Oxygen-dependent regulation of HIF-1alpha expression and function

Hypoxia is a state of reduced oxygen levels below normal values. It occurs in the pathophysiology of many diseases and plays important roles in mammalian physiology and development. The hypoxia-inducible factors (HIFs) are transcription factors that mediate adaptive cellular responses to hypoxia by activating the transcription of target genes. HIF is a heterodimer consisting of an oxygen-regulated alpha subunit and a constitutively expressed HIF-1beta (ARNT) subunit. Both HIF subunits belong to the bHLH/PAS domain-containing protein family. The bHLH and PAS domains mediate DNA binding and dimerization. HIF-alpha contains a unique oxygen-dependent degradation (ODD) domain and two transactivation domains (TADs: N-TAD and CTAD).

The oxygen-dependent degradation of HIF-alpha is mediated by PHD-dependent hydroxylation, and subsequent pVHL-dependent ubiquitylation and proteasomal degradation. Under hypoxia, HIF-alpha is stabilized and accumulates in the nucleus where it dimerizes with ARNT and activates target gene expression by recruiting coactivators. FIH-1 inhibits the transcriptional activity of HIF-alpha by inhibiting the recruitments of coactivators such as CBP/p300 through the hydroxylation of Asparagine residue in CTAD.

The aim of this study is to investigate in detail oxygen-dependent regulation of HIF-alpha degradation and function. In paper I, we identified residues that are critical for the degradation and transactivation functions of HIF-1alpha N-TAD.

In paper II, we demonstrated that the expression of pVHL-interacting peptides activates endogenous HIF-1alpha function, providing a potential pro-angiogenic stimulus. The degradation of HIF-1alpha is regulated by subcellular compartmentalization in a cell-type-dependent manner. The compartment in which degradation of HIF-1alpha occurs dictates where the pVHL-interacting peptide must be located in order to efficiently increase HIF-1alpha function at normoxia.

In paper III, we identified filamin A (FLNa) as a novel modulator of the HIF-1alpha pathway and presented a novel nuclear function of this actin-binding protein. Hypoxia enhances calpain-mediated cleavage of FLNa and the nuclear localization of the cleaved C-terminal fragment of FLNa. The C-terminal fragment of FLNa mediates the function of FLNa by facilitating the nuclear localization and enhancing the transcriptional activity of HIF-1alpha. FLNa promotes VEGF-A-mediated angiogenesis through activating HIF-1alpha-mediated hypoxic responses.

In paper IV, we demonstrated how hypoxia controls the differentiation status of a cell and demonstrates a crosstalk between the HIF-1alpha and Notch signaling pathways. Hypoxia blocks differentiation in myoblasts, satellite cells and primary neural stem cells in a Notch-dependent manner. Hypoxia stabilizes the Notch intracellular domain, activates Notch-responsive promoters and increases expression of Notch-regulated downstream genes. The modulation of the Notch pathway by hypoxia is mediated by HIF-1alpha. HIF-1alpha interacts with Notch ICD, binds to Notch-responsive promoter, and a transcriptionally active form of HIF-alpha increases Notch signaling.

The results from these studies contribute to a better understanding of the mechanism, which controls oxygen-dependent regulation and function of HIF-1alpha, and provide therapeutic implications by novel means to modulate the HIF-1alpha signaling pathway.

List of scientific papers

I. Pereira T, Zheng X, Ruas JL, Tanimoto K, Poellinger L (2003). Identification of residues critical for regulation of protein stability and the transactivation function of the hypoxia-inducible factor-1alpha by the von Hippel-Lindau tumor suppressor gene product. J Biol Chem. 278(9): 6816-23. Epub 2002 Dec 4
https://pubmed.ncbi.nlm.nih.gov/12468553

II. Zheng X, Ruas JL, Cao R, Salomons FA, Cao Y, Poellinger L, Pereira T (2006). Cell-type-specific regulation of degradation of hypoxia-inducible factor 1 alpha: role of subcellular compartmentalization. Mol Cell Biol. 26(12): 4628-41
https://pubmed.ncbi.nlm.nih.gov/16738327

III. Zheng X, Zhou X, Björndahl M, Uramoto H, Ganesh L, Nabel EG, Cao Y, Borén J, Pereira T, Poellinger L, Akyürek LM (2009). Filamin-A promotes VEGF-A activity through the HIF-1a-mediated hypoxic response. [Manuscript]

IV. Gustafsson MV, Zheng X, Pereira T, Gradin K, Jin S, Lundkvist J, Ruas JL, Poellinger L, Lendahl U, Bondesson M (2005). Hypoxia requires notch signaling to maintain the undifferentiated cell state. Dev Cell. 9(5): 617-28
https://pubmed.ncbi.nlm.nih.gov/16256737