ENZYMES INVOLVED IN HYPOXIA RESPONSE, ACTA UNIVERSITATIS OULUENSIS D Medica 1055
|Kustantaja:||Oulun yliopisto|| |
|Sijainti:||Print Tietotalo|| |
|Tekijät:||HYVÄRINEN JAANA|| |
Oxygen homeostasis is critical to all animals, as both excess (hyperoxia) and reduced (hypoxia) levels of oxygen can result in pathological changes and ultimately in the loss of cellular and organismal viability. Complex systems have evolved to sense and adapt to changes in cellular oxygen availability, and the hypoxia-inducible factor HIF plays a pivotal role in this elaborate molecular network. In normoxic conditions the á-subunit of HIF becomes hydroxylated by HIF prolyl 4-hydroxylases (HIF-P4Hs 1-3), earmarking HIF-á for proteasomal degradation. Additionally, in the presence of oxygen the hydroxylation of an asparagine residue by the HIF asparaginyl hydroxylase FIH inhibits the transactivation of HIF-target genes by blocking the interaction of HIF-á with a transcriptional coactivator. In addition to being a feature of an organism’s normal life, hypoxia is also characteristic of many common diseases such as severe anemia and myocardial infarction, and it notably decreases these hydroxylation reactions, as HIFP4Hs and FIH have an absolute requirement for oxygen as a cosubstrate. HIF-á thus escapes degradation and translocates into the nucleus, where it dimerizes with HIF-â and recruits transcriptional coactivators to the hypoxia-response elements of target genes, inducing their transcription and triggering the hypoxia response aimed at restoring cellular oxygen homeostasis.
In this study we generated a genetically modified HIF-P4H-2 hypomorphic mouse line that expresses only 8% of the wild-type HIF-P4H-2 mRNA in the heart. We showed that chronic cardiac HIF-P4H-2 deficiency leads to stabilization of HIF-1á and HIF-2á and protects the heart against acute ischemia-reperfusion injury without causing any adverse effects.
Furthermore, we identified and cloned a novel human transmembrane prolyl 4-hydroxylase P4H-TM and showed that it regulates HIF-1á protein levels in cellulo and hydroxylates HIF-1á in vitro similarly to the HIF-P4Hs, but may also have other physiological substrates. Using forward genetic tools we showed that lack of P4H-TM during development leads to basement membrane defects and compromised kidney function in zebrafish embryos.
Finally, we demonstrated that FIH displays substrate selectivity in terms of hydroxylation and binding of HIF-1á and novel substrates Notch1-3. We showed that FIH has higher affinity for oxygen with Notch1 than with HIF-1á as a substrate, implying that FIH-mediated hydroxylation of Notch can continue in oxygen concentrations where HIF-1á hydroxylation would be markedly reduced.