Genome-wide identification of transcriptional start sites in the haloarchaeon Haloferax volcanii based on differential RNA-Seq (dRNA-Seq)

Babski, Julia; Haas, Karina A.; Näther-Schindler, Daniela; Pfeiffer, Friedhelm; Förstner, Konrad U.; Hammelmann, M; Hilker, Rolf; Becker, Anke; Sharma, Cynthia M.; Marchfelder, Anita; Soppa, Jörg

doi:10.1186/s12864-016-2920-y

Cited by 126 publications

(221 citation statements)

References 93 publications

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“…Data visualization was performed using IGV v2.4.6 [90] and Gaggle Genome Browser [26]. Additionally to Halobacterium salinarum , this genome-wide dataset analysis was performed for: Haloferax volcanii DS2 (PRJNA324298) [8], Methanocaldococcus jannaschii DSM 2661 (PRJNA342613) [91], Thermococcus kodakarensis KOD1 (PRJNA242777) [10] and Thermococcus onnurineus NA1 (PRJNA339284) [92] (Table S10).…”

Section: Methodsmentioning

confidence: 99%

“…Abundant transcript signals established pervasive transcription as a general phenomenon, expanding our knowledge on the universe of non-coding RNAs [5]. Genome-wide mapping of transcription start sites (TSS) in diverse prokaryotes has confirmed that many of these signals are not artifacts but bona fide transcripts [6–8]. …”

Section: Introductionmentioning

confidence: 99%

“…The use of dRNA-seq has allowed precise TSS mapping for most annotated coding sequences (CDS), antisense RNAs (asRNAs) and new intergenic RNAs in bacteria [7,9] and archaea [8,10]. Interestingly, many TSS inside coding regions (internal TSS, iTSS) have been mapped, but to date remain mostly uncharacterized.…”

Section: Introductionmentioning

confidence: 99%

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Internal RNAs overlapping coding sequences can drive the production of alternative proteins in archaea

et al. 2018

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Prokaryotic genomes show a high level of information compaction often with different molecules transcribed from the same locus. Although antisense RNAs have been relatively well studied, RNAs in the same strand, internal RNAs (intraRNAs), are still poorly understood. The question of how common is the translation of overlapping reading frames remains open. We address this question in the model archaeon Halobacterium salinarum. In the present work we used differential RNA-seq (dRNA-seq) in H. salinarum NRC-1 to locate intraRNA signals in subsets of internal transcription start sites (iTSS) and establish the open reading frames associated to them (intraORFs). Using C-terminally flagged proteins, we experimentally observed isoforms accurately predicted by intraRNA translation for kef1, acs3 and orc4 genes. We also recovered from the literature and mass spectrometry databases several instances of protein isoforms consistent with intraRNA translation such as the gas vesicle protein gene gvpC1. We found evidence for intraRNAs in horizontally transferred genes such as the chaperone dnaK and the aerobic respiration related cydA in both H. salinarum and Escherichia coli. Also, intraRNA translation evidence in H. salinarum, E. coli and yeast of a universal elongation factor (aEF-2, fusA and eEF-2) suggests that this is an ancient phenomenon present in all domains of life.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Internal RNAs overlapping coding sequences can drive the production of alternative proteins in archaea

et al. 2018

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“…S2 in the supplemental material). This finding was not surprising, as leaderless transcripts are common to haloarchaea (20,21). An initial analysis was performed in wt cells to assess whether thiamine and/or its metabolic precursors could influence reporter gene expression.…”

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confidence: 80%

ThiN as a Versatile Domain of Transcriptional Repressors and Catalytic Enzymes of Thiamine Biosynthesis

et al. 2017

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Thiamine biosynthesis is commonly regulated by a riboswitch mechanism; however, the enzymatic steps and regulation of this pathway in archaea are poorly understood. Haloferax volcanii, one of the representative archaea, uses a eukaryote-like Thi4 (thiamine thiazole synthase) for the production of the thiazole ring and condenses this ring with a pyrimidine moiety synthesized by an apparent bacterium-like ThiC (2-methyl-4-amino-5-hydroxymethylpyrimidine [HMP] phosphate synthase) branch. Here we found that archaeal Thi4 and ThiC were encoded by leaderless transcripts, ruling out a riboswitch mechanism. Instead, a novel ThiR transcription factor that harbored an N-terminal helix-turn-helix (HTH) DNA binding domain and C-terminal ThiN (TMP synthase) domain was identified. In the presence of thiamine, ThiR was found to repress the expression of thi4 and thiC by a DNA operator sequence that was conserved across archaeal phyla. Despite having a ThiN domain, ThiR was found to be catalytically inactive in compensating for the loss of ThiE (TMP synthase) function. In contrast, bifunctional ThiDN, in which the ThiN domain is fused to an N-terminal ThiD (HMP/HMP phosphate [HMP-P] kinase) domain, was found to be interchangeable for ThiE function and, thus, active in thiamine biosynthesis. A conserved Met residue of an extended ␣-helix near the active-site His of the ThiN domain was found to be important for ThiDN catalytic activity, whereas the corresponding Met residue was absent and the ␣-helix was shorter in ThiR homologs. Thus, we provide new insight into residues that distinguish catalytic from noncatalytic ThiN domains and reveal that thiamine biosynthesis in archaea is regulated by a transcriptional repressor, ThiR, and not by a riboswitch.IMPORTANCE Thiamine pyrophosphate (TPP) is a cofactor needed for the enzymatic activity of many cellular processes, including central metabolism. In archaea, thiamine biosynthesis is an apparent chimera of eukaryote-and bacterium-type pathways that is not well defined at the level of enzymatic steps or regulatory mechanisms. Here we find that ThiN is a versatile domain of transcriptional repressors and catalytic enzymes of thiamine biosynthesis in archaea. Our study provides new insight into residues that distinguish catalytic from noncatalytic ThiN domains and reveals that archaeal thiamine biosynthesis is regulated by a ThiN domain transcriptional repressor, ThiR, and not by a riboswitch.

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“…sRNAs in bacteria and eukarya play essential roles in transcriptional regulation, chromosome replication, RNA processing and modification, mRNA stability and translation, and even protein degradation and translocation [3][4][5]. Recently, it was discovered that archaeal genomes encode for large numbers of sRNAs and that many of them are responsive to environmental stresses [1, 2,[6][7][8][9][10][11][12][13][14].…”

Section: Introductionmentioning

confidence: 99%

The Non-Coding Regulatory RNA Revolution in Archaea

Gelsinger

DiRuggiero

2018

Preprint

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Small non-coding RNAs (sRNAs) are ubiquitously found in the three domains of life playing large-scale roles in gene regulation, transposable element silencing, and defense against foreign elements. While a substantial body of experimental work has been done to uncover function of sRNAs in Bacteria and Eukarya, the functional roles of sRNAs in Archaea are still poorly understood. Recently, high throughput studies using RNA-sequencing revealed that sRNAs are broadly expressed in the Archaea, comprising thousands of transcripts within the transcriptome during non-challenged and stressed conditions. Antisense sRNAs, which overlap a portion of a gene on the opposite strand (cis-acting), are the most abundantly expressed non-coding RNAs and they can be classified based on their binding patterns to mRNAs (3' UTR, 5' UTR, CDSbinding). These antisense sRNAs target many genes and pathways, suggesting extensive roles in gene regulation. Intergenic sRNAs are less abundantly expressed and their targets are difficult to find because of a lack of complete overlap between sRNAs and target mRNAs (trans-acting).While many sRNAs have been validated experimentally, a regulatory role has only been reported for very few of them. Further work is needed to elucidate sRNA-RNA binding mechanisms, the molecular determinants of sRNA-mediated regulation, whether protein components are involved, and how sRNAs integrate with complex regulatory networks.

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Genome-wide identification of transcriptional start sites in the haloarchaeon Haloferax volcanii based on differential RNA-Seq (dRNA-Seq)

Cited by 126 publications

References 93 publications

Internal RNAs overlapping coding sequences can drive the production of alternative proteins in archaea

Internal RNAs overlapping coding sequences can drive the production of alternative proteins in archaea

ThiN as a Versatile Domain of Transcriptional Repressors and Catalytic Enzymes of Thiamine Biosynthesis

The Non-Coding Regulatory RNA Revolution in Archaea

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