A23187, lonomycin and thapsigargin upregulate mRNA of HIP-1 alpha via endoplasmic reticulum stress rather than a rise in intracellular calcium

Hypoxia inducible factor 1(HIF-1) coordinates major responses to hypoxia, with the notion that its alpha subunit is degraded under normoxia but stable under hypoxia. Recently, calcium was shown to affect expression of HIF-1 alpha. While lowering cytosolic calcium accumulates HIF-1 alpha, a calcium increase by the ionophores A23187 or ionomycin as well as the endoplasmic reticulum (ER) Ca(2+)-ATPase inhibitor thapsigargin produced inconsistent results with reports to either increase or decrease HIF-1 alpha protein. In the human hepatocyte cell line HepG2 only A23187, but neither ionomycin nor thapsigargin, accumulated HIF-1 alpha protein under normoxia. However, A23187 does so independently of a calcium increase. A23187 not only accumulated HIF-1 alpha protein but also mRNA of HIF-1 alpha, with the notion that protein but not mRNA accumulation was attenuated by blocking mitogen-activated protein kinase (MAPK), suggesting that HIF-1 alpha protein accumulation is not a direct consequence of its mRNA elevation. Indeed, protein stability determinations implied that A23187 enhanced translation of HIF-1 alpha. Interestingly, ionomycin and thapsigargin also increased the HIF-1 alpha mRNA content. Although not increasing the HIF-1 alpha protein amount under normoxia, both compounds enhanced protein accumulation of HIF-1 alpha under hypoxia. Taking into account that induction of ER stress by tunicamycin and brefeldin A, without altering intracellular calcium concentrations, also increased HIF-1 alpha mRNA, suggests that ER stress pathways enhanced transcription of HIF-1 alpha mRNA. We conclude that ER stress rather than calcium fluctuations increased HIF-1 alpha mRNA content by established calcium liberating agents, which alone is insufficient for normoxic HIF-1 alpha accumulation.