相关实验视频

Updated: Jun 30, 2025

Visualization of UV-induced Replication Intermediates in E. coli using Two-dimensional Agarose-gel Analysis

Visualization of UV-induced Replication Intermediates in E. coli using Two-dimensional Agarose-gel Analysis

Published on: December 21, 2010

通过使用单分子追踪来比较Mfd和UvrD依赖的转录合DNA修复在活体大肠杆菌中的模型

Elżbieta Kaja¹, Donata Vijande², Justyna Kowalczyk²

¹Molecular Biophysics Division, Faculty of Physics, Adam Mickiewicz University, Uniwersytetu Poznańskiego 2, Poznan 61-614, Poland; Chair and Department of Medical Chemistry and Laboratory Medicine, Poznan University of Medical Sciences, Rokietnicka 8, 60-806 Poznan, Poland.

|March 21, 2024

概括

这项研究研究了E. coli的转录合DNA修复 (TCR). 活细胞成像显示,虽然UvrA可以与RNA聚合酶 (RNAP) 相结合,但UvrD-RNAP-UvrA复合体不是核酸切割修复 (NER) 的主要DNA损伤传感器.

科学领域:

分子生物学分子生物学
微生物学微生物学
遗传学是一个遗传学.
DNA 修复机制的修复机制

背景情况:

转录合DNA修复 (TCR) 在RNA聚合酶 (RNAP) 遇到DNA损伤时启动核酸切除修复 (NER).
对于TCR存在两种模型:Mfd介导的RNAP位移或UvrD酶介导的回溯.
一个最近的假设提出UvrD和UvrA与RNAP作为损伤前传感器综合体结合.

研究的目的:

实验测试TCR在活体大肠杆菌中存在的两个主要模型.
通过使用活细胞成像来研究Mfd,UvrD,UvrA和UvrB在TCR中的作用.
为了确定UvrD-RNAP-UvrA复合体是否作为NER的主要传感器.

主要方法:

使用超分辨率显微镜 (光激活定位显微镜 - PALM) 与单颗粒追踪相结合.
测量了Mfd,UvrD,UvrA和UvrB到DNA的流动性和招聘动态.
使用紫外线 (UV) 辐射诱导的DNA损伤和用利芬素抑制的转录.

主要成果:

UvrA显示了RNAP复杂化的潜力,但UvrD没有,因为没有DNA损伤.
Mfd对DNA损伤后的招募独立于UvrA,支持其在TCR中的上游作用.
UvrD的招募依赖于UvrA,UvrD,UvrA和UvrB的招募在转录抑制后仍然存在,这表明NER功能独立于转录RNAP.

结论:

拟议的UvrD-RNAP-UvrA复合体不是NER的主要DNA损伤传感器,尽管它有可能形成.
在TCR路径中,Mfd在UvrA的上游作用.
UvrD和UvrA在NER中独立于转录RNAP的功能,挑战了回溯模型.

关键词:

在MfdMfd中使用.核酸切除修复核酸切除修复转录合修复的修复紫外线引起的DNA损伤在UvrDD中使用.

更多相关视频

Visualizing Protein-DNA Interactions in Live Bacterial Cells Using Photoactivated Single-molecule Tracking

Visualizing Protein-DNA Interactions in Live Bacterial Cells Using Photoactivated Single-molecule Tracking

Published on: March 10, 2014

Characterizing DNA Repair Processes at Transient and Long-lasting Double-strand DNA Breaks by Immunofluorescence Microscopy

Characterizing DNA Repair Processes at Transient and Long-lasting Double-strand DNA Breaks by Immunofluorescence Microscopy

Published on: June 8, 2018

相关概念视频

Mismatch Repair

Mismatch Repair

Organisms are capable of detecting and fixing nucleotide mismatches that occur during DNA replication. This sophisticated process requires identifying the new strand and replacing the erroneous bases with correct nucleotides. Mismatch repair is coordinated by many proteins in both prokaryotes and eukaryotes.
The Mutator Protein Family Plays a Key Role in DNA Mismatch Repair
The human genome has more than 3 billion base pairs of DNA per cell. Prior to cell division, that vast amount of genetic...

Nucleotide Excision Repair

Nucleotide Excision Repair

DNA Distortion and Damage
Cells are regularly exposed to mutagens—factors in the environment that can damage DNA and generate mutations. UV radiation is one of the most common mutagens and is estimated to introduce a significant number of changes in DNA. These include bends or kinks in the structure, which can block DNA replication or transcription. If these errors are not fixed, the damage can cause mutations, which in turn can result in cancer or disease depending on which sequences are...

Homologous Recombination

Homologous Recombination

The basic reaction of homologous recombination (HR) involves two chromatids that contain DNA sequences sharing a significant stretch of identity. One of these sequences uses a strand from another as a template to synthesize DNA in an enzyme-catalyzed reaction. The final product is a novel amalgamation of the two substrates. To ensure an accurate recombination of sequences, HR is restricted to the S and G2 phases of the cell cycle. At these stages, the DNA has been replicated already and the...