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Assays for the Degradation of Misfolded Proteins in Cells
Published on: August 28, 2016
Normoxic destabilization of ATF-4 depends on proteasomal degradation
M Wottawa1, J Köditz, D M Katschinski
1Department of Cardiovascular Physiology, Georg-August University Göttingen, Germany.
Acta Physiologica (Oxford, England)
|November 20, 2009
Summary
Activating transcription factor 4 (ATF-4) is degraded via oxygen-dependent proteasomal pathways under normoxia. Hypoxia stabilizes ATF-4, promoting GADD153 expression, crucial for cellular adaptation to low oxygen.
Area of Science:
- Cellular Biology
- Molecular Biology
- Physiology
Background:
- Hypoxia-inducible gene expression is a key adaptive mechanism to low oxygen.
- Activating transcription factor 4 (ATF-4) stabilization is regulated by oxygen and prolyl-4-hydroxylase (PHD)3.
- The mechanism of ATF-4 destabilization under normoxia requires further investigation.
Purpose of the Study:
- To investigate if normoxic destabilization of ATF-4 is mediated by oxygen-dependent proteasomal degradation.
- To elucidate the role of oxygen-dependent proteasomal degradation in hypoxia-induced gene expression.
Main Methods:
- Quantified ATF-4 poly-ubiquitination under normoxia versus hypoxia using immunoprecipitation and immunoblots.
- Analyzed the expression of the ATF-4 target gene GADD153 in response to varying oxygen concentrations.
- Utilized siRNA to modulate ATF-4 and PHD3 levels.
Main Results:
- ATF-4 protein was undetectable in normoxia, correlating with oxygen-dependent poly-ubiquitination.
- Hypoxic conditions hindered ATF-4 poly-ubiquitination and stabilized the protein.
- Hypoxia induced GADD153 expression, which was modulated by ATF-4 and PHD3 levels.
Conclusions:
- Oxygen-dependent proteasomal degradation of ATF-4 is involved in hypoxia-induced GADD153 expression.
- Hypoxia/PHD3-mediated stabilization of ATF-4 by inhibiting oxygen-dependent degradation is critical for linking cell fate to oxygen availability.
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