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Breast cancer stem cells (BCSCs) mitigate oxidative stress to maintain their viability and plasticity. However, the regulatory mechanism of oxidative stress in BCSCs remains unclear. We recently found that the histone reader ZMYND8 was upregulated in BCSCs. Here, we showed that ZMYND8 reduced ROS and iron to inhibit ferroptosis in aldehyde dehydrogenase-high ([ALDH.sup.hi]) BCSCs, leading to BCSC expansion and tumor initiation in mice. The underlying mechanism involved a two-fold posttranslational regulation of nuclear factor erythroid 2-related factor 2 (NRF2). ZMYND8 increased stability of NRF2 protein through KEAP1 silencing. On the other hand, ZMYND8 interacted with and recruited NRF2 to the promoters of antioxidant genes to enhance gene transcription in mammospheres. NRF2 phenocopied ZMYND8 to enhance BCSC stemness and tumor initiation by inhibiting ROS and ferroptosis. Loss of NRF2 counteracted ZMYND8's effects on antioxidant genes and ROS in mammospheres. Interestingly, ZMYND8 expression was directly controlled by NRF2 in mammospheres. Collectively, these findings uncover a positive feedback loop that amplifies the antioxidant defense mechanism sustaining BCSC survival and stemness., Introduction Breast cancer stem cells (BCSCs) are defined as a subpopulation of tumor cells with capability of self renewal and long-term maintenance of breast tumors and are responsible for breast [...]

SAP30 is a core subunit of the transcriptional corepressor SIN3 complex, but little is known about its role in gene regulation and human cancer. Here, we show that SAP30 was a nonmutational oncoprotein upregulated in more than 50% of human breast tumors and correlated with unfavorable outcomes in patients with breast cancer. In various breast cancer mouse models, we found that SAP30 promoted tumor growth and metastasis through its interaction with SIN3A/3B. Surprisingly, the canonical gene silencing role was not essential for SAP30's tumor-promoting actions. SAP30 enhanced chromatin accessibility and RNA polymerase II occupancy at promoters in breast cancer cells, acting as a coactivator for genes involved in cell motility, angiogenesis, and lymphangiogenesis, thereby driving tumor progression. Notably, SAP30 formed a homodimer with 1 subunit binding to SIN3A and another subunit recruiting MLL1 through specific Phe186/200 residues within its transactivation domain. MLL1 was required for SAP30-mediated transcriptional coactivation and breast tumor progression. Collectively, our findings reveal that SAP30 represents a transcriptional dependency in breast cancer., Introduction Breast cancer is the most common malignant disease in women, with many patients dying of metastasis. The development of breast cancer toward distant metastasis is a harsh process, wherein [...]

Altered epigenetic reprogramming contributes to breast cancer progression and metastasis. How the epigenetic reader mediates breast cancer progression remains poorly understood. Here, we showed that the epigenetic reader zinc finger MYND-type containing 8 (ZMYND8) is induced by HIF-1 and HIF-2 in breast cancer cells and also upregulated in human breast tumors, and is correlated with poor survival of patients with breast cancer. Genetic deletion of ZMYND8 decreases breast cancer cell colony formation, migration, and invasion in vitro, and inhibits breast tumor growth and metastasis to the lungs in mice. The ZMYND8's oncogenic effect in breast cancer requires HIF-1 and HIF-2. We further showed that ZMYND8 interacts with HIF-1[alpha] and HIF-2[alpha] and enhances elongation of the global HIF-induced oncogenic genes by increasing recruitment of BRD4 and subsequent release of paused RNA polymerase II in breast cancer cells. ZMYND8 acetylation at lysines 1007 and 1034 by p300 is required for HIF activation and breast cancer progression and metastasis. These findings uncover a primary epigenetic mechanism of HIF activation and HIF-mediated breast cancer progression, and discover a possible molecular target for the diagnosis and treatment of breast cancer., Introduction Metastasis is a major clinical obstacle to curative cancer therapy and is responsible for about 90% of breast cancer-related deaths in women. The development of a benign and nonmalignant [...]

Hypoxia-inducible factor (HIF) directly activates the transcription of metabolic enzymes in response to hypoxia to reprogram cellular metabolism required for tumor cell proliferation. Through analyzing glutamate-linked aminotransferases, we here identified glutamate pyruvate transaminase 2 (GPT2) as a direct HIF-2 target gene in human glioblastoma (GBM). Hypoxia upregulated GPT2 mRNA and protein levels in GBM cells, which required HIF-2 but not HIF-1. HIF-2 directly bound to the hypoxia response element of the human GPT2 gene, leading to its transcription in hypoxic GBM cells. GPT2 located at the nucleus and mitochondria and reduced α-ketoglutarate levels in GBM cells. Genetic or pharmacological inhibition of GPT2 decreased GBM cell growth and migration under normoxia and hypoxia. Knockout of GPT2 inhibited GBM tumor growth in mice. Collectively, these findings uncover a hypoxia-inducible aminotransferase GPT2 required for GBM progression. [ABSTRACT FROM AUTHOR]

Metastasis involves critical interactions between cancer and stromal cells. Intratumoral hypoxia promotes metastasis through activation of hypoxia-inducible factors (HIFs). We demonstrate that HIFs mediate paracrine signaling between breast cancer cells [...]

Cell death is a key feature of neurological diseases, including stroke and neurodegenerative disorders. Studies in a variety of ischemic/hypoxic mouse models demonstrate that poly(ADP‐ribose) polymerase 1 (PARP‐1)‐dependent cell death, also named PARthanatos, plays a pivotal role in ischemic neuronal cell death and disease progress. PARthanatos has its unique triggers, processors, and executors that convey a highly orchestrated and programmed signaling cascade. In addition to its role in gene transcription, DNA damage repair, and energy homeostasis through PARylation of its various targets, PARP‐1 activation in neuron and glia attributes to brain damage following ischemia/reperfusion. Pharmacological inhibition or genetic deletion of PARP‐1 reduces infarct volume, eliminates inflammation, and improves recovery of neurological functions in stroke. Here, we reviewed the role of PARP‐1 and PARthanatos in stroke and their therapeutic potential. [ABSTRACT FROM AUTHOR]

Increased catalytic activity of CBS (cystathionine ß-synthase) was recently shown to mediate vasodilation of the cerebral microcirculation, which is initiated within minutes of the onset of acute hypoxia. To test whether chronic hypoxia was a stimulus for increased CBS expression, U87-MG human glioblastoma and PC12 rat phaeochromocytoma cells were exposed to 1% or 20%O2 for 24-72 h. CBS mRNA and protein expression were increased in hypoxic cells. Hypoxic induction of CBS expression was abrogated in cells transfected with vector encoding shRNA targeting HIF (hypoxia-inducible factor) 1α or 2α. Exposure of rats to hypobaric hypoxia (0.35 atm; 1 atm = 101.325 kPa) for 3 days induced increased CBS mRNA, protein and catalytic activity in the cerebral cortex and cerebellum, which was blocked by administration of the HIF inhibitor digoxin. HIF-binding sites, located 0.8 and 1.2 kb 5 to the transcription start site of the human CBS and rat Cbs genes respectively, were identified by ChIP assays. A 49-bp human sequence, which encompassed an inverted repeat of the core HIF-binding site, functioned as a hypoxiaresponse element in luciferase reporter transcription assays. Thus HIFsmediate tissue-specificCBSexpression, whichmay augment cerebral vasodilation as an adaptive response to chronic hypoxia. [ABSTRACT FROM AUTHOR]

Sleep-disordered breathing with recurrent apnea is associated with intermittent hypoxia (IH). Cardiovascular morbidities caused by IH are triggered by increased generation of reactive oxygen species (ROS) by pro-oxidant enzymes, especially NADPH oxidase-2 (Nox2). Previous studies showed that (i) IH activates hypoxia-inducible factor 1 (HIF-1) in a ROS-dependent manner and (ii) HIF-1 is required for IH-induced ROS generation, indicating the existence of a feed-forward mechanism. In the present study, using multiple pharmacological and genetic approaches, we investigated whether IH-induced expression of Nox2 is mediated by HIF-1 in the central and peripheral nervous system of mice as well as in cultured cells. IH increased Nox2 mRNA, protein, and enzyme activity in PC12 pheochromocytoma cells as well as in wild-type mouse embryonic fibroblasts (MEFs). This effect was abolished or attenuated by blocking HIF-1 activity through RNA interference or pharmacologic inhibition (digoxin or YC-1) or by genetic knockout of HIF-1α in MEFs. Increasing HIF-1α expression by treating PC 12 cells with the iron chelator deferoxamine for 20 h or by transfecting them with HIF-1alpha expression vector increased Nox2 expression and enzyme activity. Exposure of wild-type mice to IH (8 h/day for 10 days) up-regulated Nox2 mRNA expression in brain cortex, brain stem, and carotid body but not in cerebellum. IH did not induce Nox2 expression in cortex, brainstem, carotid body, or cerebellum of Hif1a+/− mice, which do not manifest increased ROS or cardiovascular morbidities in response to IH. These results establish a pathogenic mechanism linking HIF-1, ROS generation, and cardiovascular pathology in response to IH. J. Cell. Physiol. 226: 2925-2933, 2011. © 2011 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Summary: MCM proteins are components of a DNA helicase that plays an essential role in DNA replication and cell proliferation. However, MCM proteins are present in excess relative to origins of replication, suggesting they may serve other functions. Decreased proliferation is a fundamental physiological response to hypoxia in many cell types, and hypoxia-inducible factor 1 (HIF-1) has been implicated in this process. Here, we demonstrate that multiple MCM proteins bind directly to the HIF-1α subunit and synergistically inhibit HIF-1 transcriptional activity via distinct O2-dependent mechanisms. MCM3 inhibits transactivation domain function, whereas MCM7 enhances HIF-1α ubiquitination and proteasomal degradation. HIF-1 activity decreases when quiescent cells re-enter the cell cycle, and this effect is MCM dependent. Exposure to hypoxia leads to MCM2–7 downregulation in diverse cell types. These studies reveal a function of MCM proteins apart from their DNA helicase activity and establish a direct link between HIF-1 and the cell-cycle machinery. [Copyright &y& Elsevier]

Summary: The pyruvate kinase isoforms PKM1 and PKM2 are alternatively spliced products of the PKM2 gene. PKM2, but not PKM1, alters glucose metabolism in cancer cells and contributes to tumorigenesis by mechanisms that are not explained by its known biochemical activity. We show that PKM2 gene transcription is activated by hypoxia-inducible factor 1 (HIF-1). PKM2 interacts directly with the HIF-1α subunit and promotes transactivation of HIF-1 target genes by enhancing HIF-1 binding and p300 recruitment to hypoxia response elements, whereas PKM1 fails to regulate HIF-1 activity. Interaction of PKM2 with prolyl hydroxylase 3 (PHD3) enhances PKM2 binding to HIF-1α and PKM2 coactivator function. Mass spectrometry and anti-hydroxyproline antibody assays demonstrate PKM2 hydroxylation on proline-403/408. PHD3 knockdown inhibits PKM2 coactivator function, reduces glucose uptake and lactate production, and increases O2 consumption in cancer cells. Thus, PKM2 participates in a positive feedback loop that promotes HIF-1 transactivation and reprograms glucose metabolism in cancer cells. [ABSTRACT FROM AUTHOR]

Fengycin, as a lipopeptide produced by Bacillus subtilis, displays potent activity against filamentous fungi, including Aspergillus flavus and Soft-rot fungus, which exhibits a wide range of potential applications in food industries, agriculture, and medicine. To better clarify the regulatory mechanism of fructose on fengycin biosynthesis, the iTRAQ-based proteomic analysis was utilized to investigate the differentially expressed proteins of B. amyloliquefaciens fmb-60 cultivated in ML (without fructose) and MLF (with fructose) medium. The results indicated that a total of 811 proteins, including 248 proteins with differential expression levels (162 which were upregulated (fold > 2) and 86, which were downregulated (fold < 0.5) were detected, and most of the proteins are associated with cellular metabolism, biosynthesis, and biological regulation process. Moreover, the target genes' relative expression was conducted using quantitative real-time PCR to validate the proteomic analysis results. Based on the results of proteome analysis, the supposed pathways of fructose enhancing fengycin biosynthesis in B. amyloliquefaciens fmb-60 can be summarized as improvement of the metabolic process, including cellular amino acid and amide, fatty acid biosynthesis, peptide and protein, nucleotide and nucleobase-containing compound, drug/toxin, cofactor, and vitamin; reinforcement of peptide/protein translation, modification, biological process, and response to a stimulus. In conclusion, this study represents a comprehensive and systematic investigation of the fructose mechanism on improving fengycin biosynthesis in B. amyloliquefaciens, which will provide a road map to facilitate the potential application of fengycin or its homolog in defending against filamentous fungi. [ABSTRACT FROM AUTHOR]

Abstract: Protease-activated receptors (PARs) are G protein-coupled receptors that regulate the cellular response to extracellular serine proteases, like thrombin, trypsin, and tryptase. The PAR family consists of four members: PAR-1, -3, and -4 as thrombin receptors and PAR-2 as the trypsin/tryptase receptor, which are abundantly expressed in the brain throughout development. Recent evidence has supported the direct involvement of PARs in brain development and function. The expression of PARs in the brain is differentially upregulated or downregulated under pathological conditions in neurodegenerative disorders, like Parkinson''s disease, Alzheimer''s disease, multiple sclerosis, stroke, and human immunodeficiency virus-associated dementia. Activation of PARs mediates cell death or cell survival in the brain, depending on the amplitude and the duration of agonist stimulation. Interference or potentiation of PAR activation is beneficial in animal models of neurodegenerative diseases. Therefore, PARs mediate either neurodegeneration or neuroprotection in neurodegenerative diseases and represent attractive therapeutic targets for treatment of brain injuries. Here, we review the abnormal expression of PARs in the brain under pathological conditions, the functions of PARs in neurodegenerative disorders, and the molecular mechanisms involved. [Copyright &y& Elsevier]

Abstract: Protease-activated receptors (PARs), G-protein-coupled receptors, are widely expressed in various tissues, where they participate in physiological and pathological processes, such as hemostasis, proliferation, tissue repair, and inflammation. Recently, we found that PARs were upregulated in the rat retina following optic nerve crush injury. However, the role of PAR in retinal ganglion cells following optic nerve crush still remains unknown. Here, we studied PAR-mediated calcium signaling in retinal ganglion cells, RGC-5. Using reverse transcription-polymerase chain reaction, we demonstrate that RGC-5 cells mainly express PAR-1 and to a lower extent PAR-2, which was further confirmed by indirect immunofluorescence. Short-term stimulation of RGC-5 cells with thrombin (0.001–1 U/ml) and trypsin (1–100 nM) concentration-dependently induced a transient increase in intracellular calcium concentration ([Ca2+]i). An increase in [Ca2+]i was also induced by both TRag (PAR-1 activating peptide) and PAR-2 activating peptide (PAR-2 AP). The EC50 values were 0.3 nM for thrombin, 12.0 nM for trypsin, 1.3 μM for TRag, and 1.6 μM for PAR-2 AP, respectively. Desensitization was studied using two successive pulses of agonists. The thrombin-induced calcium response was significantly reduced by PAR-1 desensitization caused by pre-challenging RGC-5 cells with thrombin or TRag, but not by PAR-2 desensitization. On the other hand, pretreatment with trypsin, TRag or PAR-2 AP desensitized the cells since the calcium response to a second exposure to trypsin was significantly reduced. Calcium source studies revealed that PAR-induced [Ca2+]i rise mainly comes from intracellular stores in RGC-5 cells. Thus, we demonstrate that PAR-1 and PAR-2 are functionally expressed in retinal ganglion cells, mediating calcium mobilization mainly from intracellular stores. [Copyright &y& Elsevier]

Poly(ADP-ribose) (PAR) polymerase-1 (PARP-1) acts as a DNA damage sensor. It recognizes DNA damage and facilitates DNA repair by recruiting DNA repair machinery to damage sites. Recent studies reported that PARP-1 also plays an important role in DNA replication by recognizing the unligated Okazaki fragments and controlling the speed of fork elongation. On the other hand, emerging evidence reveals that excessive activation of PARP-1 causes chromatin DNA fragmentation and triggers an intrinsic PARP-1-dependent cell death program designated parthanatos, which can be blocked by genetic deletion or pharmacological inhibition of PARP-1. Therefore, PARP-1 plays an essential role in maintaining genomic stability by either facilitating DNA repair/replication or triggering DNA fragmentation to kill cells. A group of structure-specific nucleases is crucial for executing DNA incision and fragmentation following PARP-1 activation. In this review, we will discuss how PARP-1 coordinates with its associated nucleases to maintain genomic integrity and control the decision of cell life and death. [ABSTRACT FROM AUTHOR]

Epigenetic regulator plays a pivotal role in breast cancer progression. We recently demonstrated that the epigenetic reader zinc finger MYND-type containing 8 (ZMYND8) mediates breast cancer progression and metastasis by activating hypoxia-inducible factor (HIF). This discovery provides new insights into the molecular mechanism underlying HIF activation and breast cancer progression. [ABSTRACT FROM AUTHOR]

Abstract Atherosclerosis is a complicated process comprising inflammation, accumulation of collagen matrix and aberrant DNA methylation. SMAD7 is known to play an important role in fibrosis and inflammation. In recent years, increasing research has concentrated on the connection between DNA methylation and atherosclerosis. The current study was designed to investigate methylation status of some specific gene with a focus on SMAD7 in atherosclerosis and elucidate their relationship. We found that SMAD7 expression was decreased and its promoter region was markedly methylated in atherosclerotic plaques when compared with normal artery walls. Using MALDI-TOF MS, increased DNA methylation levels of SMAD7 promoter at CpG unit 5.8.15.16 were found in peripheral blood of atherosclerosis patients relative to matched normal controls, respectively. Correlation analysis revealed that mean DNA methylation levels of SMAD7 promoter of CpG unit 5.8.15.16 were positively associated with homocysteine levels (r = 0.724, p <.001) and carotid plaque scores(r = 0.790, p <.001). SMAD7 promoter is hyper-methylated both in human atherosclerotic plaques and atherosclerosis patients, which is positively associated with homocysteine levels and carotid plaque scores. Thus, methylated SMAD7 may be a novel predicted marker and therapeutics target for atherosclerosis. Highlights • MALDI-TOF MS is an advantageous way to analyze DNA methylation. • SMAD7 promoter is hyper-methylated in human atherosclerotic plaques and peripheral blood of atherosclerosis patients. • SMAD7 methylation levels positively correlate with homocysteine levels and carotid plaque scores. • Methylated SMAD7 may be a novel predicted marker and therapeutics target for atherosclerosis. [ABSTRACT FROM AUTHOR]

Summary: T cell differentiation into distinct functional effector and inhibitory subsets is regulated, in part, by the cytokine environment present at the time of antigen recognition. Here, we show that hypoxia-inducible factor 1 (HIF-1), a key metabolic sensor, regulates the balance between regulatory T cell (Treg) and TH17 differentiation. HIF-1 enhances TH17 development through direct transcriptional activation of RORγt and via tertiary complex formation with RORγt and p300 recruitment to the IL-17 promoter, thereby regulating TH17 signature genes. Concurrently, HIF-1 attenuates Treg development by binding Foxp3 and targeting it for proteasomal degradation. Importantly, this regulation occurs under both normoxic and hypoxic conditions. Mice with HIF-1α-deficient T cells are resistant to induction of TH17-dependent experimental autoimmune encephalitis associated with diminished TH17 and increased Treg cells. These findings highlight the importance of metabolic cues in T cell fate determination and suggest that metabolic modulation could ameliorate certain T cell-based immune pathologies. [ABSTRACT FROM AUTHOR]

Traumatic brain injury (TBI), often induced by sports, car accidents, falls, or other daily occurrences, is a primary non-genetically related risk factor for the development of subsequent neurodegeneration and neuronal cell death. However, the molecular mechanisms underlying neurodegeneration, cell death, and neurobehavioral dysfunction following TBI remain unclear. Here, we found that poly(ADP-ribose) polymerase-1 (PARP-1) was hyperactivated following TBI and its inhibition reduced TBI-induced brain injury. Macrophage migration inhibitory factor (MIF), a newly identified nuclease involved in PARP-1-dependent cell death, was translocated from the cytosol to the nucleus in cortical neurons following TBI and promoted neuronal cell death in vivo. Genetic deletion of MIF protected neurons from TBI-induced dendritic spine loss, morphological complexity degeneration, and subsequent neuronal cell death in mice. Moreover, MIF knockout reduced the brain injury volume and improved long-term animal behavioral rehabilitation. These neuroprotective effects in MIF knockout mice were reversed by the expression of wild-type MIF but not nuclease-deficient MIF mutant. In contrast, genetic deletion of MIF did not alter TBI-induced neuroinflammation. These findings reveal that MIF mediates TBI-induced neurodegeneration, neuronal cell death and neurobehavioral dysfunction through its nuclease activity, but not its pro-inflammatory role. Targeting MIF’s nuclease activity may offer a novel strategy to protect neurons from TBI. [ABSTRACT FROM AUTHOR]

27-Hydroxycholesterol (27-HC) is the most abundant oxysterol that increases the risk of breast cancer progression. However, little is known about epigenetic regulation of 27-HC metabolism and its role in breast tumor initiation. Using genetic mouse mammary tumor and human breast cancer models, we showed here that the histone reader ZMYND8 was selectively expressed in breast cancer stem cells (BCSCs) and promoted epithelial-mesenchymal transition (EMT), BCSC maintenance and self-renewal, and oncogenic transformation through its epigenetic functions, leading to breast tumor initiation. Mechanistically, ZMYND8 was a master transcriptional regulator of 27-HC metabolism. It increased cholesterol biosynthesis and oxidation but blocked cholesterol efflux and 27-HC catabolism, leading to accumulation of 27-HC in BCSCs. Consequently, 27-HC promoted EMT, oncogenic transformation, and tumor initiation through activation of liver X receptor. These findings reveal that ZMYND8 is an epigenetic booster that drives breast tumor initiation through metabolic reprogramming.

Branched-chain amino acid transaminase 1 (BCAT1) is upregulated selectively in human isocitrate dehydrogenase (IDH) wildtype (WT) but not mutant glioblastoma multiforme (GBM) and promotes IDHWT GBM growth. Through a metabolic synthetic lethal screen, we report here that α-ketoglutarate (AKG) kills IDHWT GBM cells when BCAT1 protein is lost, which is reversed by reexpression of BCAT1 or supplementation with branched-chain α-ketoacids (BCKA), downstream metabolic products of BCAT1. In patient-derived IDHWT GBM tumors in vitro and in vivo, cotreatment of BCAT1 inhibitor gabapentin and AKG resulted in synthetic lethality. However, AKG failed to evoke a synthetic lethal effect with loss of BCAT2, BCKDHA, or GPT2 in IDHWT GBM cells. Mechanistically, loss of BCAT1 increased the NAD+/NADH ratio but impaired oxidative phosphorylation, mTORC1 activity, and nucleotide biosynthesis. These metabolic alterations were synergistically augmented by AKG treatment, thereby causing mitochondrial dysfunction and depletion of cellular building blocks, including ATP, nucleotides, and proteins. Partial restoration of ATP, nucleotides, proteins, and mTORC1 activity by BCKA supplementation prevented IDHWT GBM cell death conferred by the combination of BCAT1 loss and AKG. These findings define a targetable metabolic vulnerability in the most common subset of GBM that is currently incurable., Significance: Metabolic synthetic lethal screening in IDHWT glioblastoma defines a vulnerability to ΑΚG following BCAT1 loss, uncovering a therapeutic strategy to improve glioblastoma treatment. See related commentary by Meurs and Nagrath, p. 2354., (©2022 American Association for Cancer Research.)