On the Evolution of the Single-Subunit RNA Polymerases

Cermakian, Nicolas; Ikeda, Tatsuya; Miramóntes, Pedro; Lang, B. Franz; Gray, Michael W.; Cedergren, Robert

doi:10.1007/pl00006271

Cited by 150 publications

(116 citation statements)

References 73 publications

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“…This alignment is centered on the region corresponding to the T7 RNAP promoter recognition loop. The sequences flanking this element are well conserved in this family and include the G, H, and I blocks of conserved elements previously identified in a comprehensive analysis of sequence conservation patterns in the single subunit RNAP family (11). However, within the region corresponding to the T7 promoter recognition loop, conservation breaks down, although all of these RNAPs exhibit a 23-29-residue element of variable sequence between the conserved G/H and I blocks.…”

Section: All Single Subunit Rnaps Exhibit a Variable 26 ϯ 3-amino Acimentioning

confidence: 85%

“…The Ϫ3 to Ϫ11 region of its promoter is recognized primarily by the promoter specificity loop, an extended ␤-hairpin that emerges from the polymerase C-terminal domain, and the Ϫ12 to Ϫ17 region of the promoter is recognized by elements in the N-terminal domain of the polymerase (8 -10). Mitochondrial and chloroplast RNAPs are unlikely to have a structural element that is homologous or functionally analogous to the T7 RNAP N-terminal domain, since the N-terminal domains of these enzymes are highly divergent (11), and the promoter sequences recognized directly by the mitochondrial and chloroplast RNAPs do not extend upstream of Ϫ9. However, it has been suggested that the YMt RNAP has a promoter recognition loop like that seen in T7 RNAP (12), and the recently described structure of N4 RNAP exhibits a promoter loop similar to that of T7 RNAP (13), although this RNAP uses a hairpin promoter (Fig.…”

Section: S Cerevisiae Ymtmentioning

confidence: 99%

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A Promoter Recognition Mechanism Common to Yeast Mitochondrial and Phage T7 RNA Polymerases

Nayak

Guo

Sousa

2009

Journal of Biological Chemistry

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Yeast mitochondrial (YMt) and phage T7 RNA polymerases (RNAPs) are two divergent representatives of a large family of single subunit RNAPs that are also found in the mitochondria and chloroplasts of higher eukaryotes, mammalian nuclei, and many other bacteriophage. YMt and phage T7 promoters differ greatly in sequence and length, and the YMt RNAP uses an accessory factor for initiation, whereas T7 RNAP does not. We obtain evidence here that, despite these apparent differences, both the YMt and T7 RNAPs utilize a similar promoter recognition loop to bind their respective promoters. Mutations in this element in YMt RNAP specifically disrupt mitochondrial promoter utilization, and experiments with site-specifically tethered chemical nucleases indicate that this element binds the mitochondrial promoter almost identically to how the promoter recognition loop from the phage RNAP binds its promoter. Sequence comparisons reveal that the other members of the single subunit RNAP family display loops of variable sequence and size at a position corresponding to the YMt and T7 RNAP promoter recognition loops. We speculate that these elements may be involved in promoter recognition in most or all of these enzymes and that this element's structure allows it to accommodate significant sequence and length variation to provide a mechanism for rapid evolution of new promoter specificities in this RNAP family. S. cerevisiae YMt2 and phage T7 RNAPs are both members of the single subunit RNAP family, but their promoter sequences are very different. The T7 promoter is a 21-nucleotide (nt) duplex that includes 4 nt downstream of the transcription start site and 17 nt upstream of ϩ1 (1). The promoters of the closely related T3 phage or the more distantly related Sp6 phage(2) are identical in size to the T7 promoter but differ in sequence. In contrast, the YMt promoter is only 8 nt in length and is therefore similar in size to chloroplast promoters or the mitochondrial promoters of plants and other fungi, which are typically 8 -10 nt in length although highly disparate in sequence (3, 4) (Fig. 1).The mechanisms of promoter recognition by the YMt and T7 RNAP also appear to be distinct. T7 RNAP, like many of the phage polymerases, binds, melts, and initiates transcription as single subunit enzyme without any accessory transcription factor (5). In contrast, the YMt RNAP requires a specific accessory factor (Mtf1 (yeast mitochondrial transcription factor)) to initiate transcription from its promoter (6). This seeming disparity may, however, obscure underlying mechanistic similarity. For example, it was originally suggested (6) that Mtf1 might be homologous and analogous to prokaryotic RNAP -factor, which would mean that Mtf1 contributes directly to promoter recognition, a mechanism distinct from that seen with T7 RNAP, where the polymerase contains all of the promoter recognition elements. However, subsequent work showed that YMt RNAP can initiate transcription without Mtf1 if the promoter is supercoiled or contains a heterduplex ("bubble") ...

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Section: All Single Subunit Rnaps Exhibit a Variable 26 ϯ 3-amino Acimentioning

confidence: 85%

Section: S Cerevisiae Ymtmentioning

confidence: 99%

A Promoter Recognition Mechanism Common to Yeast Mitochondrial and Phage T7 RNA Polymerases

Nayak

Guo

Sousa

2009

Journal of Biological Chemistry

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“…However, vRNAP contains four short motifs (TxxGR, A, B, and C) characteristic of the T7-like RNAP family, which includes phage-, mitochondrial-, some chloroplast nuclear-, and linear plasmid-encoded enzymes (15,42). We have defined a stable and transcriptionally active 1,106-aa long domain (mini-vRNAP) located at the center of the vRNAP polypeptide, which possesses the same initiation, elongation, termination, and product displacement properties as full-length vRNAP (15).…”

Section: Ecossb Activates Vrnap Transcription At Limiting Ssdna Templatementioning

confidence: 99%

Escherichia coli single-stranded DNA-binding protein mediates template recycling during transcription by bacteriophage N4 virion RNA polymerase

Davydova

Rothman-Denes

2003

Proc. Natl. Acad. Sci. U.S.A.

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Coliphage N4 virion RNA polymerase (vRNAP), the most distantly related member of the T7-like family of RNA polymerases, is responsible for transcription of the early genes of the linear double-stranded DNA phage genome. Escherichia coli singlestranded DNA-binding protein (EcoSSB) is required for N4 early transcription in vivo, as well as for in vitro transcription on supercoiled DNA templates containing vRNAP promoters. In contrast to other DNA-dependent RNA polymerases, vRNAP initiates transcription on single-stranded, promoter-containing templates with in vivo specificity; however, the RNA product is not displaced, thus limiting template usage to one round. We show that EcoSSB activates vRNAP transcription at limiting single-stranded template concentrations through template recycling. EcoSSB binds to the template and to the nascent transcript and prevents the formation of a transcriptionally inert RNA:DNA hybrid. Using C-terminally truncated EcoSSB mutant proteins, human mitochondrial SSB (Hsmt SSB), phage P1 SSB, and F episome-encoded SSB, as well as a Hsmt-EcoSSB chimera, we have mapped a determinant of template recycling to the C-terminal amino acids of EcoSSB. T7 RNAP contains an amino-terminal domain responsible for binding the RNA product as it exits from the enzyme. No sequence similarity to this domain exists in vRNAP. Hereby, we propose a unique role for EcoSSB: It functionally substitutes in N4 vRNAP for the N-terminal domain of T7 RNAP responsible for RNA binding.B acteriophage N4 virion RNA polymerase (vRNAP), responsible for transcription of the phage early genes, is injected into the host cell with the phage genome upon infection (1). vRNAP is inactive on linear double-stranded templates but transcribes denatured genomic N4 DNA or promoter-containing single-stranded DNA (ssDNA) with in vivo specificity (2, 3). vRNAP promoters contain a 5-to 7-bp stem, 3-nt loop hairpin, and conserved sequences (3, 4).In vivo, N4 early transcription requires Escherichia coli DNA gyrase and E. coli ssDNA-binding protein (EcoSSB), which provide an ''active'' promoter structure (5). Other SSBs (T7 gp 2.5, T4 gp 32, and N4 SSB) fail to stimulate vRNAP transcription in vitro because they destabilize the promoter hairpin (6). vRNAP transcripts generated from ssDNA templates are retained as RNA⅐DNA hybrids, suggesting that vRNAP cannot displace the RNA product (2). We show that EcoSSB activates vRNAP transcription on ssDNA templates through displacement of the RNA product and, therefore, through template recycling.The 177-aa EcoSSB polypeptide consists of a 115-aa Nterminal domain responsible for DNA binding and an Ϸ50-aa Pro-and Gly-rich sequence followed by a 10-aa acidic tail, highly conserved in eubacterial SSBs (7,8). Human mitochondrial (Hsmt) SSB is similar in sequence and structure to the Nterminal domain of EcoSSB but lacks the conserved C-terminal sequence (9-11). We show that Hsmt SSB did not activate vRNAP transcription although it did not disrupt the promoter hairpin. We determined that the 10 C-terminal ami...

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“…One intriguing example is the replacement of the original, multisubunit bacteria-like RNA polymerase (inherited from the proto-mitochondrial ancestor) by a bacteriophage T3/T7-like RNA polymerase. This secondarily acquired RNA polymerase directs mitochondrial transcription in all contemporary eukaryotes, except in the minimally derived jakobid flagellates (24).…”

Section: Intracellular Gene Migration From Mitochondria To the Nucleusmentioning

confidence: 99%

Parallels in Genome Evolution in Mitochondria and Bacterial Symbionts

Burger

Lang

2003

IUBMB Life

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SummaryMitochondria, the energy-producing organelles of the eukaryotic cell, originate from an endosymbiotic a-proteobacterium. These organelles are believed to have arisen only once in evolutionary history, but despite their common ancestry, mitochondrial DNAs vary extensively throughout eukaryotes in genome architecture and gene content. New insights into early mitochondrial genome evolution come from the investigation of primitive mitochondriate eukaryotes, as well as the comparison between mitochondria and intracellular bacterial symbionts. IUBMB Life, 55: 205-212, 2003

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On the Evolution of the Single-Subunit RNA Polymerases

Cited by 150 publications

References 73 publications

A Promoter Recognition Mechanism Common to Yeast Mitochondrial and Phage T7 RNA Polymerases

A Promoter Recognition Mechanism Common to Yeast Mitochondrial and Phage T7 RNA Polymerases

Escherichia coli single-stranded DNA-binding protein mediates template recycling during transcription by bacteriophage N4 virion RNA polymerase

Parallels in Genome Evolution in Mitochondria and Bacterial Symbionts

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