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

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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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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Oxygen-sensing mechanisms in development and tissue repair

Development ◽

10.1242/dev.200030 ◽

2021 ◽

Vol 148 (23) ◽

Author(s):

Yida Jiang ◽

Li-Juan Duan ◽

Guo-Hua Fong

Keyword(s):

Clinical Relevance ◽

Tissue Repair ◽

Molecular Mechanisms ◽

Oxygen Sensing ◽

Hypoxia Inducible Factor ◽

Prolyl Hydroxylase ◽

Loss Of Function ◽

Mouse Development ◽

Α Subunits ◽

Prolyl Hydroxylase Domain

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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Hypoxia, Metabolic Reprogramming, and Drug Resistance in Liver Cancer

Cells ◽

10.3390/cells10071715 ◽

2021 ◽

Vol 10 (7) ◽

pp. 1715

Author(s):

Macus Hao-Ran Bao ◽

Carmen Chak-Lui Wong

Keyword(s):

Hepatocellular Carcinoma ◽

Drug Resistance ◽

Liver Cancer ◽

Effective Treatment ◽

Molecular Mechanisms ◽

Hypoxia Inducible Factor ◽

Metabolic Reprogramming ◽

Combination Therapies ◽

Low Oxygen ◽

Treatment Regimens

Hypoxia, low oxygen (O2) level, is a hallmark of solid cancers, especially hepatocellular carcinoma (HCC), one of the most common and fatal cancers worldwide. Hypoxia contributes to drug resistance in cancer through various molecular mechanisms. In this review, we particularly focus on the roles of hypoxia-inducible factor (HIF)-mediated metabolic reprogramming in drug resistance in HCC. Combination therapies targeting hypoxia-induced metabolic enzymes to overcome drug resistance will also be summarized. Acquisition of drug resistance is the major cause of unsatisfactory clinical outcomes of existing HCC treatments. Extra efforts to identify novel mechanisms to combat refractory hypoxic HCC are warranted for the development of more effective treatment regimens for HCC patients.

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Hypoxic tumor microenvironment: Implications for cancer therapy

Experimental Biology and Medicine ◽

10.1177/1535370220934038 ◽

2020 ◽

Vol 245 (13) ◽

pp. 1073-1086

Author(s):

Sukanya Roy ◽

Subhashree Kumaravel ◽

Ankith Sharma ◽

Camille L Duran ◽

Kayla J Bayless ◽

...

Keyword(s):

Tumor Microenvironment ◽

Cancer Cells ◽

Metabolic Regulation ◽

Molecular Mechanisms ◽

Hypoxia Inducible Factor ◽

Tumor Vasculature ◽

Therapeutic Strategies ◽

Therapeutic Resistance ◽

Low Oxygen ◽

Cancer Types

Hypoxia or low oxygen concentration in tumor microenvironment has widespread effects ranging from altered angiogenesis and lymphangiogenesis, tumor metabolism, growth, and therapeutic resistance in different cancer types. A large number of these effects are mediated by the transcription factor hypoxia inducible factor 1⍺ (HIF-1⍺) which is activated by hypoxia. HIF1⍺ induces glycolytic genes and reduces mitochondrial respiration rate in hypoxic tumoral regions through modulation of various cells in tumor microenvironment like cancer-associated fibroblasts. Immune evasion driven by HIF-1⍺ further contributes to enhanced survival of cancer cells. By altering drug target expression, metabolic regulation, and oxygen consumption, hypoxia leads to enhanced growth and survival of cancer cells. Tumor cells in hypoxic conditions thus attain aggressive phenotypes and become resistant to chemo- and radio- therapies resulting in higher mortality. While a number of new therapeutic strategies have succeeded in targeting hypoxia, a significant improvement of these needs a more detailed understanding of the various effects and molecular mechanisms regulated by hypoxia and its effects on modulation of the tumor vasculature. This review focuses on the chief hypoxia-driven molecular mechanisms and their impact on therapeutic resistance in tumors that drive an aggressive phenotype. Impact statement Hypoxia contributes to tumor aggressiveness and promotes growth of many solid tumors that are often resistant to conventional therapies. In order to achieve successful therapeutic strategies targeting different cancer types, it is necessary to understand the molecular mechanisms and signaling pathways that are induced by hypoxia. Aberrant tumor vasculature and alterations in cellular metabolism and drug resistance due to hypoxia further confound this problem. This review focuses on the implications of hypoxia in an inflammatory TME and its impact on the signaling and metabolic pathways regulating growth and progression of cancer, along with changes in lymphangiogenic and angiogenic mechanisms. Finally, the overarching role of hypoxia in mediating therapeutic resistance in cancers is discussed.

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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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Hypoxia Activates SUMO-1-HIF-1α Signaling Pathway to Upregulate Pro-inflammatory Cytokines and Permeability in Human Tonsil Epithelial Cells

10.21203/rs.3.rs-157614/v1 ◽

2021 ◽

Author(s):

Yan Lin ◽

Mingjing Wang ◽

Zhen Xiao ◽

Zhiyan Jiang

Keyword(s):

Epithelial Cells ◽

Signaling Pathway ◽

Behavioral Problems ◽

Molecular Mechanisms ◽

Inflammatory Responses ◽

Hypoxia Inducible Factor ◽

Hypoxic Condition ◽

Human Tonsil ◽

Vitro Model

Abstract Adenoid hypertrophy (AH) can cause harmful effects on untreated children, which include mouth breathing, chronic intermittent hypoxia, sleep disordered breathing (SDB), and even some behavioral problems. However, the molecular mechanisms underlying this pathophysiological process have remained poorly understood. In this study, with use of a variety of biochemical approaches including gene silencing and transiently ectopic protein expression, we examined the molecular effectors involved in this process in an in vitro model of human tonsil epithelial cells (HTECs). We found that a hypoxic condition caused a dramatic upregulation of SUMO-1 expression, a member of the ubiquitin-like protein family, which in turn stabilized hypoxia-inducible factor (HIF)-1α by sumoylating this HIF subunit and thus preventing its ubiquitination and degradation in HTECs. We also found that activating HIF-1α promoted permeability of HTEC cells as well as production and secretion of a variety of proinflammatory cytokines including IL-6, IL-8, and TNF-α, and pro-angiogenic growth factor VEGF. Furthermore, our data showed that hypoxia-induced inflammation was markedly inhibited by M2 macrophages that possess potent anti-inflammatory function. Our results suggest that selectively inhibiting the SUMO-1-HIF-1α signaling pathway leads to inflammatory responses in human tonsil epithelial cells, which might be a novel therapeutic approach for managing hypoxia-induced SDB resulting from AH.

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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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