Normoxic destabilization of ATF-4 depends on proteasomal degradation.
OBJECTIVE
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.


CONCLUSIONS
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.
Wottawa
M
M
Department of Cardiovascular Physiology, Georg-August University Göttingen, Germany.
Köditz
J
J
Katschinski
D M
DM
D051701:Activating Transcription Factor 4 / Q000378:metabolism / Q000502:physiology*; D002460:Cell Line; D045744:Cell Line, Tumor; D006367:HeLa Cells; D006801:Humans; D010100:Oxygen / Q000378:metabolism*; D010101:Oxygen Consumption / Q000502:physiology*; D011393:Procollagen-Proline Dioxygenase / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D034741:RNA, Small Interfering; D051742:Transcription Factor CHOP / Q000187:drug effects / Q000235:genetics*; D015533:Transcriptional Activation / Q000502:physiology; D054875:Ubiquitination / Q000502:physiology
C497249:DDIT3 protein, human; D034741:RNA, Small Interfering; D051701:Activating Transcription Factor 4; D051742:Transcription Factor CHOP; D011393:Procollagen-Proline Dioxygenase; C030606:proline, 2-oxoglutarate 3-dioxygenase; D046988:Proteasome Endopeptidase Complex; D010100:Oxygen