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Updated: Jun 18, 2026

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.
Aim:
Hypoxia-inducible gene expression is an important physiological adaptive mechanism in response to a decreased oxygen supply. We have recently described an oxygen- and prolyl-4-hydroxylase (PHD)3-dependent stabilization of the activating transcription factor 4 (ATF-4). The aim of the present study was to examine if the normoxic destabilization of ATF-4 is regulated by oxygen-dependent proteasomal degradation.
Methods:
We determined poly-ubiquitination of ATF-4 in normoxia compared to hypoxia by immunoprecipitation and immunoblots. Furthermore, we analysed the expression of the ATF-4 target gene GADD153 as a function of oxygen concentration.
Results:
ATF-4 protein levels were not detectable in normoxia. Normoxic degradation correlated with an oxygen-dependent poly-ubiquitination of ATF-4, which was hindered by hypoxic incubation of the cells. As a result of hypoxia, GADD153 was expressed. The hypoxic GADD153 expression was attenuated or increased by transfecting the cells with ATF-4 siRNA or PHD3 siRNA respectively.
Conclusion:
Our results demonstrate the involvement of oxygen-dependent proteasomal degradation of ATF-4 in the hypoxia-induced expression of GADD153. Taken together, hypoxia/PHD3-regulated stabilization of ATF-4 by hindering oxygen-dependent degradation may play a critical role in linking cell fate decisions to oxygen availability.
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