Chapter 9 : Noncoding RNA in Mycobacteria

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In the 15 years since the publication of the genome sequence of , analysis of transcriptional regulation has provided a dominant framework for rendering genomic information into understanding of the functional biology of the organism. The powerful combination of molecular biology tools for characterization of individual genetic loci alongside the genome-wide perspective provided by microarray analysis has led to the use of transcription profiles as a surrogate for key phenotypic states implicated in pathogenesis, immunogenicity, and response to drug treatment. As technologies move toward definition of phenotypes by more direct measurement of metabolic status and high-resolution ultrastructural imaging, it is important to develop a rigorous understanding of the quantitative relationship between RNA transcript abundance and broader aspects of cellular physiology, both at the level of bulk populations and at the level of individual cells. The growing awareness—driven largely by the application of high-throughput sequencing technologies to the analysis of RNA (RNA-seq)—that bacteria transcribe much more RNA than is required for direct translation into proteins is likely to be important in this context.

Citation: Arnvig K, Cortes T, Young D. 2014. Noncoding RNA in Mycobacteria, p 183-207. In Hatfull G, Jacobs W (ed), Molecular Genetics of Mycobacteria, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.MGM2-0029-2013
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Figure 1

Venn diagram illustrating how the (nomenclature of) different types of ncRNA and regulatory RNAs overlap and, in particular, how sRNAs can be assigned to more than one category. The sRNA subcategories shown are sRNA (purely regulatory function), sRNA (dual function, i.e., regulatory potential as well as encoding small peptide), sRNA (purely coding, i.e., no function as ribo-regulator). Thus, sRNAs can be coding or noncoding, regulatory or not regulatory. Figure modified from reference .

Citation: Arnvig K, Cortes T, Young D. 2014. Noncoding RNA in Mycobacteria, p 183-207. In Hatfull G, Jacobs W (ed), Molecular Genetics of Mycobacteria, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.MGM2-0029-2013
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Figure 2

Transcription termination in mycobacteria. The top panel illustrates the consensus sequence and structure of the mycobacterial terminator, TRIT (tuberculosis rho-independent terminator). TRIT is a novel rho-independent terminator with high sequence conservation identified in and specific for mycobacteria ( ). The bottom panel illustrates the expression of two converging genes in , according to RNA-seq and visualized in the Artemis genome browser; blue represents expression from the forward strand (), and red represents expression from the reverse strand (); the height of the trace represents the normalized expression level (reads) over the entire region. The traces demonstrate the termination efficiency exerted by TRIT between the two converging genes.

Citation: Arnvig K, Cortes T, Young D. 2014. Noncoding RNA in Mycobacteria, p 183-207. In Hatfull G, Jacobs W (ed), Molecular Genetics of Mycobacteria, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.MGM2-0029-2013
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Figure 3

Ribo-regulation associated with methionine biosynthesis in . At least three enzymes synthesize methionine in ; the expression of two of these is regulated by riboswitches. expression is regulated by a SAM-IV riboswitch, and the MetC enzyme uses homoserine as substrate. expression is regulated by a B riboswitch, and the MetE enzyme uses homocysteine as substrate. The third enzyme, MetH, is a B-dependent isoform of MetE, and MetH also uses homocysteine as substrate; the mRNA of this gene belongs to the category of naturally leaderless mRNAs, which are unusually widespread in ( ). Riboswitch ligands and enzyme cofactors are shown as “stars,” genes are shown in blue, and enzymes are shown as green/yellow “clouds.”

Citation: Arnvig K, Cortes T, Young D. 2014. Noncoding RNA in Mycobacteria, p 183-207. In Hatfull G, Jacobs W (ed), Molecular Genetics of Mycobacteria, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.MGM2-0029-2013
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Figure 4

Magnesium-sensing riboswitches in . The figure illustrates the genomic context of the two identified Mboxes (magnesium-sensing riboswitches) in . Genes shown in green are conserved hypotheticals, those in orange are information pathways, gray represents PE-PPE genes, and genes in blue are cell wall associated. Black arrows indicate relevant transcription start sites.

Citation: Arnvig K, Cortes T, Young D. 2014. Noncoding RNA in Mycobacteria, p 183-207. In Hatfull G, Jacobs W (ed), Molecular Genetics of Mycobacteria, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.MGM2-0029-2013
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Figure 5

The RNA is highly expressed during exponential growth and significantly downregulated in stationary phase in H37Rv. The RNA is expressed in the majority of lineage 4 strains (here represented by H37Rv) and in some lineage 1 strains (represented by N0157), but not in other lineages (N0052—lineage 2—has been shown for comparison). RNA-seq data is visualized in Artemis. All reads are normalized to total reads and adjusted to the same scale.

Citation: Arnvig K, Cortes T, Young D. 2014. Noncoding RNA in Mycobacteria, p 183-207. In Hatfull G, Jacobs W (ed), Molecular Genetics of Mycobacteria, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.MGM2-0029-2013
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Figure 6

-encoded RNAs with -regulating potential. A transcript encoded opposite (antisense) to the gene (ASpks) shows high complementarity to three other mRNAs, namely , , and . The figure illustrates the predicted base-pairing between ASpks and the three mRNAs. From reference .

Citation: Arnvig K, Cortes T, Young D. 2014. Noncoding RNA in Mycobacteria, p 183-207. In Hatfull G, Jacobs W (ed), Molecular Genetics of Mycobacteria, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.MGM2-0029-2013
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Figure 7

RNA-seq data (visualized in Artemis) of the CRISPR locus. The upper trace records TSS mapping (i.e., enriched for primary transcripts) from the right-hand side of the CRISPR locus, showing overlapping start sites for the Rv2816c antisense transcript (forward direction in blue) and the single CRISPR-RNA (crRNA) transcript (reverse, in red). The lower trace records sequencing of total RNA, showing the antisense transcript covering Rv2816c and Rv2817c (blue) and illustrating how the single crRNA transcript is processed into a series of mature, smaller crRNAs (red). A similar crRNA profile is seen on the left-hand side of the CRISPR locus, upstream of the Rv2614c/Rv2615c IS6110 insertion sequence (not shown).

Citation: Arnvig K, Cortes T, Young D. 2014. Noncoding RNA in Mycobacteria, p 183-207. In Hatfull G, Jacobs W (ed), Molecular Genetics of Mycobacteria, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.MGM2-0029-2013
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Table 1

Intergenic sRNAs

Citation: Arnvig K, Cortes T, Young D. 2014. Noncoding RNA in Mycobacteria, p 183-207. In Hatfull G, Jacobs W (ed), Molecular Genetics of Mycobacteria, Second Edition. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.MGM2-0029-2013

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