ROS-generating oxidases Nox1 and Nox4 contribute to oncogenic Ras-induced premature senescence
Ryo Kodama1, Masayoshi Kato, Shuichi Furuta
1Department of Molecular Biology and Biochemistry, Shinshu University Graduate School of Medicine, Matsumoto, Nagano, 390-8621, Japan.
Abstract:
Activated oncogenes induce premature cellular senescence, a permanent state of proliferative arrest in primary rodent and human fibroblasts. Recent studies suggest that generation of reactive oxygen species (ROS) is involved in oncogenic Ras-induced premature senescence. However, the signaling mechanism controlling this oxidant-mediated irreversible growth arrest is not fully understood. Here, we show that through the Ras/MEK pathway, Ras oncogene up-regulated the expression of superoxide-generating oxidases, Nox1 in rat REF52 cells and Nox4 in primary human lung TIG-3 cells, leading to an increase in intracellular level of ROS. Ablation of Nox1 and Nox4 by small interfering RNAs (siRNAs) blocked the RasV12 senescent phenotype including β-galactosidase activity, growth arrest and accumulation of tumor suppressors such as p53 and p16Ink4a. This suggests that Nox-generated ROS transduce senescence signals by activating the p53 and p16Ink4a pathway. Furthermore, Nox1 and Nox4 siRNAs inhibited both Ras-induced DNA damage response and p38MAPK activation, whereas overexpression of Nox1 and Nox4 alone was able to induce senescence. The involvement of Nox1 in Ras-induced senescence was also confirmed with embryonic fibroblasts derived from Nox1 knockout mice. Together, these findings suggest that Nox1- and Nox4-generated ROS play an important role in Ras-induced premature senescence, which may involve DNA damage response and p38MAPK signaling pathways.
Insights
Oncogene-activated cells undergo premature senescence, a process driven by reactive oxygen species (ROS) generated by Nox1 and Nox4 enzymes. Inhibiting these enzymes halts senescence, implicating ROS in Ras-induced cell cycle arrest.
Area of Science:
- Cell Biology
- Molecular Biology
- Oncology
Background:
- Oncogene activation triggers premature cellular senescence, a permanent growth arrest.
- Reactive oxygen species (ROS) are implicated in oncogenic Ras-induced senescence, but the underlying mechanisms remain unclear.
Purpose of the Study:
- To elucidate the signaling pathways connecting Ras oncogenes to ROS production and subsequent senescence.
- To investigate the role of superoxide-generating oxidases (Nox1 and Nox4) in Ras-induced premature senescence.
Main Methods:
- Utilized Ras/MEK pathway activation in rat REF52 and human TIG-3 cells.
- Employed small interfering RNAs (siRNAs) to ablate Nox1 and Nox4 expression.
- Assessed senescence markers (β-galactosidase activity, growth arrest, p53, p16Ink4a accumulation).
- Investigated DNA damage response and p38MAPK activation.
- Confirmed findings using Nox1 knockout mouse embryonic fibroblasts.
Main Results:
- Ras oncogene up-regulated Nox1 and Nox4 expression, increasing intracellular ROS levels.
- Ablation of Nox1 and Nox4 using siRNAs prevented RasV12-induced senescence phenotypes.
- Nox1/Nox4 knockdown inhibited Ras-induced DNA damage response and p38MAPK activation.
- Overexpression of Nox1 or Nox4 alone induced senescence.
- Nox1 deficiency in mice confirmed its role in Ras-induced senescence.
Conclusions:
- Nox1 and Nox4-generated ROS are crucial mediators of Ras-induced premature senescence.
- These ROS likely signal through DNA damage response and p38MAPK pathways to enforce growth arrest.
- Targeting Nox enzymes may offer therapeutic strategies for cancers driven by oncogenic Ras.
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