Hypoxia-inducible Transcription Factor Research Articles

Erythropoietin Production in Embryonic Neural Cells is Controlled by Hypoxia Signaling and Histone Deacetylases with an Undifferentiated Cellular State.

During mammalian development, production sites of the erythroid growth factor erythropoietin (EPO) shift from the neural tissues to the liver in embryos and to the kidneys in adults. Embryonic neural EPO-producing (NEP) cells, a subpopulation of neuroepithelial and neural crest cells, express the Epo gene between embryonic day (E) 8.5 and E11.5 to promote primitive erythropoiesis in mice. While Epo gene expression in the liver and kidneys is induced under hypoxic conditions through hypoxia-inducible transcription factors (HIFs), the Epo gene regulatory mechanisms in NEP cells remain to be elucidated. Here, we confirmed the presence of cells co-expressing EPO and HIFs in mouse neural tubes, where the hypoxic microenvironment activates HIFs. Chemical activation and inhibition of HIFs demonstrated the hypoxic regulation of EPO expression in human fetal neural progenitors and mouse embryonic neural tissues. In addition, we found that histone deacetylase inhibitors can reactivate EPO production in cell lines derived from NEP cells and human neuroblastoma, as well as in mouse primary neural crest cells, while rejuvenating these cells. Furthermore, the ability of the rejuvenated cells to produce EPO was maintained in hypoxia. Thus, EPO production is controlled by epigenetic mechanisms and hypoxia signaling in the immature state of hypoxic NEP cells.

A closer look at the role of deubiquitinating enzymes in the Hypoxia Inducible Factor pathway.

Hypoxia Inducible transcription Factors (HIFs) are central to the metazoan oxygen-sensing response. Under low oxygen conditions (hypoxia), HIFs are stabilised and govern an adaptive transcriptional programme to cope with prolonged oxygen starvation. However, when oxygen is present, HIFs are continuously degraded by the proteasome in a process involving prolyl hydroxylation and subsequent ubiquitination by the Von Hippel Lindau (VHL) E3 ligase. The essential nature of VHL in the HIF response is well established but the role of other enzymes involved in ubiquitination is less clear. Deubiquitinating enzymes (DUBs) counteract ubiquitination and provide an important regulatory aspect to many signalling pathways involving ubiquitination. In this review, we look at the complex network of ubiquitination and deubiquitination in controlling HIF signalling in normal and low oxygen tensions. We discuss the relative importance of DUBs in opposing VHL, and explore roles of DUBs more broadly in hypoxia, in both VHL and HIF independent contexts. We also consider the catalytic and non-catalytic roles of DUBs, and elaborate on the potential benefits and challenges of inhibiting these enzymes for therapeutic use.

Clinical characteristics of patients with P4HTM variant-associated epilepsy and therapeutic exploration: a case report and literature review.

The P4HTM gene encodes a transmembrane prolyl 4-hydroxylase, which is responsible for the degradation of hypoxia-inducible transcription factors (HIF) under normoxia. Clinically, biallelic P4HTM variants have been identified in patients with hypotonia, hypoventilation, intellectual disabilities, dysautonomia, epilepsy, and eye abnormalities (HIDEA syndrome). Seizure was one of the most prominent symptoms. However, the clinical features of patients with epilepsy associated with P4HTM variants remain unclear. In this report, we describe a one-month-old infant with HIDEA syndrome caused by compound heterozygous P4HTM variants (c.300dupG/p.Gly103Argfs*22 and c.488C > T/p.Ala163Val). The infant presented with clonic seizures of focal onset that responded well to valproate, but with profound intellectual disability and global developmental delay at the last follow-up at 3 years old. A review of the existing literature indicates that seizures in this population typically begin early in infancy, manifest in multiple types, and are relatively well controlled. Epilepsy seemed unrelated to developmental outcomes or disease progression. Valproate, which has HIF-1α inhibiting properties, may be a promising treatment avenue for this population.

Human prolyl hydroxylase domain 2 reacts with O2 and 2-oxoglutarate to enable formation of inactive Fe(III).2OG.hypoxia-inducible-factor α complexes

Hypoxia inducible transcription factors (HIFs) mediate the hypoxic response in metazoans. When sufficient O2 is present, Fe(II)/2-oxoglutarate (2OG)-dependent oxygenases (human PHD1-3) promote HIFα degradation via prolyl-hydroxylation. We report crystallographic, spectroscopic, and biochemical characterization of stable and inactive PHD2.Fe(III).2OG complexes. Aerobic incubation of PHD2 with Fe(II) and 2OG enables formation of PHD2.Fe(III).2OG complexes which bind HIF1-2α to give inactive PHD2.Fe(III).2OG.HIF1-2α complexes. The Fe(III) oxidation state in the inactive complexes was shown by EPR spectroscopy. L-Ascorbate hinders formation of the PHD2.Fe(III).2OG.(+/-HIFα) complexes and slowly regenerates them to give the catalytically active PHD2.Fe(II).2OG complex. Crystallographic comparison of the PHD2.Fe(III).2OG.HIF2α complex with the analogous anaerobic Fe(II) complex reveals near identical structures. Exposure of the anaerobic PHD2.Fe(II).2OG.HIF2α crystals to O2 enables in crystallo hydroxylation. The resulting PHD2.product structure, manifests conformational changes compared to the substrate structures. The results have implications for the role of the PHDs in hypoxia sensing and open new opportunities for inhibition of the PHDs and other 2OG dependent oxygenases by promoting formation of stable Fe(III) complexes.

Gating residues govern ligand unbinding kinetics from the buried cavity in HIF-2α PAS-B.

While transcription factors have been generally perceived as "undruggable," an exception is the HIF-2 hypoxia-inducible transcription factor, which contains an internal cavity that is sufficiently large to accommodate a range of small-molecules, including the therapeutically used inhibitor belzutifan. Given the relatively long ligand residence times of these small molecules and the lack of any experimentally observed pathway connecting the cavity to solvent, there has been great interest in understanding how these drug ligands exit the buried receptor cavity. Here, we focus on the relevant PAS-B domain of hypoxia-inducible factor 2α (HIF-2α) and examine how one such small molecule (THS-017) exits from the buried cavity within this domain on the seconds-timescale using atomistic simulations and ZZ-exchange NMR. To enable the simulations, we applied the weighted ensemble path sampling strategy, which generates continuous pathways for a rare-event process [e.g., ligand (un)binding] with rigorous kinetics in orders of magnitude less computing time compared to conventional simulations. Results reveal the formation of an encounter complex intermediate and two distinct classes of pathways for ligand exit. Based on these pathways, we identified two pairs of conformational gating residues in the receptor: one for the major class (N288 and S304) and another for the minor class (L272 and M309). ZZ-exchange NMR validated the kinetic importance of N288 for ligand unbinding. Our results provide an ideal simulation dataset for rational manipulation of ligand unbinding kinetics.

Gas Tunnel Engineering of Prolyl Hydroxylase Reprograms Hypoxia Signaling in Cells

AbstractCells have evolved intricate mechanisms for recognizing and responding to changes in oxygen (O2) concentrations. Here, we have reprogrammed cellular hypoxia (low O2) signaling via gas tunnel engineering of prolyl hydroxylase 2 (PHD2), a non‐heme iron dependent O2 sensor. Using computational modeling and protein engineering techniques, we identify a gas tunnel and critical residues therein that limit the flow of O2 to PHD2’s catalytic core. We show that systematic modification of these residues can open the constriction topology of PHD2’s gas tunnel. Using kinetic stopped‐flow measurements with NO as a surrogate diatomic gas, we demonstrate up to 3.5‐fold enhancement in its association rate to the iron center of tunnel‐engineered mutants. Our most effectively designed mutant displays 9‐fold enhanced catalytic efficiency (kcat/KM=830±40 M−1 s−1) in hydroxylating a peptide mimic of hypoxia inducible transcription factor HIF‐1α, as compared to WT PHD2 (kcat/KM=90±9 M−1 s−1). Furthermore, transfection of plasmids that express designed PHD2 mutants in HEK‐293T mammalian cells reveal significant reduction of HIF‐1α and downstream hypoxia response transcripts under hypoxic conditions of 1 % O2. Overall, these studies highlight activation of PHD2 as a new pathway to reprogram hypoxia responses and HIF signaling in cells.

Gas Tunnel Engineering of Prolyl Hydroxylase Reprograms Hypoxia Signaling in Cells.

Cells have evolved intricate mechanisms for recognizing and responding to changes in oxygen (O2) concentrations. Here, we have reprogrammed cellular hypoxia (low O2) signaling via gas tunnel engineering of prolyl hydroxylase 2 (PHD2), a non-heme iron dependent O2 sensor. Using computational modeling and protein engineering techniques, we identify a gas tunnel and critical residues therein that limit the flow of O2 to PHD2's catalytic core. We show that systematic modification of these residues can open the constriction topology of PHD2's gas tunnel. Using kinetic stopped-flow measurements with NO as a surrogate diatomic gas, we demonstrate up to 3.5-fold enhancement in its association rate to the iron center of tunnel-engineered mutants. Our most effectively designed mutant displays 9-fold enhanced catalytic efficiency (kcat/KM=830±40 M-1 s-1) in hydroxylating a peptide mimic of hypoxia inducible transcription factor HIF-1α, as compared to WT PHD2 (kcat/KM=90±9 M-1 s-1). Furthermore, transfection of plasmids that express designed PHD2 mutants in HEK-293T mammalian cells reveal significant reduction of HIF-1α and downstream hypoxia response transcripts under hypoxic conditions of 1 % O2. Overall, these studies highlight activation of PHD2 as a new pathway to reprogram hypoxia responses and HIF signaling in cells.

Abstract B085: The HIF-2 transcription factor mediates resistance to ferroptosis in pancreatic adenocarcinoma

Abstract Ferroptosis is an iron-dependent form of cell death converging on lipid peroxidation, first identified by examining compounds with enhanced lethality to KRAS mutant cells. Two compounds widely used as ferroptosis inducers are erastin, which acts as an inhibitor of cystine import and thereby depletes cellular glutathione, and RSL-3, which inhibits glutathione peroxidase 4. Despite over 90% of pancreatic adenocarcinoma (PDAC) tumors harboring KRAS mutations, PDAC exhibits relative resistance to ferroptosis compared to other tumor types, and the mechanisms behind this resistance remain unclear. The pancreatic tumor environment is marked by severe hypoxia, due to poor vascularity and a dense fibrotic stroma, and so we hypothesized that hypoxia might mediate resistance to ferroptosis. Here we report that across a panel of human and murine PDAC cell lines, hypoxia induces resistance to both ferroptosis inducing agents, erastin and RSL-3. The effect of hypoxia was synergistic with exposure to pancreatic cancer tumor interstitial fluid, which induces a hypoxic-like gene expression profile even under normoxia and enhances hypoxic gene expression changes when combined with hypoxia. Our findings reveal that hypoxia-mediated resistance to ferroptosis is orchestrated by the hypoxia-inducible transcription factor HIF-2, as evidenced by its complete abolition in HIF-2 knockout cell lines, while the protective effect of hypoxia was maintained in HIF-1 knockout lines. RNA-seq analyses and chromatin immunoprecipitation studies (chIP) identified several HIF-2 target genes responsible for coordinating ferroptosis resistance mechanisms. Cysteine is a precursor for glutathione, and HIF-2 increased cystine import through upregulation of both components of the cystine-glutamate antiporter system Xc (SLC7A11 and SLC3A2) and increased cysteine biosynthesis via upregulation of transsulfuration pathway enzymes (CBS and CTH). In addition, HIF-2 upregulated Parkin, which we identify as a novel regulator of ferroptosis, thereby increasing mitophagy and decreasing reactive oxygen species. In vivo studies using human and murine PDAC xenograft models confirmed that HIF-2 knockout impaired tumor growth and increased susceptibility to the pro-ferroptotic agent sulfasalazine. Collectively, these data identify a mechanism by which PDAC evades ferroptosis and identifies a possible therapeutic strategy of combining HIF-2 inhibitors with pro-ferroptotic agents in pancreatic cancer. Citation Format: Maimon E Hubbi, Catherine Wang, Erin Hollander, Yasir Suhail, Kshitiz, Alexander Muir, Ben Z Stanger, Chi V Dang. The HIF-2 transcription factor mediates resistance to ferroptosis in pancreatic adenocarcinoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B085.

HIF-2α-dependent induction of miR-29a restrains TH1 activity during T cell dependent colitis

Metabolic imbalance leading to inflammatory hypoxia and stabilization of hypoxia-inducible transcription factors (HIFs) is a hallmark of inflammatory bowel diseases. We hypothesize that HIF could be stabilized in CD4+ T cells during intestinal inflammation and alter the functional responses of T cells via regulation of microRNAs. Our assays reveal markedly increased T cell-intrinsic hypoxia and stabilization of HIF protein during experimental colitis. microRNA screen in primary CD4+ T cells points us towards miR-29a and our subsequent studies identify a selective role for HIF-2α in CD4-cell-intrinsic induction of miR-29a during hypoxia. Mice with T cell-intrinsic HIF-2α deletion display elevated T-bet (target of miR-29a) levels and exacerbated intestinal inflammation. Mice with miR-29a deficiency in T cells show enhanced intestinal inflammation. T cell-intrinsic overexpression of HIF-2α or delivery of miR-29a mimetic dampen TH1-driven colitis. In this work, we show a previously unrecognized function for hypoxia-dependent induction of miR-29a in attenuating TH1-mediated inflammation.

The transcription factor HIF-1α in NKp46+ ILCs limits chronic intestinal inflammation and fibrosis.

Innate lymphoid cells (ILCs) are critical for intestinal adaptation to microenvironmental challenges, and the gut mucosa is characterized by low oxygen. Adaptation to low oxygen is mediated by hypoxia-inducible transcription factors (HIFs), and the HIF-1α subunit shapes an ILC phenotype upon acute colitis that contributes to intestinal damage. However, the impact of HIF signaling in NKp46+ ILCs in the context of repetitive mucosal damage and chronic inflammation, as it typically occurs during inflammatory bowel disease, is unknown. In chronic colitis, mice lacking the HIF-1α isoform in NKp46+ ILCs show a decrease in NKp46+ ILC1s but a concomitant rise in neutrophils and Ly6Chigh macrophages. Single-nucleus RNA sequencing suggests enhanced interaction of mesenchymal cells with other cell compartments in the colon of HIF-1α KO mice and a loss of mucus-producing enterocytes and intestinal stem cells. This was, furthermore, associated with increased bone morphogenetic pathway-integrin signaling, expansion of fibroblast subsets, and intestinal fibrosis. In summary, this suggests that HIF-1α-mediated ILC1 activation, although detrimental upon acute colitis, protects against excessive inflammation and fibrosis during chronic intestinal damage.

Gut bacteria are essential for development of an invasive bark beetle by regulating glucose transport

Insects and their gut bacteria form a tight and beneficial relationship, especially in utilization of host nutrients. The red turpentine beetle (RTB), a destructive and invasive pine pest, employs mutualistic microbes to facilitate its invasion success. However, the molecular mechanism underlying the utilization of nutrients remains unknown. In this study, we found that gut bacteria are crucial for the utilization of D-glucose, a main carbon source for RTB development. Downstream assays revealed that gut bacteria-induced gut hypoxia and the secretion of riboflavin are responsible for RTB development by regulating D-glucose transport via the activation of a hypoxia-induced transcription factor 1 (Hif-1α). Further functional investigations confirmed that Hif-1α mediates glucose transport by direct upregulation of two glucose transporters (ST10 and ST27), thereby promoting RTB development. Our findings reveal how gut bacteria regulate the development of RTB, and promote our understanding of the mutualistic relationship of animals and their gut bacteria.

Hif-2α programs oxygen chemosensitivity in chromaffin cells.

The study of transcription factors that determine specialized neuronal functions has provided invaluable insights into the physiology of the nervous system. Peripheral chemoreceptors are neurone-like electrophysiologically excitable cells that link the oxygen concentration of arterial blood to the neuronal control of breathing. In the adult, this oxygen chemosensitivity is exemplified by type I cells of the carotid body, and recent work has revealed one isoform of the hypoxia-inducible transcription factor (HIF), HIF-2α, as having a nonredundant role in the development and function of that organ. Here, we show that activation of HIF-2α, including isolated overexpression of HIF-2α but not HIF-1α, is sufficient to induce oxygen chemosensitivity in adult adrenal medulla. This phenotypic change in the adrenal medulla was associated with retention of extra-adrenal paraganglioma-like tissues resembling the fetal organ of Zuckerkandl, which also manifests oxygen chemosensitivity. Acquisition of chemosensitivity was associated with changes in the adrenal medullary expression of gene classes that are ordinarily characteristic of the carotid body, including G protein regulators and atypical subunits of mitochondrial cytochrome oxidase. Overall, the findings suggest that, at least in certain tissues, HIF-2α acts as a phenotypic driver for cells that display oxygen chemosensitivity, thus linking 2 major oxygen-sensing systems.

Induction of the Mdm2 gene and protein by kinase signaling pathways is repressed by the pVHL tumor suppressor

The tumor suppressor von Hippel-Lindau, pVHL, is a multifaceted protein. One function is to dock to the hypoxia-inducible transcription factor (HIF) and recruit a larger protein complex that destabilizes HIF via ubiquitination, preventing angiogenesis and tumor development. pVHL also binds to the tumor suppressor p53 to activate specific p53 target genes. The oncogene Mdm2 impairs the formation of the p53-pVHL complex and activation of downstream genes by conjugating nedd8 to pVHL. While Mdm2 can impact p53 and pVHL, how pVHL may impact Mdm2 is unclear. Like p53 somatic mutations, point mutations are evident in pVHL that are common in renal clear cell carcinomas (RCC). In patients with RCC, Mdm2 levels are elevated, and we examined whether there was a relationship between Mdm2 and pVHL. TCGA and DepMap analysis revealed that mdm2 gene expression was elevated in RCC with vhl point mutations or copy number loss. In pVHL reconstituted or deleted isogenetically match RCC or MEF cell lines, Mdm2 was decreased in the presence of pVHL. Furthermore, through analysis using genetic and pharmacological approaches, we show that pVHL represses Mdm2 gene expression by blocking the MAPK-Ets signaling pathway and blocks Akt-mediated phosphorylation and stabilization of Mdm2. Mdm2 inhibition results in an increase in the p53-p21 pathway to impede cell growth. This finding shows how pVHL can indirectly impact the function of Mdm2 by regulating signaling pathways to restrict cell growth.

Bhlhe40 Regulates Proliferation and Angiogenesis in Mouse Embryoid Bodies under Hypoxia.

Knowledge of the molecular mechanisms that underlie the regulation of major adaptive responses to an unbalanced oxygen tension is central to understanding tissue homeostasis and disease. Hypoxia-inducible transcription factors (HIFs) coordinate changes in the transcriptome that control these adaptive responses. Here, we focused on the functional role of the transcriptional repressor basic-helix-loop-helix family member e40 (Bhlhe40), which we previously identified in a meta-analysis as one of the most consistently upregulated genes in response to hypoxia across various cell types. We investigated the role of Bhlhe40 in controlling proliferation and angiogenesis using a gene editing strategy in mouse embryonic stem cells (mESCs) that we differentiated in embryoid bodies (EBs). We observed that hypoxia-induced Bhlhe40 expression was compatible with the rapid proliferation of pluripotent mESCs under low oxygen tension. However, in EBs, hypoxia triggered a Bhlhe40-dependent cell cycle arrest in most progenitor cells and endothelial cells within vascular structures. Furthermore, Bhlhe40 knockout increased the basal vascularization of the EBs in normoxia and exacerbated the hypoxia-induced vascularization, supporting a novel role for Bhlhe40 as a negative regulator of blood vessel formation. Our findings implicate Bhlhe40 in mediating key functional adaptive responses to hypoxia, such as proliferation arrest and angiogenesis.

Transcriptional landscape of a hypoxia response identifies cell-specific pathways for adaptation.

How the HIF-1 (Hypoxia-Inducible) transcription factor drives and coordinates distinct responses to low oxygen across diverse cell types is poorly understood. We present a multi-tissue single-cell gene-expression atlas of the hypoxia response of the nematode Caenorhabditis elegans . This atlas highlights how cell-type-specific HIF-1 responses overlap and diverge among and within neuronal, intestinal, and muscle tissues. Using the atlas to guide functional analyses of candidate muscle-specific HIF-1 effectors, we discovered that HIF-1 activation drives downregulation of the tspo-1 ( TSPO, Translocator Protein) gene in vulval muscle cells to modulate a hypoxia-driven change in locomotion caused by contraction of body-wall muscle cells. We further showed that in human cardiomyocytes HIF-1 activation decreases levels of TSPO and thereby alters intracellular cholesterol transport and the mitochondrial network. We suggest that TSPO-1 is an evolutionarily conserved mediator of HIF-1-dependent modulation of muscle and conclude that our gene-expression atlas can help reveal how HIF-1 drives cell-specific adaptations to hypoxia.

Effects of Cinnamon on Cancer Prevention and Progression

Cinnamon has been used medicinally for centuries, but recently research has suggested it may have a role in cancer prevention and potentially treatment. The search for alternative and subjunctive therapies is essential due to the public demand and the increasing cost of healthcare. Here we review the biologically active components of cinnamon and discuss the methods of potential cinnamon activity against cancer including: transcription factor regulation and kinase activity. Nuclear Factor kappa B (NF-\(𝜿\)B) is a stress sensitive transcription factor that regulates transcription of genes involved in tumor progression and is inhibited by cinnamon components. Another way that cinnamon inhibits tumor growth is by suppression of transcription factor activator protein -1 (AP-1) which interacts with genes responsible for apoptosis, metastasis and inflammation. Hypoxia-inducible transcription factor-1 (HIF-1) and vascular endothelial growth factor (VEGF) are involved in angiogenesis, especially in the tumor microenvironment. The HIF-1-VEGF pathway are targeted by cinnamaldehyde, a compound found in cinnamon. Nuclear factor erythroid related factor 2 (Nrf2) is also examined and has been indicated to both potentially prevent cancer as well as induce it; various cinnamon derivatives target Nrf2. A cinnamaldehyde derivative has been implicated in a reduction of the mitogen-activated protein kinases (MAPKs), which are a group of kinases that regulate proliferation. Additionally, cinnamon components have been tied to cancer prevention by positively affecting the gut microbiome and inhibiting inflammation. The review concludes with a discussion of the future research needed and potential risk associated with cinnamon intake.

The special adaptation to hypoxia facilitated the expansion of the Asian house rat (Rattus tanezumi) into Tibet but not other Rattus species.

Rattus species are thought to live only at altitudes less than 2500 m, but the Asian house rat (R. tanezumi) (RT) has recently expanded to altitudes greater than 3500 m in China. Other Rattus species, especially brown rats (R. norvegicus) (RN), still reach only low altitudes on the Tibetan Plateau. Comparative genomics revealed the positive selection of hypoxia-inducible transcription factors 1 and 2 (HIFs) in RT, with the rapid evolution of HIF pathway genes in RT and Mus musculus (MM) but not RN or R. rattus. Population genomics revealed that genes associated with energy metabolism and oxygen transport were positively selected in RT compared with the other four Rattus species, and two specific substitutions (arginine 31 serine and leucine 33 methionine) were identified in the hemoglobin subunit beta (HBB) in RT. The above results suggested that RT possesses unique genetic adaptations to hypoxia, which was further confirmed by behavioral experiments on RT and RN. Normobaric hypoxia significantly reduced locomotion in RN but not in RT. Moreover, through intraspecific transcriptome analysis, the expression of Hbb and genes related to angiogenesis, oxygen transport, and glycolysis was upregulated, and the expression of genes associated with immunological functions in the liver, lungs, and/or sperm was downregulated in RT compared to those in RN. Interspecific transcriptome analysis further revealed that HIF-1α plays a role in modulating the hypoxic adaptation of RT rather than RN. Our work provides genomic, behavioral, and physiological insights into why RT, but not other Rattus species, could invade the Tibetan Plateau.

HIF3A gene disruption causes abnormal alveoli structure and early neonatal death.

Transcriptional response to changes in oxygen concentration is mainly controlled by hypoxia-inducible transcription factors (HIFs). Besides regulation of hypoxia-responsible gene expression, HIF-3α has recently been shown to be involved in lung development and in the metabolic process of fat tissue. However, the precise mechanism for such properties of HIF-3α is still largely unknown. To this end, we generated HIF3A gene-disrupted mice by means of genome editing technology to explore the pleiotropic role of HIF-3α in development and physiology. We obtained adult mice carrying homozygous HIF3A gene mutations with comparable body weight and height to wild-type mice. However, the number of litters and ratio of homozygous mutation carriers born from the mating between homozygous mutant mice was lower than expected due to sporadic deaths on postnatal day 1. HIF3A gene-disrupted mice exhibited abnormal configuration of the lung such as a reduced number of alveoli and thickened alveolar walls. Transcriptome analysis showed, as well as genes associated with lung development, an upregulation of stearoyl-Coenzyme A desaturase 1, a pivotal enzyme for fatty acid metabolism. Analysis of fatty acid composition in the lung employing gas chromatography indicated an elevation in palmitoleic acid and a reduction in oleic acid, suggesting an imbalance in distribution of fatty acid, a constituent of lung surfactant. Accordingly, administration of glucocorticoid injections during pregnancy resulted in a restoration of normal alveolar counts and a decrease in neonatal mortality. In conclusion, these observations provide novel insights into a pivotal role of HIF-3α in the preservation of critically important structure and function of alveoli beyond the regulation of hypoxia-mediated gene expression.

Hypoxia-Inducible Factor-Dependent and Independent Mechanisms Underlying Chemoresistance of Hypoxic Cancer Cells.

In hypoxic regions of malignant solid tumors, cancer cells acquire resistance to conventional therapies, such as chemotherapy and radiotherapy, causing poor prognosis in patients with cancer. It is widely recognized that some of the key genes behind this are hypoxia-inducible transcription factors, e.g., hypoxia-inducible factor 1 (HIF-1). Since HIF-1 activity is suppressed by two representative 2-oxoglutarate-dependent dioxygenases (2-OGDDs), PHDs (prolyl-4-hydroxylases), and FIH-1 (factor inhibiting hypoxia-inducible factor 1), the inactivation of 2-OGDD has been associated with cancer therapy resistance by the activation of HIF-1. Recent studies have also revealed the importance of hypoxia-responsive mechanisms independent of HIF-1 and its isoforms (collectively, HIFs). In this article, we collate the accumulated knowledge of HIF-1-dependent and independent mechanisms responsible for resistance of hypoxic cancer cells to anticancer drugs and briefly discuss the interplay between hypoxia responses, like EMT and UPR, and chemoresistance. In addition, we introduce a novel HIF-independent mechanism, which is epigenetically mediated by an acetylated histone reader protein, ATAD2, which we recently clarified.

The drug-specific properties of hypoxia-inducible factor-prolyl hydroxylase inhibitors in mice reveal a significant contribution of the kidney compared to the liver to erythropoietin induction

AimsKidney disease often leads to anemia due to a defect in the renal production of the erythroid growth factor erythropoietin (EPO), which is produced under the positive regulation of hypoxia-inducible transcription factors (HIFs). Chemical compounds that inhibit HIF-prolyl hydroxylases (HIF-PHs), which suppress HIFs, have been developed to reactivate renal EPO production in renal anemia patients. Currently, multiple HIF-PH inhibitors, in addition to conventional recombinant EPO reagents, are used for renal anemia treatment. This study aimed to elucidate the therapeutic mechanisms and drug-specific properties of HIF-PH inhibitors. Methods and key findingsGene expression analyses and mass spectrometry revealed that HIF-PH inhibitors (daprodustat, enarodustat, molidustat, and vadadustat) alter Epo gene expression levels in the kidney and liver in a drug-specific manner, with different pharmacokinetics in the plasma and urine after oral administration to mice. The drug specificity revealed the dominant contribution of EPO induction in the kidneys rather than in the liver to plasma EPO levels after HIF-PH inhibitor administration. We also found that several HIF-PH inhibitors directly induce duodenal gene expression related to iron intake, while these drugs indirectly suppress hepatic hepcidin expression to mobilize stored iron for hemoglobin synthesis through induction of the EPO-erythroferrone axis. SignificanceRenal EPO induction is the major target of HIF-PH inhibitors for their therapeutic effects on erythropoiesis. Additionally, the drug-specific properties of HIF-PH inhibitors in EPO induction and iron metabolism have been shown in mice, providing useful information for selecting the proper HIF-PH inhibitor for each renal anemia patient.