been detected in Methanosarcina thermophila (12). In M. acetivorans, both of these operons appear to be regulated in response to substrate, although the extent of the regulation is greater in one (MA3680) than in the other (MA1016) (17). Northern analysis of CODH/ACS mRNA from Methanosarcina thermophila indicates that the regulation of this CODH/ACS ortholog in response to substrate occurs, at least in part, at the level of mRNA (28). The regulation of this ortholog was corroborated with translational fusions of the M. thermophila CODH/ACS promoter to lacZ (2) and by peptide fragment analysis of a CODH/ACS component, CdhA, in M. mazei (9). This CODH/ACS operon has an unusually long 371-bp 5Ј leader region (28). In contrast, the 5Ј leader sequence of CODH/ACS in the obligately aceticlastic Methanosaeta concilii (formerly "Methanothrix soehgenii") is less than 100 bases long and has no significant sequence simi...

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

“…functions in transcription pretermination of the gene in Methanosarcina thermophila (48). Moreover, the large 5= UTRs are predicted to play multiple roles.…”

sRNA41 affects ribosome binding sites within polycistronic mRNAs in Methanosarcina mazei Gö1

“…5). ACDS is biochemically well characterized in methanoarchaea (Ferry, 2015) and its expression has been shown to be highly regulated on the transcriptional level in a substrate dependent manner (Hovey et al, 2005;Anderson et al, 2009). Besides transcriptional regulation, indications for additional post-transcriptional regulation in response to the substrate were obtained in several methanoarchaea (Anderson et al, 2009;Matschiavelli et al, 2012).…”

“…5, Supporting Information Fig. S5A), is known to be transcriptionally regulated in response to substrate availability (i.e., acetate) (Hovey et al, 2005;Anderson et al, 2009;Matschiavelli et al, 2012). Since M. mazei G€ o1 3A requires long adaptation times for growth on acetate, as sole energy and carbon source, methylotrophic growth on methanol under acetate depletion was chosen to evaluate acetate dependent regulation.…”

Contribution of Transcriptomics to Systems-Level Understanding of MethanogenicArchaea

“…A 5′ UTR was important in the methanogenic substrate-dependent regulation of CODH/ACS activity in Methanosarcina spp. by an uncharacterized mechanism that was likely either early transcriptional termination or endoribonuclease activity against the 5′ UTR [61]. In M. acetivorans , 5′ UTRs are also known to be involved in regulating the expression of different isozymes of methanol specific methyltransferases [62].…”