Mingjiang Zhu

The Lysosomal v-ATPase-Ragulator Complex is a Common Activator for AMPK and mTORC1, Acting as a Switch between Catabolism and Anabolism

AMPK and mTOR play principal roles in governing metabolic programs; however, mechanisms underlying the coordination of the two inversely regulated kinases remain unclear. In this study we found, most surprisingly, that the late endosomal/lysosomal protein complex v-ATPase-Ragulator, essential for activation of mTORC1, is also required for AMPK activation. We also uncovered that AMPK is a residential protein of late endosome/lysosome. Under glucose starvation, the v-ATPase-Ragulator complex is accessible to AXIN/LKB1 for AMPK activation. Concurrently, the guanine nucleotide exchange factor (GEF) activity of Ragulator toward RAG is inhibited by AXIN, causing dissociation from endosome and inactivation of mTORC1. We have thus revealed that the v-ATPase-Ragulator complex is also an initiating sensor for energy stress and meanwhile serves as an endosomal docking site for LKB1-mediated AMPK activation by forming the v-ATPase-Ragulator-AXIN/LKB1-AMPK complex, thereby providing a switch between catabolism and anabolism. Our current study also emphasizes a general role of late endosome/lysosome in controlling metabolic programs.

Free radical oxidation of cardiolipin: chemical mechanisms, detection and implication in apoptosis, mitochondrial dysfunction and human diseases

Abstract Cardiolipin (CL) is a mitochondria-specific phospholipid and is critical for maintaining the integrity of mitochondrial membrane and mitochondrial function. CL also plays an active role in mitochondria-dependent apoptosis by interacting with cytochrome c (cyt c), tBid and other important Bcl-2 proteins. The unique structure of CL with four linoleic acid side chains in the same molecule and its cellular location make it extremely susceptible to free radical oxidation by reactive oxygen species including free radicals derived from peroxidase activity of cyt c/CL complex, singlet oxygen and hydroxyl radical. The free radical oxidation products of CL have been emerged as important mediators in apoptosis. In this review, we summarize the free radical chemical mechanisms that lead to CL oxidation, recent development in detection of oxidation products of CL by mass spectrometry and the implication of CL oxidation in mitochondria-mediated apoptosis, mitochondrial dysfunction and human diseases.

Fructose-1,6-bisphosphate and aldolase mediate glucose sensing by AMPK.

The major energy source for most cells is glucose, from which ATP is generated via glycolysis and/or oxidative metabolism. Glucose deprivation activates AMP-activated protein kinase (AMPK), but it is unclear whether this activation occurs solely via changes in AMP or ADP, the classical activators of AMPK. Here, we describe an AMP/ADP-independent mechanism that triggers AMPK activation by sensing the absence of fructose-1,6-bisphosphate (FBP), with AMPK being progressively activated as extracellular glucose and intracellular FBP decrease. When unoccupied by FBP, aldolases promote the formation of a lysosomal complex containing at least v-ATPase, ragulator, axin, liver kinase B1 (LKB1) and AMPK, which has previously been shown to be required for AMPK activation. Knockdown of aldolases activates AMPK even in cells with abundant glucose, whereas the catalysis-defective D34S aldolase mutant, which still binds FBP, blocks AMPK activation. Cell-free reconstitution assays show that addition of FBP disrupts the association of axin and LKB1 with v-ATPase and ragulator. Importantly, in some cell types AMP/ATP and ADP/ATP ratios remain unchanged during acute glucose starvation, and intact AMP-binding sites on AMPK are not required for AMPK activation. These results establish that aldolase, as well as being a glycolytic enzyme, is a sensor of glucose availability that regulates AMPK.

Mass spectrometry-based metabolomic profiling identifies alterations in salivary redox status and fatty acid metabolism in response to inflammation and oxidative stress in periodontal disease

Periodontal diseases represent the most common chronic inflammatory diseases in humans and a major cause of tooth loss. Combining mass spectrometry-based ionomics and targeted lipidomics on fatty acid metabolites, we identified significant alterations in redox status and fatty acid metabolism in saliva in response to chronic inflammation and oxidative stress in periodontal disease in a cohort of nonsmoker subjects with chronic periodontitis. For the first time, ionomic profiling of around 30 ions in saliva revealed significantly decreased levels of redox-active metal ions including Mn, Cu, and Zn in the periodontal group, which is consistent with decreased levels of superoxide dismutases in saliva and serum. A targeted lipidomic approach was employed to monitor the major metabolites of arachidonic acid and linoleic acid in saliva. We observed increased levels of cyclooxygenase products including PGE2, PGD2, and PGF2α and TXB2, but decreased level of PGI2 in the periodontal group. A unique pattern of the lipoxygenase products of arachidonic acid and linoleic acid was observed with increased level of 5-HETE but decreased levels of 13-HODE and 9-HODE. Levels of salivary F2-isoprostanes, free radical lipid peroxidation products, and a gold standard for oxidative stress in vivo were also significantly elevated. Taking these data together, our study using multiple powerful omics techniques demonstrates that local redox alteration contributes significantly to periodontitis through the modulation of fatty acid metabolism in response to inflammation and oxidative stress. This study highlights the importance of redox status in periodontitis and provides a rationale for preventing periodontal disease by dietary interventions aiming to restore redox balance.

Role of mitochondria in programmed cell death mediated by arachidonic acid-derived eicosanoids

Arachidonic acid-derived eicosanoids from cyclooxygenases, lipoxygenases, and cytochrome P450 are important lipid mediators involved in numerous homeostatic and pathophysiological processes. Most eicosanoids act primarily on their respective cell surface G-protein coupled receptors to elicit downstream signaling in an autocrine and paracrine fashion. Emerging evidence indicates that these hormones are also critical in apoptosis in a cell/tissue specific manner. In this review, we summarize the formation of eicosanoids and their roles as mediators in apoptosis, specifically on the roles of mitochondria in mediating these events and the signaling pathways involved. The biological relevance of eicosanoid-mediated apoptosis is also discussed.

Mitochondrial control of apoptosis through modulation of cardiolipin oxidation in hepatocellular carcinoma: A novel link between oxidative stress and cancer

Abstract Altered redox status in cancer cells has been linked to lipid peroxidation induced by reactive oxygen species (ROS) and subsequent formation of reactive lipid electrophiles, especially 4‐hydroxy‐nonenal (4‐HNE). Emerging evidence suggests that cancer cells manipulate redox status to acquire anti‐apoptotic phenotype but the underlying mechanisms are poorly understood. Cardiolipin (CL), a mitochondria‐specific inner membrane phospholipid, is critical for maintaining mitochondrial function. Paradoxically, liver tissues contain tetralinoleoyl cardiolipin (TLCL) as the major CL in mitochondria yet emerging evidence suggests that ROS generated in mitochondria may lead to CL peroxidation and activation of intrinsic apoptosis. It remains unclear how CL oxidation leads to apoptosis and its relevance to the pathogenesis of hepatocellular carcinoma (HCC). We employed a mass spectrometry‐based lipidomic approach to profile lipids in human tissues of HCC and found that CL was gradually decreased in tumor comparing to peripheral non‐cancerous tissues, accompanied by a concomitant decrease of oxidized CL and its oxidation product, 4‐HNE. Incubation of liver cancer cells with TLCL significantly restored apoptotic sensitivity accompanied by an increase of CL and its oxidation products when treated with staurosporine (STS) or Sorafenib (the standard treatment for late stage HCC patients). Our studies uncovered a novel mechanism by which cancer cells adopt to evade apoptosis, highlighting the importance of mitochondrial control of apoptosis through modulation of CL oxidation and subsequent 4‐HNE formation in HCC. Thus manipulation of mitochondrial CL oxidation and lipid electrophile formation may have potential therapeutic value for diseases linked to oxidative stress and mitochondrial dysfunctions. Graphical abstract Figure. No Caption available. HighlightsLipidomics identified distinct patterns of mitochondrial CL in human HCC tumor.Tumor tissue has more diverse fatty acids especially saturated fatty acids in CL.Stage‐dependent decrease of 4‐HNE protein adducts and CL oxidation in HCC tumor.Incubation of liver cancer cells with TLCL restored apoptotic sensitivity.

Formation of electrophilic oxidation products from mitochondrial cardiolipin in vitro and in vivo in the context of apoptosis and atherosclerosis.

Emerging evidence indicates that mitochondrial cardiolipins (CL) are prone to free radical oxidation and this process appears to be intimately associated with multiple biological functions of mitochondria. Our previous work demonstrated that a significant amount of potent lipid electrophiles including 4-hydroxy-nonenal (4-HNE) was generated from CL oxidation through a novel chemical mechanism. Here we provide further evidence that a characteristic class of CL oxidation products, epoxyalcohol-aldehyde-CL (EAA-CL), is formed through this novel mechanism in isolated mice liver mitochondria when treated with the pro-apoptotic protein t-Bid to induce cyt c release. Generation of these oxidation products are dose-dependently attenuated by a peroxidase inhibitor acetaminophen (ApAP). Using a mouse model of atherosclerosis, we detected significant amount of these CL oxidation products in liver tissue of low density lipoprotein receptor knockout (LDLR −/−) mice after Western diet feeding. Our studies highlight the importance of lipid electrophiles formation from CL oxidation in the settings of apoptosis and atherosclerosis as inhibition of CL oxidation and lipid electrophiles formation may have potential therapeutic value in diseases linked to oxidant stress and mitochondrial dysfunctions.

T-Cell Mineralocorticoid Receptor Controls Blood Pressure by Regulating Interferon-GammaNovelty and Significance

Rationale: Hypertension remains to be a global public health burden and demands novel intervention strategies such as targeting T cells and T-cell–derived cytokines. Mineralocorticoid receptor (MR) antagonists have been clinically used to treat hypertension. However, the function of T-cell MR in blood pressure (BP) regulation has not been elucidated. Objective: We aim to determine the role of T-cell MR in BP regulation and to explore the mechanism. Methods and Results: Using T-cell MR knockout mouse in combination with angiotensin II–induced hypertensive mouse model, we demonstrated that MR deficiency in T cells strikingly decreased both systolic and diastolic BP and attenuated renal and vascular damage. Flow cytometric analysis showed that T-cell MR knockout mitigated angiotensin II–induced accumulation of interferon-gamma (IFN-&ggr;)–producing T cells, particularly CD8+ population, in both kidneys and aortas. Similarly, eplerenone attenuated angiotensin II–induced elevation of BP and accumulation of IFN-&ggr;–producing T cells in wild-type mice. In cultured CD8+ T cells, T-cell MR knockout suppressed IFN-&ggr; expression whereas T-cell MR overexpression and aldosterone both enhanced IFN-&ggr; expression. At the molecular level, MR interacted with NFAT1 (nuclear factor of activated T-cells 1) and activator protein-1 in T cells. Finally, T-cell MR overexpressing mice manifested more elevated BP compared with control mice after angiotensin II infusion and such difference was abolished by IFN-&ggr;–neutralizing antibodies. Conclusions: MR may interact with NFAT1 and activator protein-1 to control IFN-&ggr; in T cells and to regulate target organ damage and ultimately BP. Targeting MR in T cells specifically may be an effective novel approach for hypertension treatment.

Identification of a novel series of anti-inflammatory and anti-oxidative phospholipid oxidation products containing the cyclopentenone moietyin vitroandin vivo: Implication in atherosclerosis

Oxidative stress and inflammation are two major contributing factors to atherosclerosis, a leading cause of cardiovascular disease. Oxidation of phospholipids on the surface of low density lipoprotein (LDL) particles generated under oxidative stress has been associated with the progression of atherosclerosis, but the underlying molecular mechanisms remain poorly defined. We identified a novel series of oxidation products containing the cyclopentenone moiety, termed deoxy-A2/J2-isoprostanes-phosphocholine, from 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine in vivo using mass spectrometry and by comparison to a chemically synthesized standard. Transcriptomic analysis (RNA-seq) demonstrated that these compounds affected >200 genes in bone marrow-derived macrophages, and genes associated with inflammatory and anti-oxidative responses are among the top 5 differentially expressed. To further investigate the biological relevance of these novel oxidized phospholipids in atherosclerosis, we chemically synthesized a representative compound 1-palmitoyl-2–15-deoxy-δ-12,14-prostaglandin J2-sn-glycero-3-phosphocholine (15d-PGJ2-PC) and found that it induced anti-inflammatory and anti-oxidant responses in macrophages through modulation of NF-κB, peroxisome proliferator-activated receptor γ (PPARγ), and Nrf2 pathways; this compound also showed potent anti-inflammatory properties in a mice model of LPS-induced systematic inflammatory response syndrome. Additionally, 15d-PGJ2-PC inhibited macrophage foam cell formation, suggesting a beneficial role against atherosclerosis. These properties were consistent with decreased levels of these compounds in the plasma of patients with coronary heart disease compared with control subjects. Our findings uncovered a novel molecular mechanism for the negative regulation of inflammation and positive enhancement of anti-oxidative responses in macrophages by these oxidized phospholipids in LDL in the context of atherosclerosis.

Discovery of novel lipid profiles in PCOS: Do insulin and androgen oppositely regulate bioactive lipid production?

We found serum lipid profiles changed in women with PCOS. Obesity and compensatory hyperinsulinemia promoted the metabolism of arachidonic acid and other PUFAs, whereas androgen had an inhibitory effect.