Project description:Cis-encoded antisense RNAs (asRNAs) are widespread along bacterial transcriptomes. However, the role of the vast majority of these RNAs remains unknown, and there is an ongoing discussion as to what extent these transcripts are the result of transcriptional noise. We show, by comparative transcriptomics of 20 bacterial species and one chloroplast, that the number of asRNAs is exponentially dependent on the genomic AT content, and that expression of asRNA at low levels exerts little impact in terms of energy consumption. A transcription model simulating mRNA and asRNA production indicates that the asRNA regulatory effect is only observed above certain expression thresholds, substantially higher than physiological transcript levels. These predictions were verified experimentally by overexpressing 9 different asRNAs in M. pneumoniae. Our results suggest that most of the antisense transcripts found in bacteria are the consequence of transcriptional noise, arising at spurious promoters throughout the genome.
Project description:Cis-encoded antisense RNAs (asRNAs) are widespread along bacterial transcriptomes. However, the role of the vast majority of these RNAs remains unknown, and there is an ongoing discussion as to what extent these transcripts are the result of transcriptional noise. We show, by comparative transcriptomics of 20 bacterial species and one chloroplast, that the number of asRNAs is exponentially dependent on the genomic AT content, and that expression of asRNA at low levels exerts little impact in terms of energy consumption. A transcription model simulating mRNA and asRNA production indicates that the asRNA regulatory effect is only observed above certain expression thresholds, substantially higher than physiological transcript levels. These predictions were verified experimentally by overexpressing 9 different asRNAs in M. pneumoniae. Our results suggest that most of the antisense transcripts found in bacteria are the consequence of transcriptional noise, arising at spurious promoters throughout the genome.
Project description:LlorénsRico2016 - Effects of cis-Encoded antisense RNAs (asRNAs) - Case1
Three
putative effects of the asRNAs were considered in this study: in
case 1
(this
model)
,
the binding of the asRNA to the corresponding mRNA induces
degradation of the duplex. In case 2, the binding of the asRNA to
the mRNA induces degradation of the mRNA, but not of the asRNA.
In case 3, the mRNA and the asRNA bind reversibly to form a
stable duplex, preventing translation of the mRNA. In all the
three cases, binding to the ribosome protects the mRNA from the
effect of the asRNA.
This model is described in the article:
Bacterial antisense RNAs are
mainly the product of transcriptional noise.
Lloréns-Rico V, Cano J,
Kamminga T, Gil R, Latorre A, Chen WH, Bork P, Glass JI, Serrano
L, Lluch-Senar M.
Sci Adv 2016 Mar; 2(3): e1501363
Abstract:
cis-Encoded antisense RNAs (asRNAs) are widespread along
bacterial transcriptomes. However, the role of most of these
RNAs remains unknown, and there is an ongoing discussion as to
what extent these transcripts are the result of transcriptional
noise. We show, by comparative transcriptomics of 20 bacterial
species and one chloroplast, that the number of asRNAs is
exponentially dependent on the genomic AT content and that
expression of asRNA at low levels exerts little impact in terms
of energy consumption. A transcription model simulating mRNA
and asRNA production indicates that the asRNA regulatory effect
is only observed above certain expression thresholds,
substantially higher than physiological transcript levels.
These predictions were verified experimentally by
overexpressing nine different asRNAs in Mycoplasma pneumoniae.
Our results suggest that most of the antisense transcripts
found in bacteria are the consequence of transcriptional noise,
arising at spurious promoters throughout the genome.
This model is hosted on
BioModels Database
and identified by:
MODEL1511170000.
To cite BioModels Database, please use:
BioModels Database:
An enhanced, curated and annotated resource for published
quantitative kinetic models.
To the extent possible under law, all copyright and related or
neighbouring rights to this encoded model have been dedicated to
the public domain worldwide. Please refer to
CC0
Public Domain Dedication for more information.
Project description:LlorénsRico2016 - Effects of cis-Encoded
antisense RNAs (asRNAs) - Case1
Three
putative effects of the asRNAs were considered in this study: in
case 1
,
the binding of the asRNA to the corresponding mRNA induces
degradation of the duplex. In case 2
(this
model)
the binding of the asRNA to the mRNA induces degradation of the
mRNA, but not of the asRNA. In case 3, the mRNA and the asRNA
bind reversibly to form a stable duplex, preventing translation
of the mRNA. In all the three cases, binding to the ribosome
protects the mRNA from the effect of the asRNA.
This model is described in the article:
Bacterial antisense RNAs are
mainly the product of transcriptional noise.
Lloréns-Rico V, Cano J,
Kamminga T, Gil R, Latorre A, Chen WH, Bork P, Glass JI, Serrano
L, Lluch-Senar M.
Sci Adv 2016 Mar; 2(3): e1501363
Abstract:
cis-Encoded antisense RNAs (asRNAs) are widespread along
bacterial transcriptomes. However, the role of most of these
RNAs remains unknown, and there is an ongoing discussion as to
what extent these transcripts are the result of transcriptional
noise. We show, by comparative transcriptomics of 20 bacterial
species and one chloroplast, that the number of asRNAs is
exponentially dependent on the genomic AT content and that
expression of asRNA at low levels exerts little impact in terms
of energy consumption. A transcription model simulating mRNA
and asRNA production indicates that the asRNA regulatory effect
is only observed above certain expression thresholds,
substantially higher than physiological transcript levels.
These predictions were verified experimentally by
overexpressing nine different asRNAs in Mycoplasma pneumoniae.
Our results suggest that most of the antisense transcripts
found in bacteria are the consequence of transcriptional noise,
arising at spurious promoters throughout the genome.
This model is hosted on
BioModels Database
and identified by:
MODEL1511170001.
To cite BioModels Database, please use:
BioModels Database:
An enhanced, curated and annotated resource for published
quantitative kinetic models.
To the extent possible under law, all copyright and related or
neighbouring rights to this encoded model have been dedicated to
the public domain worldwide. Please refer to
CC0
Public Domain Dedication for more information.
Project description:LlorénsRico2016 - Effects of cis-Encoded
antisense RNAs (asRNAs) - Case3
Three putative
effects of the asRNAs were considered in this study: in case 1,
the binding of the asRNA to the corresponding mRNA induces
degradation of the duplex. In case 2, the binding of the asRNA
to the mRNA induces degradation of the mRNA, but not of the
asRNA. In case 3 (this model), the mRNA and the asRNA bind
reversibly to form a stable duplex, preventing translation of
the mRNA. In all the three cases, binding to the ribosome
protects the mRNA from the effect of the asRNA.
This model is described in the article:
Bacterial antisense RNAs are
mainly the product of transcriptional noise.
Lloréns-Rico V, Cano J,
Kamminga T, Gil R, Latorre A, Chen WH, Bork P, Glass JI, Serrano
L, Lluch-Senar M.
Sci Adv 2016 Mar; 2(3): e1501363
Abstract:
cis-Encoded antisense RNAs (asRNAs) are widespread along
bacterial transcriptomes. However, the role of most of these
RNAs remains unknown, and there is an ongoing discussion as to
what extent these transcripts are the result of transcriptional
noise. We show, by comparative transcriptomics of 20 bacterial
species and one chloroplast, that the number of asRNAs is
exponentially dependent on the genomic AT content and that
expression of asRNA at low levels exerts little impact in terms
of energy consumption. A transcription model simulating mRNA
and asRNA production indicates that the asRNA regulatory effect
is only observed above certain expression thresholds,
substantially higher than physiological transcript levels.
These predictions were verified experimentally by
overexpressing nine different asRNAs in Mycoplasma pneumoniae.
Our results suggest that most of the antisense transcripts
found in bacteria are the consequence of transcriptional noise,
arising at spurious promoters throughout the genome.
This model is hosted on
BioModels Database
and identified by:
MODEL1511170002.
To cite BioModels Database, please use:
BioModels Database:
An enhanced, curated and annotated resource for published
quantitative kinetic models.
To the extent possible under law, all copyright and related or
neighbouring rights to this encoded model have been dedicated to
the public domain worldwide. Please refer to
CC0
Public Domain Dedication for more information.
Project description:A major challenge for RNA-seq analysis of gene expression is to achieve sufficient coverage of informative non-ribosomal transcripts. In eukaryotic samples, this is typically achieved by selective oligo(dT)-priming of messenger RNAs to exclude ribosomal RNA (rRNA) during cDNA synthesis. However, this strategy is not compatible with prokaryotes in which functional transcripts are generally not polyadenylated. To overcome this, we adopted DASH (Depletion of Abundant Sequences by Hybridization), initially developed for eukaryotic cells, to improve both the sensitivity and depth of bacterial RNA-seq. DASH employs the Cas9 nuclease to remove unwanted cDNA sequences prior to library amplification. We report the design, evaluation, and optimization of DASH experiments for standard bacterial short-read sequencing approaches, including software for automated guide RNA (gRNA) design for Cas9-mediated cleavage in bacterial rDNA sequences. Using these gRNA pools, we effectively removed rRNA reads (56-86%) in RNA-seq libraries from two different model bacteria, the Gram-negative pathogen Salmonella enterica and the anaerobic gut commensal Bacteroides thetaiotaomicron. DASH works robustly, even with sub-nanogram amounts of input cDNA. Its efficiency, high sensitivity, ease of implementation, and low cost (~$5 per sample) render DASH an attractive alternative to rRNA removal protocols, in particular for material-constrained studies where conventional ribodepletion techniques fail.
Project description:To explore the potential functions of exogenous RNAs in circulation, we transfected several synthetic, double-stranded mature microRNA-like molecules selected from observed exogenous miRNA sequences and some highly abundant exogenous sequences (bacterial rRNAs) that have potential to form pre-miRNA-like secondary structures into a mouse, Dicer-deficient, fibroblast cell line. This provides a good tool for studying the function of miRNAs. By introducing individual miRNA into these cells and avoiding multiple interactions of microRNA and mRNA (Wang et al in preparation) it is possible to interrogate the cells for mRNA levels, which are informative as to specific miRNA function.