Oxygen-sensing mechanisms in development and tissue repair

ABSTRACT Under normoxia, hypoxia inducible factor (HIF) α subunits are hydroxylated by PHDs (prolyl hydroxylase domain proteins) and subsequently undergo polyubiquitylation and degradation. Normal embryogenesis occurs under hypoxia, which suppresses PHD activities and allows HIFα to stabilize and regulate development. In this Primer, we explain molecular mechanisms of the oxygen-sensing pathway, summarize HIF-regulated downstream events, discuss loss-of-function phenotypes primarily in mouse development, and highlight clinical relevance to angiogenesis and tissue repair.

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The Hypoxia-Inducible Factor Pathway, Prolyl Hydroxylase Domain Protein Inhibitors, and Their Roles in Bone Repair and Regeneration

BioMed Research International ◽

10.1155/2014/239356 ◽

2014 ◽

Vol 2014 ◽

pp. 1-11 ◽

Cited By ~ 25

Author(s):

Lihong Fan ◽

Jia Li ◽

Zefeng Yu ◽

Xiaoqian Dang ◽

Kunzheng Wang

Keyword(s):

Cell Fate ◽

Bone Repair ◽

Molecular Mechanisms ◽

Hypoxia Inducible Factor ◽

Prolyl Hydroxylase ◽

Bone Tumours ◽

Cell Fate Decisions ◽

Tightly Coupled ◽

Hif Pathway ◽

Prolyl Hydroxylase Domain

Hypoxia-inducible factors (HIFs) are oxygen-dependent transcriptional activators that play crucial roles in angiogenesis, erythropoiesis, energy metabolism, and cell fate decisions. The group of enzymes that can catalyse the hydroxylation reaction of HIF-1 is prolyl hydroxylase domain proteins (PHDs). PHD inhibitors (PHIs) activate the HIF pathway by preventing degradation of HIF-αvia inhibiting PHDs. Osteogenesis and angiogenesis are tightly coupled during bone repair and regeneration. Numerous studies suggest that HIFs and their target gene, vascular endothelial growth factor (VEGF), are critical regulators of angiogenic-osteogenic coupling. In this brief perspective, we review current studies about the HIF pathway and its role in bone repair and regeneration, as well as the cellular and molecular mechanisms involved. Additionally, we briefly discuss the therapeutic manipulation of HIFs and VEGF in bone repair and bone tumours. This review will expand our knowledge of biology of HIFs, PHDs, PHD inhibitors, and bone regeneration, and it may also aid the design of novel therapies for accelerating bone repair and regeneration or inhibiting bone tumours.

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Molecular Characterization and Response of Prolyl Hydroxylase Domain (PHD) Genes to Hypoxia Stress in Hypophthalmichthys molitrix

Animals ◽

10.3390/ani12020131 ◽

2022 ◽

Vol 12 (2) ◽

pp. 131

Author(s):

Xiaohui Li ◽

Meidong Zhang ◽

Chen Ling ◽

Hang Sha ◽

Guiwei Zou ◽

...

Keyword(s):

Molecular Mechanisms ◽

Silver Carp ◽

Hypoxia Inducible Factor ◽

Hypoxic Condition ◽

Early Response ◽

Prolyl Hydroxylase ◽

Alpha Subunit ◽

Hypophthalmichthys Molitrix ◽

Low Oxygen ◽

Prolyl Hydroxylase Domain

As an economically and ecologically important freshwater fish, silver carp (Hypophthalmichthys molitrix) is sensitive to low oxygen tension. Prolyl hydroxylase domain (PHD) proteins are critical regulators of adaptive responses to hypoxia for their function of regulating the hypoxia inducible factor-1 alpha subunit (HIF-1α) stability via hydroxylation reaction. In the present study, three PHD genes were cloned from H. molitrix by rapid amplification of cDNA ends (RACE). The total length of HmPHD1, HmPHD2, and HmPHD3 were 2981, 1954, and 1847 base pair (bp), and contained 1449, 1080, and 738 bp open reading frames (ORFs) that encoded 482, 359, and 245 amino acids (aa), respectively. Amino acid sequence analysis showed that HmPHD1, HmPHD2, and HmPHD3 had the conserved prolyl 4-hydroxylase alpha subunit homolog domains at their C-termini. Meanwhile, the evaluation of phylogeny revealed PHD2 and PHD3 of H. molitrix were more closely related as they belonged to sister clades, whereas the clade of PHD1 was relatively distant from these two. The transcripts of PHD genes are ubiquitously distributed in H. molitrix tissues, with the highest expressional level of HmPHD1 and HmPHD3 in liver, and HmPHD2 in muscle. After acute hypoxic treatment for 0.5 h, PHD genes of H. molitrix were induced mainly in liver and brain, and different from HmPHD1 and HmPHD2, the expression of HmPHD3 showed no overt tissue specificity. Furthermore, under continued hypoxic condition, PHD genes exhibited an obviously rapid but gradually attenuated response from 3 h to 24 h, and upon reoxygenation, the transcriptional expression of PHD genes showed a decreasing trend in most of the tissues. These results indicate that the PHD genes of H. molitrix are involved in the early response to hypoxic stress, and they show tissue-specific transcript expression when performing physiological regulation functions. This study is of great relevance for advancing our understanding of how PHD genes are regulated when addressing the hypoxic challenge and provides a reference for the subsequent research of the molecular mechanisms underlying hypoxia adaptation in silver carp.

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Regulation of adult erythropoiesis by prolyl hydroxylase domain proteins

Blood ◽

10.1182/blood-2007-09-114561 ◽

2008 ◽

Vol 111 (6) ◽

pp. 3229-3235 ◽

Cited By ~ 180

Author(s):

Kotaro Takeda ◽

Hector L. Aguila ◽

Nehal S. Parikh ◽

Xiping Li ◽

Katie Lamothe ◽

...

Keyword(s):

Hypoxia Inducible Factor ◽

Prolyl Hydroxylase ◽

Heterozygous Mutation ◽

Adult Liver ◽

Double Knockout ◽

Adult Mice ◽

Liver And Kidney ◽

Prolyl Hydroxylation ◽

Α Subunits ◽

Prolyl Hydroxylase Domain

Abstract Polycythemia is often associated with erythropoietin (EPO) overexpression and defective oxygen sensing. In normal cells, intracellular oxygen concentrations are directly sensed by prolyl hydroxylase domain (PHD)–containing proteins, which tag hypoxia-inducible factor (HIF) α subunits for polyubiquitination and proteasomal degradation by oxygen-dependent prolyl hydroxylation. Here we show that different PHD isoforms differentially regulate HIF-α stability in the adult liver and kidney and suppress Epo expression and erythropoiesis through distinct mechanisms. Although Phd1−/− or Phd3−/− mice had no apparent defects, double knockout of Phd1 and Phd3 led to moderate erythrocytosis. HIF-2α, which is known to activate Epo expression, accumulated in the liver. In adult mice deficient for PHD2, the prototypic Epo transcriptional activator HIF-1α accumulated in both the kidney and liver. Elevated HIF-1α levels were associated with dramatically increased concentrations of both Epo mRNA in the kidney and Epo protein in the serum, which led to severe erythrocytosis. In contrast, heterozygous mutation of Phd2 had no detectable effects on blood homeostasis. These findings suggest that PHD1/3 double deficiency leads to erythrocytosis partly by activating the hepatic HIF-2α/Epo pathway, whereas PHD2 deficiency leads to erythrocytosis by activating the renal Epo pathway.

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The Zinc Finger of Prolyl Hydroxylase Domain Protein 2 Is Essential for Efficient Hydroxylation of Hypoxia-Inducible Factor α

Molecular and Cellular Biology ◽

10.1128/mcb.00090-16 ◽

2016 ◽

Vol 36 (18) ◽

pp. 2328-2343 ◽

Cited By ~ 7

Author(s):

Patrick R. Arsenault ◽

Daisheng Song ◽

Yu Jin Chung ◽

Tejvir S. Khurana ◽

Frank S. Lee

Keyword(s):

High Altitude ◽

Zinc Finger ◽

Hypoxia Inducible Factor ◽

Prolyl Hydroxylase ◽

Loss Of Function ◽

Domain Protein ◽

Factor Α ◽

Prolyl Hydroxylase Domain

Prolyl hydroxylase domain protein 2 (PHD2) (also known as EGLN1) is a key oxygen sensor in mammals that posttranslationally modifies hypoxia-inducible factor α (HIF-α) and targets it for degradation. In addition to its catalytic domain, PHD2 contains an evolutionarily conserved zinc finger domain, which we have previously proposed recruits PHD2 to the HSP90 pathway to promote HIF-α hydroxylation. Here, we provide evidence that this recruitment is critical bothin vitroandin vivo. We show thatin vitro, the zinc finger can function as an autonomous recruitment domain to facilitate interaction with HIF-α.In vivo, ablation of zinc finger function by a C36S/C42SEgln1knock-in mutation results in upregulation of theerythropoietingene, erythrocytosis, and augmented hypoxic ventilatory response, all hallmarks ofEgln1loss of function and HIF stabilization. Hence, the zinc finger ordinarily performs a critical positive regulatory function. Intriguingly, the function of this zinc finger is impaired in high-altitude-adapted Tibetans, suggesting that their adaptation to high altitude may, in part, be due to a loss-of-functionEGLN1allele. Thus, these findings have important implications for understanding both the molecular mechanism of the hypoxic response and human adaptation to high altitude.

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Enhancement of Angiogenesis Through Stabilization of Hypoxia-inducible Factor-1 by Silencing Prolyl Hydroxylase Domain-2 Gene

Molecular Therapy ◽

10.1038/mt.2008.90 ◽

2008 ◽

Vol 16 (7) ◽

pp. 1227-1234 ◽

Cited By ~ 28

Author(s):

Shourong Wu ◽

Nobuhiro Nishiyama ◽

Mitsunobu R Kano ◽

Yasuyuki Morishita ◽

Kohei Miyazono ◽

...

Keyword(s):

Hypoxia Inducible Factor ◽

Prolyl Hydroxylase ◽

Hypoxia Inducible Factor 1 ◽

Prolyl Hydroxylase Domain

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Prolyl hydroxylase domain-containing protein inhibitors as stabilizers of hypoxia-inducible factor: small molecule-based therapeutics for anemia

Expert Opinion on Therapeutic Patents ◽

10.1517/13543776.2010.510836 ◽

2010 ◽

Vol 20 (9) ◽

pp. 1219-1245 ◽

Cited By ~ 59

Author(s):

Lin Yan ◽

Vincent J Colandrea ◽

Jeffrey J Hale

Keyword(s):

Small Molecule ◽

Hypoxia Inducible Factor ◽

Prolyl Hydroxylase ◽

Protein Inhibitors ◽

Prolyl Hydroxylase Domain

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Abstract 396: Inhibition of Prolyl Hydroxylase Domain-containing Protein Downregulates Vascular Angiotensin II Type 1 Receptor

Hypertension ◽

10.1161/hyp.60.suppl_1.a396 ◽

2012 ◽

Vol 60 (suppl_1) ◽

Author(s):

Toshihiro Ichiki

Keyword(s):

Cellular Response ◽

Target Genes ◽

Interstitial Fibrosis ◽

Critical Role ◽

Hypoxia Inducible Factor ◽

Renin Angiotensin System ◽

Prolyl Hydroxylase ◽

Ang Ii ◽

Prolyl Hydroxylase Domain

Background: Prolyl hydroxylase domain-containing protein (PHD) mediates hydroxylation of hypoxia-inducible factor (HIF)-1α and thereby induces proteasomal degradation of HIF-1α. Inhibition of PHD by hypoxia or hypoxia mimetics such as cobalt chloride (CoCl2) stabilizes HIF-1 and increases the expression of target genes such as vascular endothelial growth factor (VEGF). Although hypoxia activates the systemic renin angiotensin system (RAS), the role of PHD in regulating RAS remains unknown. We examined the effect of PHD inhibition on the expression of angiotensin (Ang) II type 1 receptor (AT1R) and its signaling. Methods and Results: Hypoxia (1% O2), CoCl2 (100-300 μmol/L), and dimethyloxalylglycine (0.25-1.0 mmol/L), all known to inhibit PHD, reduced AT1R expression by 37.7±7.6, 39.6±8.4-69.7±9.9, and 13.4±6.1-25.2±7.0%, respectively (p<0.01) in cultured vascular smooth muscle cell. The same stimuli increased the expression of nuclear HIF-1α and VEGF (p<0.05), suggesting that PHD activity is inhibited. Knockdown of PHD2, a major isoform of PHDs, by RNA interference also reduced AT1R expression by 55.3±6.0% (p<0.01). CoCl2 decreased AT1R mRNA through transcriptional and posttranscriptional mechanisms (p<0.01 and <0.05, respectively). CoCl2 and PHD2 knockdown diminished Ang II-induced ERK phosphorylation (P<0.01). Over-expression of the constitutively active HIF-1α did not impact the AT1R gene promoter activity. Oral administration of CoCl2 (14 mg/kg/day) to C57BL/6J mice receiving Ang II infusion (490 ng/kg/min) for 4 weeks significantly reduced the expression of AT1R in the aorta by 60.9±11.3% (p<0.05) and attenuated coronary perivascular fibrosis by 85% (p<0.01) without affecting blood pressure. However, CoCl2 did not affect Ang II-induced renal interstitial fibrosis. Conclusion: PHD inhibition downregulates AT1R expression independently of HIF-1α, reduces the cellular response to Ang II, and attenuates profibrotic effect of Ang II on the coronary arteries. PHD inhibition may be beneficial for the treatment of cardiovascular diseases, in which activation of RAS plays a critical role.

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