Hypoxia-mediated control of HIF/ARNT machinery in epidermal keratinocytes

Transcriptional activity of hypoxia-induced factor 1 (HIF1) – a heterodimer of HIF1α and ARNT (HIF1β) – is essential for cellular adaptation to environmental stress and plays an important role in skin development, wound healing, tumorigenesis and barrier function. Using primary mouse and human epidermal keratinocytes at ambient or hypoxic (1% O2) conditions we studied effects of hypoxia upon HIF protein expression. Significant nuclear levels of ARNT and HIF1α along with high HIF1 activity in normoxic keratinocytes suggest an as yet uncharacterised oxygen-independent role for HIF pathway in the epidermis. Acute hypoxia results in an instant but transient increase of HIF1α protein accompanied by a gradual decrease in its mRNA, while ARNT expression remains unchanged. In prolonged (chronic) hypoxia both HIF1α and Arnt are downregulated along with decline of HIF1 activity. However, expression of classical HIF1 targets such as Selenbp1 and Vegfa remains high. Thus, keratinocytes respond to acute hypoxia with immediate block of HIF1α protein degradation and concomitant increase of HIF activity, while under chronic hypoxia pro-angiogenic signalling is maintained through HIF1-independent pathway(s). Decline of HIF1α during chronic exposure is controlled at both mRNA and protein levels, while Arnt is downregulated post-translationally. Distinct transcription levels of Hif1α and Hif3α splice variants under normoxia and their differential response to hypoxia suggest functional diversity of Hif-α isoforms and highlight the complexity of HIF machinery control in epidermal keratinocytes.

Research highlights
► Hif1α and Arnt proteins have hypoxia-independent functions in the epidermis.
► The keratinocytes respond to acute and chronic hypoxia through different mechanisms.
► Hif1 activity in keratinocytes is regulated by hypoxia on transcriptional level as well.
► Protein-coding splice variants of Hif1α and Hif3α are differentially regulated by hypoxia.
► Hif1 activity in keratinocytes is regulated by hypoxia through multiple transcriptional and translational mechanisms.