DNA topoisomerase I inhibition by camptothecin induces escape of RNA polymerase II from promoter-proximal pause site, antisense transcription and histone acetylation at the human HIF-1α gene locus

A34.5, a phosphoprotein copurifying with RNA polymerase I (Pol I), lacks homology to any component of the Pol II or Pol III transcription complexes. Cells devoid of A34.5 hardly affect growth and rRNA synthesis and generate a catalytically active but structurally modified enzyme also lacking subunit A49 upon in vitro purification. Other Pol I-specific subunits (A49, A14, and A12.2) are nonessential for growth at 30 degrees C but are essential (A49 and A12.2) or helpful (A14) at 25 or 37 degrees C. Triple mutants without A34.5, A49, and A12.2 are viable, but inactivating any of these subunits together with A14 is lethal. Lethality is rescued by expressing pre-rRNA from a Pol II-specific promoter, demonstrating that these subunits are collectively essential but individually dispensable for rRNA synthesis. A14 and A34.5 single deletions affect the subunit composition of the purified enzyme in pleiotropic but nonoverlapping ways which, if accumulated in the double mutants, provide a structural explanation for their strict synthetic lethality. A34.5 (but not A14) becomes quasi-essential in strains lacking DNA topoisomerase I, suggesting a specific role of this subunit in helping Pol I to overcome the topological constraints imposed on ribosomal DNA by transcription.

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Isolation of mutants of Saccharomyces cerevisiae requiring DNA topoisomerase I.

Genetics ◽

10.1093/genetics/141.2.465 ◽

1995 ◽

Vol 141 (2) ◽

pp. 465-479 ◽

Cited By ~ 4

Author(s):

B U Sadoff ◽

S Heath-Pagliuso ◽

I B Castaño ◽

Y Zhu ◽

F S Kieff ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Rna Polymerase Ii ◽

Topoisomerase I ◽

Growth Defect ◽

Dna Topoisomerase I ◽

Dna Topoisomerase ◽

Acid Protein ◽

Complementation Groups ◽

Topo Ii ◽

Null Mutants

Abstract Despite evidence that DNA topoisomerase I is required to relieve torsional stress during DNA replication and transcription, yeast strains with a top1 null mutation are viable and display no gross defects in DNA or RNA synthesis, possibly because other proteins provide overlapping functions. We isolated mutants whose inviablility or growth defect is relieved when TOP1 is expressed [trf mutants (topoisomerase one-requiring function)]. The TRF genes define at least four complementation groups. TRF3 is allelic to TOP2. TRF1 is allelic to HPR1, previously shown to be homologous to TOP1 over two short regions. TRF4 encodes a novel 584-amino acid protein with homology to the N-terminus of Saccharomyces cerevisiae topo I. Like top1 mutants, trf4 mutants have elevated rDNA recombination and fail to shut off RNA polymerase II transcription in stationary phase. trf4 null mutants are cs for viability, display reduced expression of GAL1 and Cell Cycle Box UAS::LacZ fusions, and are inviable in combination with trfI null mutants, indicating that both proteins may share a common function with DNA topoisomerase I. The existence of multiple TRF complementation groups suggests that not all biological functions of topo I can be carried out by topo II.

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Genome-wide proximity between RNA polymerase and DNA topoisomerase I supports transcription in Streptococcus pneumoniae

PLoS Genetics ◽

10.1371/journal.pgen.1009542 ◽

2021 ◽

Vol 17 (4) ◽

pp. e1009542

Author(s):

María-José Ferrándiz ◽

Pablo Hernández ◽

Adela G. de la Campa

Keyword(s):

Streptococcus Pneumoniae ◽

Rna Polymerase ◽

Topoisomerase I ◽

Type I ◽

Dna Topoisomerase I ◽

Sequence Motifs ◽

Dna Topoisomerase ◽

Genome Wide

Streptococcus pneumoniae is a major cause of disease and death that develops resistance to multiple antibiotics. DNA topoisomerase I (TopoI) is a novel pneumococcal drug target. TopoI is the sole type-I pneumococcal topoisomerase that regulates supercoiling homeostasis in this bacterium. In this study, a direct in vitro interaction between TopoI and RNA polymerase (RNAP) was detected by surface plasmon resonance. To understand the interplay between transcription and supercoiling regulation in vivo, genome-wide association of RNAP and TopoI was studied by ChIP-Seq. RNAP and TopoI were enriched at the promoters of 435 and 356 genes, respectively. Higher levels of expression were consistently measured in those genes whose promoters recruit both RNAP and TopoI, in contrast with those enriched in only one of them. Both enzymes occupied a narrow region close to the ATG codon. In addition, RNAP displayed a regular distribution throughout the coding regions. Likewise, the summits of peaks called with MACS tool, mapped around the ATG codon in both cases. However, RNAP showed a broader distribution towards ATG-downstream positions. Remarkably, inhibition of RNAP with rifampicin prevented the localization of TopoI at promoters and, vice versa, inhibition of TopoI with seconeolitsine prevented the binding of RNAP to promoters. This indicates a functional interplay between RNAP and TopoI. To determine the molecular factors responsible for RNAP and TopoI co-recruitment, we looked for DNA sequence motifs. We identified a motif corresponding to a -10-extended promoter for TopoI and for RNAP. Furthermore, RNAP was preferentially recruited to genes co-directionally oriented with replication, while TopoI was more abundant in head-on genes. TopoI was located in the intergenic regions of divergent genes pairs, near the promoter of the head-on gene of the pair. These results suggest a role for TopoI in the formation/stability of the RNAP-DNA complex at the promoter and during transcript elongation.

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DNA Topoisomerase I and PC4 Can Interact with Human TFIIIC to Promote Both Accurate Termination and Transcription Reinitiation by RNA Polymerase III

Molecular Cell ◽

10.1016/s1097-2765(00)80074-x ◽

1998 ◽

Vol 1 (5) ◽

pp. 749-757 ◽

Cited By ~ 57

Author(s):

Zhengxin Wang ◽

Robert G Roeder

Keyword(s):

Rna Polymerase ◽

Topoisomerase I ◽

Rna Polymerase Iii ◽

Dna Topoisomerase I ◽

Dna Topoisomerase ◽

Polymerase Iii ◽

Transcription Reinitiation

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Why Should DNA Topoisomerase I Have a Scaffold Activity?

Biology ◽

10.3390/biology10030190 ◽

2021 ◽

Vol 10 (3) ◽

pp. 190

Author(s):

Francesca Di Felice ◽

Giorgio Camilloni

Keyword(s):

Catalytic Activity ◽

Rna Polymerase Ii ◽

Topoisomerase I ◽

Ribosomal Genes ◽

Dna Topoisomerase I ◽

Dna Topoisomerase ◽

Macromolecular Complexes ◽

Rna Polymerase Ii Transcription

Since the early 1990s, in vitro studies have demonstrated that DNA topoisomerase I promotes RNA polymerase II transcription, acting as a cofactor, regardless of its catalytic activity. Recent studies, carried in vivo, using yeast as a model system, also demonstrate that DNA topoisomerase I is able to recruit, without the involvement of its catalytic activity, the Sir2p deacetylase on ribosomal genes thus contributes to achieve their silencing. In this review, the DNA topoisomerase I capability, acting as a scaffold protein, as well as its involvement and role in several macromolecular complexes, will be discussed, in light of several observations reported in the literature, pointing out how its role goes far beyond its well-known ability to relax DNA.

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