scholarly journals Author response: Complete dissection of transcription elongation reveals slow translocation of RNA polymerase II in a linear ratchet mechanism

2013 ◽  
Author(s):  
Manchuta Dangkulwanich ◽  
Toyotaka Ishibashi ◽  
Shixin Liu ◽  
Maria L Kireeva ◽  
Lucyna Lubkowska ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Elongation ◽  
Author Response ◽  
Ratchet Mechanism

scholarly journals Complete Dissection of Transcription Elongation Reveals Slow Translocation of RNA Polymerase II in a Linear Ratchet Mechanism

2014 ◽  
Vol 106 (2) ◽  
pp. 485a-486a ◽  
Author(s):  
Manchuta Dangkulwanich ◽  
Toyotaka Ishibashi ◽  
Shixin Liu ◽  
Maria L. Kireeva ◽  
Lucyna Lubkowska ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Elongation ◽  
Ratchet Mechanism

scholarly journals Complete dissection of transcription elongation reveals slow translocation of RNA polymerase II in a linear ratchet mechanism

eLife ◽  
2013 ◽  
Vol 2 ◽  
Author(s):  
Manchuta Dangkulwanich ◽  
Toyotaka Ishibashi ◽  
Shixin Liu ◽  
Maria L Kireeva ◽  
Lucyna Lubkowska ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Single Molecule ◽  
Transcription Elongation ◽  
Energy Landscape ◽  
Ratchet Mechanism ◽  
Elongation Cycle ◽  
Nucleotide Addition ◽  
Single Molecule Studies ◽  
Rate Limiting

During transcription elongation, RNA polymerase has been assumed to attain equilibrium between pre- and post-translocated states rapidly relative to the subsequent catalysis. Under this assumption, recent single-molecule studies proposed a branched Brownian ratchet mechanism that necessitates a putative secondary nucleotide binding site on the enzyme. By challenging individual yeast RNA polymerase II with a nucleosomal barrier, we separately measured the forward and reverse translocation rates. Surprisingly, we found that the forward translocation rate is comparable to the catalysis rate. This finding reveals a linear, non-branched ratchet mechanism for the nucleotide addition cycle in which translocation is one of the rate-limiting steps. We further determined all the major on- and off-pathway kinetic parameters in the elongation cycle. The resulting translocation energy landscape shows that the off-pathway states are favored thermodynamically but not kinetically over the on-pathway states, conferring the enzyme its propensity to pause and furnishing the physical basis for transcriptional regulation.

Crystal structure of the human transcription elongation factor DSIF hSpt4 subunit in complex with the hSpt5 dimerization interface

2009 ◽  
Vol 425 (2) ◽  
pp. 373-380 ◽  
Author(s):  
Sabine Wenzel ◽  
Berta M. Martins ◽  
Paul Rösch ◽  
Birgitta M. Wöhrl
Keyword(s):  
Crystal Structure ◽  
Escherichia Coli ◽  
Transcription Factor ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Transcription Elongation ◽  
Elongation Factor ◽  
Structural Similarity ◽  
Transcription Elongation Factor

The eukaryotic transcription elongation factor DSIF [DRB (5,6-dichloro-1-β-D-ribofuranosylbenzimidazole) sensitivity-inducing factor] is composed of two subunits, hSpt4 and hSpt5, which are homologous to the yeast factors Spt4 and Spt5. DSIF is involved in regulating the processivity of RNA polymerase II and plays an essential role in transcriptional activation of eukaryotes. At several eukaryotic promoters, DSIF, together with NELF (negative elongation factor), leads to promoter-proximal pausing of RNA polymerase II. In the present paper we describe the crystal structure of hSpt4 in complex with the dimerization region of hSpt5 (amino acids 176–273) at a resolution of 1.55 Å (1 Å=0.1 nm). The heterodimer shows high structural similarity to its homologue from Saccharomyces cerevisiae. Furthermore, hSpt5-NGN is structurally similar to the NTD (N-terminal domain) of the bacterial transcription factor NusG. A homologue for hSpt4 has not yet been found in bacteria. However, the archaeal transcription factor RpoE” appears to be distantly related. Although a comparison of the NusG-NTD of Escherichia coli with hSpt5 revealed a similarity of the three-dimensional structures, interaction of E. coli NusG-NTD with hSpt4 could not be observed by NMR titration experiments. A conserved glutamate residue, which was shown to be crucial for dimerization in yeast, is also involved in the human heterodimer, but is substituted for a glutamine residue in Escherichia coli NusG. However, exchanging the glutamine for glutamate proved not to be sufficient to induce hSpt4 binding.

scholarly journals BRCA2 Regulates Transcription Elongation by RNA Polymerase II to Prevent R-Loop Accumulation

Cell Reports ◽  
2018 ◽  
Vol 22 (4) ◽  
pp. 1031-1039 ◽  
Author(s):  
Mahmud K.K. Shivji ◽  
Xavier Renaudin ◽  
Çiğdem H. Williams ◽  
Ashok R. Venkitaraman
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Elongation

scholarly journals The Rpb6 Subunit of Fission Yeast RNA Polymerase II Is a Contact Target of the Transcription Elongation Factor TFIIS

2000 ◽  
Vol 20 (4) ◽  
pp. 1263-1270 ◽  
Author(s):  
Akira Ishiguro ◽  
Yasuhisa Nogi ◽  
Koji Hisatake ◽  
Masami Muramatsu ◽  
Akira Ishihama
Keyword(s):  
Rna Polymerase ◽  
Fission Yeast ◽  
Rna Polymerase Ii ◽  
Transcription Elongation ◽  
Elongation Factor ◽  
Direct Interaction ◽  
Temperature Sensitive ◽  
Single Mutation ◽  
Transcription Elongation Factor ◽  
Essential Region

ABSTRACT The Rpb6 subunit of RNA polymerase II is one of the five subunits common to three forms of eukaryotic RNA polymerase. Deletion and truncation analyses of the rpb6 gene in the fission yeastSchizosaccharomyces pombe indicated that Rpb6, consisting of 142 amino acid residues, is an essential protein for cell viability, and the essential region is located in the C-terminal half between residues 61 and 139. After random mutagenesis, a total of 14 temperature-sensitive mutants were isolated, each carrying a single (or double in three cases and triple in one) mutation. Four mutants each carrying a single mutation in the essential region were sensitive to 6-azauracil (6AU), which inhibits transcription elongation by depleting the intracellular pool of GTP and UTP. Both 6AU sensitivity and temperature-sensitive phenotypes of these rpb6 mutants were suppressed by overexpression of TFIIS, a transcription elongation factor. In agreement with the genetic studies, the mutant RNA polymerases containing the mutant Rpb6 subunits showed reduced affinity for TFIIS, as measured by a pull-down assay of TFIIS-RNA polymerase II complexes using a fusion form of TFIIS with glutathioneS-transferase. Moreover, the direct interaction between TFIIS and RNA polymerase II was competed by the addition of Rpb6. Taken together, the results lead us to propose that Rpb6 plays a role in the interaction between RNA polymerase II and the transcription elongation factor TFIIS.

scholarly journals A Role for SSU72 in Balancing RNA Polymerase II Transcription Elongation and Termination

2002 ◽  
Vol 10 (5) ◽  
pp. 1139-1150 ◽  
Author(s):  
Bernhard Dichtl ◽  
Diana Blank ◽  
Martin Ohnacker ◽  
Arno Friedlein ◽  
Daniel Roeder ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Elongation ◽  
Rna Polymerase Ii Transcription

scholarly journals Sub1 associates with Spt5 and influences RNA polymerase II transcription elongation rate

2012 ◽  
Vol 23 (21) ◽  
pp. 4297-4312 ◽  
Author(s):  
Alicia García ◽  
Alejandro Collin ◽  
Olga Calvo
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Elongation ◽  
Elongation Factor ◽  
Molecular Data ◽  
Elongation Rate ◽  
Dependent Manner ◽  
Gene Encoding ◽  
Mrna Metabolism ◽  
Carboxy Terminal

The transcriptional coactivator Sub1 has been implicated in several steps of mRNA metabolism in yeast, such as the activation of transcription, termination, and 3′-end formation. In addition, Sub1 globally regulates RNA polymerase II phosphorylation, and most recently it has been shown that it is a functional component of the preinitiation complex. Here we present evidence that Sub1 plays a significant role in transcription elongation by RNA polymerase II (RNAPII). We show that SUB1 genetically interacts with the gene encoding the elongation factor Spt5, that Sub1 influences Spt5 phosphorylation of the carboxy-terminal domain of RNAPII largest subunit by the kinase Bur1, and that both Sub1 and Spt5 copurify in the same complex, likely during early transcription elongation. Indeed, our data indicate that Sub1 influences Spt5–Rpb1 interaction. In addition, biochemical and molecular data show that Sub1 influences transcription elongation of constitutive and inducible genes and associates with coding regions in a transcription-dependent manner. Taken together, our results indicate that Sub1 associates with Spt5 and influences Spt5–Rpb1 complex levels and consequently transcription elongation rate.

Sign in / Sign up

Export Citation Format

Share Document