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Published on: August 4, 2019
Activation of p53 in anoxic freshwater crayfish, Faxonius virilis
Aakriti Gupta1, Sarah A Breedon1, Kenneth B Storey1
1Department of Biology, Carleton University, Ottawa, ON, Canada, K1S 5B6.
Abstract:
Tumor suppressing transcription factor p53 regulates multiple pathways including DNA repair, cell survival, apoptosis and autophagy. Here, we studied the stress-induced activation of p53 in anoxic crayfish (Faxonius virilis). Relative levels of target proteins and mRNAs involved in the DNA damage response were measured in normoxic control and anoxic hepatopancreas and tail muscle. Phosphorylation levels of p53 were assessed using immunoblotting at sites known to be phosphorylated (serine 15 and 37) in response to DNA damage or reduced oxygen signaling. The capacity for DNA binding by phosphorylated p53 (p-p53) was also measured, followed by transcript analysis of a potentially pro-apoptotic downstream target, the etoposide induced (ei24) gene. Following this, both inhibitor (MDM2) and activator (p19-ARF) protein levels in response to low-oxygen stress were studied. The results showed an increase in p-p53 levels during anoxia in both hepatopancreas and tail muscle. Increased transcript levels of ei24 support the activation of p53 under anoxic stress. Cytoplasmic accumulation of Ser15 phosphorylated p53 was observed during anoxia when proteins from cytoplasmic and nuclear fractions were measured. Increased cytoplasmic concentration is known to initiate an apoptotic response, which can be assumed as a preparatory step to prevent autophagy. The results suggest that p53 might play a protective role in crayfish defense against low-oxygen stress. Understanding how anoxia-tolerant organisms are able to protect themselves against DNA damage could provide important clues towards survival under metabolic rate depression and preparation for recovery to minimize damage.
Insights
The tumor suppressor p53 is activated by low oxygen in crayfish, increasing DNA repair and potentially preventing cell death. This suggests p53 plays a protective role in anoxia-tolerant species.
Area of Science:
- * Molecular Biology
- * Stress Physiology
- * Eukaryotic Gene Regulation
Background:
- * The tumor suppressor p53 is a key regulator of cellular responses to stress, including DNA repair, apoptosis, and autophagy.
- * Anoxia (low oxygen) is a significant environmental stressor that can lead to cellular damage, particularly DNA damage.
- * Understanding how organisms tolerate anoxia is crucial for insights into metabolic depression and recovery.
Purpose of the Study:
- * To investigate the stress-induced activation of transcription factor p53 in anoxic crayfish (Faxonius virilis).
- * To determine the role of p53 in the DNA damage response and cellular protection under low-oxygen conditions.
- * To explore the potential protective mechanisms employed by anoxia-tolerant organisms.
Main Methods:
- * Measurement of target protein and mRNA levels in normoxic and anoxic crayfish tissues (hepatopancreas and tail muscle).
- * Assessment of p53 phosphorylation at key sites (serine 15 and 37) using immunoblotting.
- * Analysis of phosphorylated p53 DNA-binding capacity and transcript levels of the downstream target gene, ei24.
- * Quantification of inhibitor (MDM2) and activator (p19-ARF) protein levels.
Main Results:
- * Increased levels of phosphorylated p53 (p-p53) were observed in both hepatopancreas and tail muscle during anoxia.
- * Elevated transcript levels of the ei24 gene indicated p53 activation under anoxic stress.
- * Cytoplasmic accumulation of Ser15 phosphorylated p53 was detected, suggesting a potential apoptotic response initiation.
- * Changes in MDM2 and p19-ARF levels indicated modulation of p53 activity.
Conclusions:
- * The tumor suppressor p53 is activated by anoxic stress in crayfish, suggesting a role in cellular defense.
- * p53 activation may initiate an apoptotic response to prevent autophagy and protect against DNA damage.
- * These findings highlight a potential protective mechanism in anoxia-tolerant organisms, offering insights into survival strategies.
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