Glucose Utilization Is Essential for Hypoxia-Inducible Factor 1α-Dependent Phosphorylation of c-Jun

Hypoxia and anoxia are important microenvironmental stresses that contribute to pathological events such as solid-tumor development. We have been investigating the effects of hypoxia and anoxia on expression of the proto-oncogene c-jun and the regulation of c-Jun/AP-1 transcription factors. In earlier work using genetically manipulated mouse embryo fibroblasts (mEFs), we found a functional relationship among c-jun expression, c-Jun N-terminal phosphorylation, and the presence of hypoxia-inducible factor 1α (HIF-1α), the oxygen-regulated subunit of the HIF-1 transcription factor. Both the induction of c-jun mRNA expression and c-Jun N-terminal phosphorylation in cells exposed to hypoxia or anoxia were found to be dependent on the presence of HIF-1α, but this was not the case in cells exposed to less-severe hypoxia. Here we describe new findings concerning HIF-1-dependent c-Jun N-terminal phosphorylation in cells exposed to hypoxia or anoxia. Specifically, we report that hypoxia-inducible c-Jun N-terminal kinase (JNK) activity, which involves JNKs or stress-activated protein kinases (SAPKs), is dependent on enhanced glucose utilization mediated by HIF-1. These results suggest a model in which hypoxia-inducible JNK activity is connected to oxygen sensing through increased glucose absorption and/or glycolytic activity regulated by the HIF-1 system. We also found that basal threonine and tyrosine phosphorylation (within the TEY motif) of extracellular signal-regulated kinases 1 and 2 (ERK1/2) and the corresponding ERK1/2 activity were defective in hypoxic HIF-1α-null mEFs but not in wild-type mEFs, independently of glucose uptake. Therefore, the activities of both JNKs/SAPKs and ERK1/2 are sensitive to HIF-1-dependent processes in cells exposed to hypoxia or anoxia

The Response of c-Jun/AP-1 to Chronic Hypoxia Is Hypoxia-Inducible Factor 1α Dependent

Hypoxia (low-oxygen tension) is an important physiological stress that influences responses to a wide range of pathologies, including stroke, infarction, and tumorigenesis. Prolonged or chronic hypoxia stimulates expression of the stress-inducible transcription factor gene c-jun and transient activation of protein kinase and phosphatase activities that regulate c-Jun/AP-1 activity. Here we describe evidence obtained by using wild-type and HIF-1α nullizygous mouse embryonic fibroblasts (mEFs) that the induction of c-jun mRNA expression and c-Jun phosphorylation by prolonged hypoxia are completely dependent on the presence of the oxygen-regulated transcription factor hypoxia-inducible factor 1α (HIF-1α). In contrast, transient hypoxia induced c-jun expression in both types of mEFs, showing that the early or rapid induction of this gene is independent of HIF-1α. These findings indicate that the c-jun gene has a biphasic response to hypoxia consisting of inductions that depend on the degree or duration of exposure. To more completely define the relationship between prolonged hypoxia and c-Jun phosphorylation, we used mEFs from mice containing inactivating mutations of critical phosphorylation sites in the c-Jun N-terminal region (serines 63 and 73 or threonines 91 and 93). Exposure of these mEFs to prolonged hypoxia demonstrated an absolute requirement for N-terminal sites for HIF-1α-dependent phosphorylation of c-Jun. Taken together, these findings suggest that c-Jun/AP-1 and HIF-1 cooperate to regulate gene expression in pathophysiological microenvironments