Intrinsic terminators in Mycoplasma hyopneumoniae transcription

Fritsch, Tiago Ebert; Siqueira, Franciele Maboni; Schrank, Irene Silveira

doi:10.1186/s12864-015-1468-6

Cited by 14 publications

(13 citation statements)

References 34 publications

(61 reference statements)

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“…Although the putative stem loops displayed relatively high values of Gibbs free energy (median of −7.5 kcal/mol), these values were comparable to the genome-wide predicted intrinsic terminators in Mycoplasma hyopneumoniae (median of −8.0 kcal/mol), an organism with a similarly high AT content (Fritsch et al 2015). Further, RNA stem loops of bacterial intrinsic transcription terminators are usually followed by the typical 7-8 nt U-tract that promotes RNAP pausing at weak dA-rU DNA-RNA hybrid (Martin and Tinoco 1980;d'Aubenton Carafa et al 1990;Gusarov and Nudler 1999).…”

Section: Transcription Terminationmentioning

confidence: 63%

Transcription apparatus of the yeast killer DNA plasmids: Architecture, function, and evolutionary origin

Sýkora

Pospíšek

Novak

et al. 2018

Preprint

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Transcription of extrachromosomal elements such as organelles, viruses, and plasmids is dependent on cellular RNA polymerase (RNAP) or intrinsic RNAP encoded by these elements. The yeast Kluyveromyces lactis contains killer DNA plasmids that bear putative non-canonical RNAP genes. Here, we describe the architecture and evolutionary origin of this transcription machinery. We show that the two RNAP subunits interact in vivo, and this complex interacts with another two plasmid-encoded proteins -the mRNA capping enzyme, and a putative helicase which interacts with plasmid-specific DNA. Further, we identify a promoter element that causes 5′ polyadenylation of plasmid-specific transcripts via RNAP slippage during transcription initiation, and structural elements that precede the termination sites. As a result, we present a first model of the yeast killer plasmid transcription initiation and intrinsic termination. Finally, we demonstrate that plasmid RNAP and its promoters display high similarity to poxviral RNAP and promoters of early poxviral genes, respectively.K2ORF6p regardless of the presence or absence of nucleic acids which was confirmed by PCR ( Supplementary Figure S3).Taken together, the immunoprecipitation, mass spectrometry, and Western blot results demonstrated the existence of the hypothesized plasmid-specific transcription complex because K2ORF3p, K2ORF4p, K2ORF6p, and K2ORF7p were specifically associated in vivo.This association was independent of nucleic acids. Finally, K2ORF3p, K2ORF6p, and K2ORF7p appeared to form a core transcription complex with relatively strong mutual interactions to which K2ORF4p bound relatively weakly. Putative helicase is associated with plasmid-specific DNA in vivoThe weak binding of K2ORF4p to the other three proteins suggested that the putative helicase may also interact with plasmid DNA. Hence, we performed in vivo chromatin immunoprecipitation. We used the IFO1267_pRKL2-13 strain expressing HA-K2ORF4p and 10.1007/BF00326291, PMID: 2065362 Kämper J, Meinhardt F, Gunge N, Esser K. 1989. In vivo construction of linear vectors based on killer plasmids from Kluyveromyces lactis: selection of a nuclear gene results in attachment of telomeres. Mol Cell Biol 9:3931-3937. 24 Kettenberger H, Armache KJ, Cramer P. 2004. Complete RNA polymerase II elongation complex structure and its interactions with NTP and TFIIS. Mol Cell 16:955-965. Kuznedelov K, Korzheva N, Mustaev A, Severinov K. 2002. Structure-based analysis of RNA polymerase function: the largest subunit's rudder contributes critically to elongation complex stability and is not involved in the maintenance of RNA-DNA hybrid length.

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Section: Transcription Terminationmentioning

confidence: 63%

Transcription apparatus of the yeast killer DNA plasmids: Architecture, function, and evolutionary origin

Sýkora

Pospíšek

Novak

et al. 2018

Preprint

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“…The in silico analysis of transcription regulatory elements was performed in M. hyopneumoniae 7448 genome (INSDC AE017244.1). The identification data previously described for promoter motifs [ 19 ], intrinsic terminators [ 20 ], DNA repeats [ 21 ], and sRNAs [ 23 ] were utilized in this study to evaluate their role in transcription regulation. Each regulatory element was manually mapped in the respective predicted sRNA target gene using Artemis software [ 27 ].…”

Section: Methodsmentioning

confidence: 99%

“…The relative expression was calculated by the 2 -ΔΔCt method [ 31 ]. The threshold cycle (CT) values were normalized to the reference gene MHP7448_0333 [ 19 , 20 ]. Three technical and biological replicates were undertaken for each sRNA or gene evaluated.…”

Section: Methodsmentioning

confidence: 99%

“…M. hyopneumoniae genes are organized in large transcriptional units (TUs) that are continuously transcribed (co-transcribed), with each TU being transcribed in the same direction, with no intervening genes transcribed in the opposite [ 16 , 17 ]. The occurrence of promoter motifs [ 18 , 19 ], intrinsic terminators [ 20 ] and DNA repeat sequences [ 21 ] has already been described in this species. These regulatory elements are located both at the boundaries of TUs and also in the internal genes between several TUs.…”

Section: Introductionmentioning

confidence: 99%

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Global analysis of sRNA target genes in Mycoplasma hyopneumoniae

2018

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BackgroundSmall RNAs (sRNAs) are noncoding molecules that regulate different cellular activities in several bacteria. The role of sRNAs in gene expression regulation is poorly characterized in the etiological agent of porcine enzootic pneumonia Mycoplasma hyopneumoniae. We performed a global analysis of the sRNAs, sRNA target genes and regulatory elements previously identified in their genome and analyzed the expression of some sRNAs and their target genes by quantitative RT-PCR (qPCR) in three different culture conditions.ResultsSeven of the 145 sRNA target genes are organized as monocistronic genes (mCs) while the other 138 sRNA target genes are organized into transcriptional units (TU). The identification of transcriptional regulatory elements (promoter motif, DNA repeat sequence or intrinsic terminator) was verified in 116 of the 145 sRNA target genes. Moreover, the 29 sRNA target genes without regulatory elements revealed the presence of at least one regulatory element in the boundaries of the TU or in other internal genes of the TU. We verified that 16 sRNAs showed differential expression, seven in heat shock condition and 14 in oxidative stress condition. Analysis of the differential expression of the sRNA target genes showed that the tested sRNAs possibly regulate gene expression. The sRNA target genes were up- or down-regulated possibly in response to sRNA only under oxidative stress condition. Moreover, the sRNA target genes are involved in diverse processes of the cell, some of which could be linked to transcription processes and cell homeostasis.ConclusionOur results indicate that bacterial sRNAs could regulate a number of targets with various outcomes, and different correlations between the levels of sRNA transcripts and their target gene mRNAs were found, which suggest that the regulation of gene expression via sRNAs may play an important role in mycoplasma.Electronic supplementary materialThe online version of this article (10.1186/s12864-018-5136-5) contains supplementary material, which is available to authorized users.

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“…hyopneumoniae . The occurrence of transcription units [4, 9, 10], promoters [11, 12] and terminators [13] has already been described in this species, but the existence of other regulatory sequences remains to be elucidated.…”

Section: Introductionmentioning

confidence: 99%

Repetitive Elements in Mycoplasma hyopneumoniae Transcriptional Regulation

et al. 2016

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Transcriptional regulation, a multiple-step process, is still poorly understood in the important pig pathogen Mycoplasma hyopneumoniae. Basic motifs like promoters and terminators have already been described, but no other cis-regulatory elements have been found. DNA repeat sequences have been shown to be an interesting potential source of cis-regulatory elements. In this work, a genome-wide search for tandem and palindromic repetitive elements was performed in the intergenic regions of all coding sequences from M. hyopneumoniae strain 7448. Computational analysis demonstrated the presence of 144 tandem repeats and 1,171 palindromic elements. The DNA repeat sequences were distributed within the 5’ upstream regions of 86% of transcriptional units of M. hyopneumoniae strain 7448. Comparative analysis between distinct repetitive sequences found in related mycoplasma genomes demonstrated different percentages of conservation among pathogenic and nonpathogenic strains. qPCR assays revealed differential expression among genes showing variable numbers of repetitive elements. In addition, repeats found in 206 genes already described to be differentially regulated under different culture conditions of M. hyopneumoniae strain 232 showed almost 80% conservation in relation to M. hyopneumoniae strain 7448 repeats. Altogether, these findings suggest a potential regulatory role of tandem and palindromic DNA repeats in the M. hyopneumoniae transcriptional profile.

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Intrinsic terminators in Mycoplasma hyopneumoniae transcription

Cited by 14 publications

References 34 publications

Transcription apparatus of the yeast killer DNA plasmids: Architecture, function, and evolutionary origin

Transcription apparatus of the yeast killer DNA plasmids: Architecture, function, and evolutionary origin

Global analysis of sRNA target genes in Mycoplasma hyopneumoniae

Repetitive Elements in Mycoplasma hyopneumoniae Transcriptional Regulation

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