Molecular basis of transcription initiation in Archaea

Carlo, Sacha De; Lin, Shih-Chieh; Taatjes, Dylan J.; Hoenger, Andreas

doi:10.4161/trns.1.2.13189

Cited by 14 publications

(9 citation statements)

References 49 publications

(70 reference statements)

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“…The cryo-EM structure of the P. furiosus RNAP has been reported by two different groups (24,25), one of which used an RNAP sample provided by us (24). In both cryo-EM maps, the P. furiosus RNAP resembles a "crab claw" with a protruding "stalk," similar to the crystal structure of Sulfolobus sulfataricus RNAP (Fig.…”

supporting

confidence: 56%

RNA polymerase and transcription elongation factor Spt4/5 complex structure

Klein

Bose

Baker

et al. 2010

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

130

145

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Spt4/5 in archaea and eukaryote and its bacterial homolog NusG is the only elongation factor conserved in all three domains of life and plays many key roles in cotranscriptional regulation and in recruiting other factors to the elongating RNA polymerase. Here, we present the crystal structure of Spt4/5 as well as the structure of RNA polymerase-Spt4/5 complex using cryoelectron microscopy reconstruction and single particle analysis. The Spt4/5 binds in the middle of RNA polymerase claw and encloses the DNA, reminiscent of the DNA polymerase clamp and ring helicases. The transcription elongation complex model reveals that the Spt4/5 is an upstream DNA holder and contacts the nontemplate DNA in the transcription bubble. These structures reveal that the cellular RNA polymerases also use a strategy of encircling DNA to enhance its processivity as commonly observed for many nucleic acid processing enzymes including DNA polymerases and helicases.cryo-EM | Spt4/5-DSIF-NusG | X-ray crystallography T ranscription by RNA polymerase (RNAP) plays a central role in gene expression, and this process is highly regulated at many steps, including promoter recognition, transcription activation, elongation, and termination. During transcription, several transcription elongation factors communicate with RNAP and regulate the velocity of RNA synthesis, transcriptional pausing, and termination (1). Recent genome-wide analyses of transcription elongation revealed the importance of transcription elongation in the regulation of gene expression (2-4). For example, a large fraction of transcribing eukaryotic RNAP II (Pol II) pauses near promoters to facilitate rapid changes in gene expression during cell development.Bacterial NusG is perhaps the best characterized transcription elongation factor in vivo and in vitro. Diverse functions of NusG have been reported, e.g., Escherichia coli NusG reduces RNAP pausing and intrinsic termination (5, 6), whereas Bacillus subtilis and Thermus thermophilus NusG enhance pausing (7,8). NusG consists of the NusG amino-terminal (NGN) domain and Kyprides-Onzonis-Woese (KOW) motif at the C-terminal domain (hence also called KOW domain), and these two domains fold independently and are connected by a flexible linker of 13 amino acids (9, 10) ( Fig. 1B and Fig. S1C). The NGN domain has been assigned the function for regulating transcription elongation. The KOW domain, on the other hand, plays important roles in interacting with other proteins. For example, it contacts the elongation factor NusE, which is also the ribosomal S10 subunit. This interaction is essential for forming rRNA and λ gene antitermination complexes, as well as for the coupling of transcription and translation. Furthermore, the KOW contacts Rho for transcription termination (9, 11).Eukaryotic Spt5 is the functional and structural counterpart of bacterial NusG. In addition to the NGN and KOW domains, eukaryotic Spt5 contains additional motifs including the N-terminal acidic region, four to five additional KOW motifs, and C-terminal repeats t...

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supporting

confidence: 56%

RNA polymerase and transcription elongation factor Spt4/5 complex structure

Klein

Bose

Baker

et al. 2010

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

130

145

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“…TFIIB also plays roles at several key steps during the transcription cycle that include transcription start site selection (TSS), promoter opening, abortive initiation, promoter clearance, and termination 20,23-30 . TBP is also universally required for transcription initiation by archaeal Pol and each of the three major eukaryotic Pols 2,3,14,31-33 . TBP binds specifically to TATA or TATA-like DNA sequences upstream of the transcription start site of promoter DNA, and induces a sharp 90° bend in DNA 7,3334 .…”

Section: Introductionmentioning

confidence: 99%

Emergence and expansion of TFIIB-like factors in the plant kingdom

Knutson

2013

Gene

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Many gene families in higher plants have expanded in number, giving rise to diverse protein paralogs with specialized biochemical functions. For instance, plant general transcription factors such as TFIIB have expanded in number and in some cases perform specialized transcriptional functions in the plant cell. To date, no comprehensive genome-wide identification of the TFIIB gene family has been conducted in the plant kingdom. To determine the extent of TFIIB expansion in plants, I used the remote homology program HHPred to search for TFIIB homologs in the plant kingdom and performed a comprehensive analysis of eukaryotic TFIIB gene families. I discovered that higher plants encode more than 10 different TFIIB-like proteins. In particular, Arabidopsis thaliana encodes 14 different TFIIB-like proteins and predicted domain architectures of the newly identified TFIIB-like proteins revealed that they have unique modular domain structures that are divergent in sequence and size. Phylogenetic analysis of selected eukaryotic organisms showed that most life forms encode three major TFIIB subfamilies that include TFIIB, Brf, Rrn7/TAF1B/MEE12 subfamilies, while all plants and some algae species encode one or two additional TFIIB-related protein subfamilies. A subset of A. thaliana GTFs have also expanded in number, indicating that GTF diversification and expansion is a general phenomenon in higher plants. Together, these findings were used to generate a model for the evolutionary history of TFIIB-like proteins in eukaryotes.

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“…Weakly binding RyR ligands generally do not form sufficiently stable complexes with RyR in vitro for study by single particle cryo-EM. Glutaraldehyde has been used to stabilize large fragile cytoplasmic complexes for single particle cryo-EM (Carlo et al, 2010; Lee et al, 2003; Richter et al, 2010; Southworth and Agar, 2011; Yu et al, 2008), and could similarly be used to stabilize RyR or components of ECC. However this approach has not been tried with RyR before.…”

Section: Introductionmentioning

confidence: 99%

Structure of glutaraldehyde cross-linked ryanodine receptor

Strauss

Wagenknecht

2013

Journal of Structural Biology

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The ryanodine receptor (RyR) family of calcium release channels plays a vital role in excitation-contraction coupling (ECC). Along with the dihydropyridine receptor (DHPR), calsequestrin, and several other smaller regulatory and adaptor proteins, RyRs form a large dynamic complex referred to as ECC machinery. Here we describe a simple cross-linking procedure that can be used to stabilize fragile components of the ECC machinery, for the purpose of structural elucidation by single particle cryo-electron microscopy (cryo-EM). As a model system, the complex of the FK506-binding protein (FKBP12) and RyR1 was used to test the cross-linking protocol. Glutaraldehyde fixation led to complete cross-linking of receptor-bound FKBP12 to RyR1, and also to extensive cross-linking of the four subunits comprising RyR to one another without compromising the RyR1 ultrastructure. FKBP12 cross-linked with RyR1 was visualized in 2D averages by single particle cryo-EM. Comparison of control RyR1 and cross-linked RyR1 3D reconstructions revealed minor conformational changes at the transmembrane assembly and at the cytoplasmic region. Intersubunit cross-linking enhanced [3H]ryanodine binding to RyR1. Based on our findings we propose that intersubunit cross-linking of RyR1 by glutaraldehyde induced RyR1 to adopt an open like conformation.

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Molecular basis of transcription initiation in Archaea

Cited by 14 publications

References 49 publications

RNA polymerase and transcription elongation factor Spt4/5 complex structure

RNA polymerase and transcription elongation factor Spt4/5 complex structure

Emergence and expansion of TFIIB-like factors in the plant kingdom

Structure of glutaraldehyde cross-linked ryanodine receptor

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