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Co-immunoprecipitation Assay Using Endogenous Nuclear Proteins from Cells Cultured Under Hypoxic Conditions
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Published on: August 2, 2018

Oxidative DNA modifications in hypoxic signaling.

Mark N Gillespie1, Viktor Pastukh, Mykhaylo V Ruchko

  • 1Department of Pharmacology, College of Medicine, University of South Alabama, Mobile, AL 36688-0002, USA. mgillesp@jaguar1.usouthal.edu

Annals of the New York Academy of Sciences
|October 23, 2009
PubMed
Summary

Hypoxia triggers DNA base modifications via reactive oxygen species (ROS), enhancing gene expression. DNA repair pathways then resolve these modifications, crucial for hypoxia-induced transcription.

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Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • Hypoxia is a critical biological stimulus.
  • Reactive oxygen species (ROS) act as second messengers in cellular signaling.
  • Hypoxia-inducible genes, like vascular endothelial growth factor (VEGF), are key targets in cellular responses.

Purpose of the Study:

  • To investigate the role of ROS-mediated DNA base modifications in hypoxia-induced gene transcription.
  • To explore the involvement of DNA repair pathways in regulating hypoxia-responsive gene expression.
  • To elucidate the mechanism by which ROS influence transcription factor binding and gene activation.

Main Methods:

  • Analysis of oxidative base modifications within hypoxic response elements (HREs) of VEGF and other hypoxia-inducible genes.
  • Localization of modifications to transcriptionally active mononucleosomes.
  • Investigating the role of Ref-1/Ape1 in transcription complex assembly and DNA repair.
  • Utilizing oligonucleotide-based models to mimic hypoxia effects by introducing abasic sites.

Main Results:

  • Hypoxia-generated ROS directly target specific DNA bases within HREs.
  • Oxidative modifications correlate with increased mRNA accumulation and are found in active chromatin.
  • The base excision DNA repair pathway, involving Ref-1/Ape1, is implicated in removing these modifications.
  • Simulating abasic sites in HREs alters transcription factor binding and enhances reporter gene expression.

Conclusions:

  • Controlled DNA damage by ROS and subsequent repair via the base excision pathway are essential for hypoxia-induced transcriptional activation.
  • ROS act as signaling molecules that initiate a cascade involving DNA modification and repair to regulate gene expression.
  • This mechanism highlights a novel interplay between oxidative stress, DNA integrity, and transcriptional control in response to hypoxia.