Laser controlled singlet oxygen generation in mitochondria to promote mitochondrial DNA replication in vitro

Sci Rep. 2015 Nov 18;5:16925. doi: 10.1038/srep16925.

Abstract

Reports have shown that a certain level of reactive oxygen species (ROS) can promote mitochondrial DNA (mtDNA) replication. However, it is unclear whether it is the mitochondrial ROS that stimulate mtDNA replication and this requires further investigation. Here we employed a photodynamic system to achieve controlled mitochondrial singlet oxygen ((1)O2) generation. HeLa cells incubated with 5-aminolevulinic acid (ALA) were exposed to laser irradiation to induce (1)O2 generation within mitochondria. Increased mtDNA copy number was detected after low doses of 630 nm laser light in ALA-treated cells. The stimulated mtDNA replication was directly linked to mitochondrial (1)O2 generation, as verified using specific ROS scavengers. The stimulated mtDNA replication was regulated by mitochondrial transcription factor A (TFAM) and mtDNA polymerase γ. MtDNA control region modifications were induced by (1)O2 generation in mitochondria. A marked increase in 8-Oxoguanine (8-oxoG) level was detected in ALA-treated cells after irradiation. HeLa cell growth stimulation and G1-S cell cycle transition were also observed after laser irradiation in ALA-treated cells. These cellular responses could be due to a second wave of ROS generation detected in mitochondria. In summary, we describe a controllable method of inducing mtDNA replication in vitro.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Proliferation
  • DNA Polymerase gamma
  • DNA Replication / radiation effects*
  • DNA, Mitochondrial / radiation effects*
  • DNA-Binding Proteins / metabolism
  • DNA-Directed DNA Polymerase / metabolism
  • Gene Dosage
  • Guanine / analogs & derivatives
  • Guanine / metabolism
  • HeLa Cells
  • Humans
  • Lasers*
  • Mitochondria / drug effects
  • Mitochondria / physiology*
  • Mitochondria / radiation effects*
  • Mitochondrial Proteins / metabolism
  • Oxidative Stress / genetics
  • Photochemotherapy
  • Photosensitizing Agents / pharmacology
  • Protoporphyrins / pharmacology
  • Reactive Oxygen Species / metabolism
  • Singlet Oxygen / metabolism*
  • Transcription Factors / metabolism

Substances

  • DNA, Mitochondrial
  • DNA-Binding Proteins
  • Mitochondrial Proteins
  • Photosensitizing Agents
  • Protoporphyrins
  • Reactive Oxygen Species
  • TFAM protein, human
  • Transcription Factors
  • Singlet Oxygen
  • 8-hydroxyguanine
  • Guanine
  • protoporphyrin IX
  • DNA Polymerase gamma
  • DNA-Directed DNA Polymerase