SUMMARY Immune cells can metabolize glucose, amino acids, and fatty acids (FAs) to generate energ... more SUMMARY Immune cells can metabolize glucose, amino acids, and fatty acids (FAs) to generate energy. The roles of different FA species and their impacts on humoral immunity remain poorly understood. Here, we report that proliferating B cells require monounsaturated FAs (MUFAs) to maintain mitochondrial metabolism and mTOR activity and to prevent excessive autophagy and endoplasmic reticulum (ER) stress. Furthermore, B cell-extrinsic stearoyl-CoA desaturase (SCD) activity generates MUFA to support early B cell development and germinal center (GC) formation in vivo during immunization and influenza infection. Thus, SCD-mediated MUFA production is critical for humoral immunity.
bioRxiv (Cold Spring Harbor Laboratory), Jun 18, 2024
Mitochondrial creatine kinases are key players in maintaining energy homeostasis in cells by work... more Mitochondrial creatine kinases are key players in maintaining energy homeostasis in cells by working in conjunction with cytosolic creatine kinases for energy transport from mitochondria to cytoplasm. High levels of MtCK observed in Her2+ breast cancer and inhibition of breast cancer cell growth by substrate analog, cyclocreatine, indicate dependence of cancer cells on the 'energy shuttle' for cell growth and survival. Hence, understanding the key mechanistic features of creatine kinases and their inhibition plays an important role in the development of cancer therapeutics. Herein, we present the mutational and structural investigation on understudied ubiquitous mitochondrial creatine kinase (uMtCK). Our cryo-EM structures and biochemical data on uMtCK showed closure of the loop comprising residue His61 is specific to and relies on creatine binding and the reaction mechanism of phosphoryl transfer depends on electrostatics in the active site. In addition, the previously identified covalent inhibitor CKi showed inhibition in breast cancer BT474 cells, however our biochemical and structural data indicated that CKi is not a potent inhibitor for breast cancer due to strong dependency on the covalent link formation and inability to induce conformational changes upon binding.
Alveolar macrophages (AMs) are critical for normal lung homeostasis, surfactant metabolism, and h... more Alveolar macrophages (AMs) are critical for normal lung homeostasis, surfactant metabolism, and host defense against various respiratory pathogens. Despite being terminally differentiated cells, AMs are able to proliferate and self-renew to maintain their compartment without the input of the hematopoietic system in the adulthood during homeostasis. However, the molecular and metabolic mechanisms modulating AM proliferative responses are still incompletely understood. Here we have investigated the metabolic regulation of AM proliferation and self-renewal. Inhibition of glucose uptake or fatty acid oxidation did not significantly impact AM proliferation. Rather, inhibition of the glutamine uptake and/or glutaminase activity impaired AM mitochondrial respiration and cellular proliferation in vitro and in vivo in response to growth factor stimulation. Furthermore, mice with a genetic deletion of glutaminase in macrophages showed decreased proliferation. Our data indicate that glutamine is a critical substrate for fueling mitochondrial metabolism that is required for AM proliferation. Overall, our study is expected to shed light on the AM maintenance and repopulation by glutamine during homeostasis and following acute respiratory viral infection.
BackgroundOverexpression of c-Myc is required for the progression of pre-malignant plasma cells i... more BackgroundOverexpression of c-Myc is required for the progression of pre-malignant plasma cells in monoclonal gammopathy of undetermined significance (MGUS) to malignant plasma cells in multiple myeloma (MM). c-Myc also increases glutamine anaplerosis into the tricarboxylic acid (TCA) cycle within cancer cells. Whether increased glutamine anaplerosis is associated with the progression of pre-malignant to malignant plasma cells is unknown.MethodsHuman volunteers (N = 7) and patients with MGUS (N = 11) and MM (N = 12) were prospectively recruited to undergo an intravenous infusion of 13C-labelled glutamine followed by a bone marrow aspiration to obtain bone marrow cells and plasma.ResultsDespite notable heterogeneity, stable isotope resolved metabolomics (SIRM) revealed that the mean 13C-labelled glutamine anaplerosis into the TCA cycle was higher in malignant compared to pre-malignant bone marrow plasma cells relative to the remainder of their paired bone marrow mononuclear cells. RN...
BRCA1 plays a key role in homologous recombination (HR) DNA repair. Accordingly, changes that dow... more BRCA1 plays a key role in homologous recombination (HR) DNA repair. Accordingly, changes that downregulate BRCA1, including BRCA1 mutations and reduced BRCA1 transcription, due to promoter hypermethylation or loss of the BRCA1 transcriptional regulator CDK12, disrupt HR in multiple cancers. In addition, BRCA1 has also been implicated in the regulation of metabolism. Here, we show that reducing BRCA1 expression, either by CDK12 or BRCA1 depletion, led to metabolic reprogramming of ovarian cancer cells, causing decreased mitochondrial respiration and reduced ATP levels. BRCA1 depletion drove this reprogramming by upregulating nicotinamide N-methyltransferase (NNMT). Notably, the metabolic alterations caused by BRCA1 depletion and NNMT upregulation sensitized ovarian cancer cells to agents that inhibit mitochondrial metabolism (VLX600 and tigecycline) and to agents that inhibit glucose import (WZB117). These observations suggest that inhibition of energy metabolism may be a potential s...
The Warburg effect: a phenomenon of cancer cell metabolism yet poorly understood • The Warburg ef... more The Warburg effect: a phenomenon of cancer cell metabolism yet poorly understood • The Warburg effect in cancer cells consists of an increase in aerobic glycolysis and enhanced lactate production.
MYC gene amplification and somatic mutations are frequent in both adult and pediatric AML althoug... more MYC gene amplification and somatic mutations are frequent in both adult and pediatric AML although how MYC drives and contributes to the development and maintenance of AML has not been resolved. Transcription factor EB (TFEB) is a master regulator of genes that control autophagy and lysosome biogenesis, a central catabolic recycling pathway that regulates cell survival. Given the oncogenic effects of MYC in AML and that the induction of autophagy compromises AML cell growth and survival, we tested if the oncogenic effect of MYC depends on its suppression of TFEB transcription programs in AML. In support of this hypothesis, inducible MYC expression in K562 and THP-1 leukemia cells was sufficient to suppress expression of TFEB and its target genes. Further and conversely, MYC knockdown in NB4 AML cells provoked increased expression of TFEB mRNA and protein, as well as increased expression of TFEB target genes. Notably, dose response studies demonstrated that expression of TFEBS211A, c...
Lysine succinylation (Ksucc), defined as a transfer of a succinyl group to a lysine residue of a ... more Lysine succinylation (Ksucc), defined as a transfer of a succinyl group to a lysine residue of a protein, is a newly identified protein post-translational modification 1e3. This chemical modification is reversible, dynamic, and evolutionarily conserved 4 where it has been comprehensively studied in both bacterial and mammalian cells 5e7. Numerous proteins involved in the regulation of various cellular and biological processes have been shown to be heavily succinylated 5e7. Emerging clinical data provides evidence that dysregulation of Ksucc is correlated with the development of several diseases, including cardiovascular diseases and cancer 7e9. Therefore, an in-depth understanding of Ksucc and its regulation is important not only for understanding its physiological function but also for developing drug therapies and targeted agents for these diseases. In this review, we highlight some of the recent advances in understanding the role of Ksucc and desuccinylation under physiological and pathological conditions.
Highlights d Bhlhe40 is required for Trm cell and TIL fitness and function d Bhlhe40 is critical ... more Highlights d Bhlhe40 is required for Trm cell and TIL fitness and function d Bhlhe40 is critical for TIL reinvigoration following anti-PD-L1 blockade d Bhlhe40 programs Trm cell and TIL mitochondrial metabolism and active chromatin state d Epigenetic targeting Trm cell and TIL functional program promotes tumor control
Lysine succinylation was recently identified as a post-translational modification in cells. Howev... more Lysine succinylation was recently identified as a post-translational modification in cells. However, the molecular mechanism underlying lysine succinylation remains unclear. Here, we show that carnitine palmitoyltransferase 1A (CPT1A) has lysine succinyl-transferase (LSTase) activity in vivo and in vitro. Using a stable isotope labeling by amino acid in cell culture (SILAC)-based proteomics approach, we found that 101 proteins were more succinylated in cells expressing wildtype (WT) CPT1A compared with vector control cells. One of the most heavily succinylated proteins in this analysis was enolase 1. We found that CPT1A WT succinylated enolase 1 and reduced enolase enzymatic activity in cells and in vitro. Importantly, mutation of CPT1A Gly710 (G710E) selectively inactivated carnitine palmitoyltransferase (CPTase) activity but not the LSTase activity that decreased enolase activity in cells and promoted cell proliferation under glutamine depletion. These findings suggest that CPT1A acts as an LSTase that can regulate
The Warburg Effect, or aerobic glycolysis, is one of the major metabolic alterations observed in ... more The Warburg Effect, or aerobic glycolysis, is one of the major metabolic alterations observed in cancer. Hypothesized to increase a cell's proliferative capacity via regenerating NAD + , increasing the pool of glycolytic biosynthetic intermediates, and increasing lactate production that affects the tumor microenvironment, the Warburg Effect is important for the growth and proliferation of tumor cells. The mechanisms by which a cell acquires the Warburg Effect phenotype are regulated by the expression of numerous oncogenes, including oncogenic tyrosine kinases. Oncogenic tyrosine kinases play a significant role in phosphorylating and regulating the activity of numerous metabolic enzymes. Tyrosine phosphorylation of glycolytic enzymes increases the activities of a majority of glycolytic enzymes, thus promoting increased glycolytic rate and tumor cell proliferation. Paradoxically however, tyrosine phosphorylation of pyruvate kinase M2 isoform (PKM2) results in decreased PKM2 activity, and this decrease in PKM2 activity promotes the Warburg Effect. Furthermore, recent studies have shown that PKM2 is also able to act as a protein kinase using phosphoenolpyruvate (PEP) as a substrate to promote tumorigenesis. Therefore, numerous recent studies have investigated both the role of the classical and non-canonical activity of PKM2 in promoting the Warburg Effect and tumor growth, which raise further interesting questions. In this review, we will summarize these recent advances revealing the importance of tyrosine kinases in the regulation of the Warburg Effect as well as the role of PKM2 in the promotion of tumor growth.
How mitochondrial metabolism is altered by oncogenic tyrosine kinases to promote tumor growth is ... more How mitochondrial metabolism is altered by oncogenic tyrosine kinases to promote tumor growth is incompletely understood. Here, we show that oncogenic HER2 tyrosine kinase signaling induces phosphorylation of mitochondrial creatine kinase 1 (MtCK1) on tyrosine 153 (Y153) in an ABL-dependent manner in breast cancer cells. Y153 phosphorylation, which is commonly upregulated in HER2 breast cancers, stabilizes MtCK1 to increase the phosphocreatine energy shuttle and promote proliferation. Inhibition of the phosphocreatine energy shuttle by MtCK1 knockdown or with the creatine analog cyclocreatine decreases proliferation of trastuzumab-sensitive and -resistant HER2 cell lines in culture and in xenografts. Finally, we show that cyclocreatine in combination with the HER2 kinase inhibitor lapatinib reduces the growth of a trastuzumab-resistant HER2 patient-derived xenograft. These findings suggest that activation of the phosphocreatine energy shuttle by MtCK1 Y153 phosphorylation creates a ...
The production of the oncometabolite 2-hydroxyglutarate (2-HG) has been associated with c-MYC ove... more The production of the oncometabolite 2-hydroxyglutarate (2-HG) has been associated with c-MYC overexpression. c-MYC also regulates glutamine metabolism and drives progression of asymptomatic precursor plasma cell (PC) malignancies to symptomatic multiple myeloma (MM). However, the presence of 2-HG and its clinical significance in PC malignancies is unknown. By performing 13C stable isotope resolved metabolomics (SIRM) using U[13C6]Glucose and U[13C5]Glutamine in human myeloma cell lines (HMCLs), we show that 2-HG is produced in clonal PCs and is derived predominantly from glutamine anaplerosis into the TCA cycle. Furthermore, the 13C SIRM studies in HMCLs also demonstrate that glutamine is preferentially utilized by the TCA cycle compared with glucose. Finally, measuring the levels of 2-HG in the BM supernatant and peripheral blood plasma from patients with precursor PC malignancies such as smoldering MM (SMM) demonstrates that relatively elevated levels of 2-HG are associated with ...
Tumor tissues are chronically exposed to hypoxia owing to aberrant vascularity. Lipid droplet (LD... more Tumor tissues are chronically exposed to hypoxia owing to aberrant vascularity. Lipid droplet (LD) accumulation is a hallmark of hypoxic cancer cells, yet how LDs form and function during hypoxia remains poorly understood. Herein, we report that upon oxygen deprivation HIF-1 activation down-modulates LD catabolism mediated by adipose triglyceride lipase (ATGL), the key enzyme for intracellular lipolysis, in various cancer cells. Proteomics and functional analyses identified hypoxia-inducible gene 2 (HIG2), a HIF-1 target, as a new inhibitor of ATGL. Knockout of HIG2 enhanced LD breakdown and fatty acid (FA) oxidation, leading to increased ROS production and apoptosis in hypoxic cancer cells as well as impaired growth of tumor xenografts. All of these effects were reversed by co-ablation of ATGL. Thus, by inhibiting ATGL, HIG2 acts downstream of HIF-1 to sequester FAs in LDs away from the mitochondrial pathways for oxidation and ROS generation, thereby sustaining cancer cell survival...
The oxidative pentose phosphate pathway (PPP) contributes to tumour growth, but the precise contr... more The oxidative pentose phosphate pathway (PPP) contributes to tumour growth, but the precise contribution of 6-phosphogluconate dehydrogenase (6PGD), the third enzyme in this pathway, to tumorigenesis remains unclear. We found that suppression of 6PGD decreased lipogenesis and RNA biosynthesis and elevated ROS levels in cancer cells, attenuating cell proliferation and tumour growth. 6PGD-mediated production of ribulose-5-phosphate (Ru-5-P) inhibits AMPK activation by disrupting the active LKB1 complex, thereby activating acetyl-CoA carboxylase 1 and lipogenesis. Ru-5-P and NADPH are thought to be precursors in RNA biosynthesis and lipogenesis, respectively; thus, our findings provide an additional link between the oxidative PPP and lipogenesis through Ru-5-P-dependent inhibition of LKB1-AMPK signalling. Moreover, we identified and developed 6PGD inhibitors, physcion and its derivative S3, that effectively inhibited 6PGD, cancer cell proliferation and tumour growth in nude mice xenogr...
Many cancer cells rely more on aerobic glycolysis (the Warburg effect) than mitochondrial oxidati... more Many cancer cells rely more on aerobic glycolysis (the Warburg effect) than mitochondrial oxidative phosphorylation and catabolize glucose at a high rate. Such a metabolic switch is suggested to be due in part to functional attenuation of mitochondria in cancer cells. However, how oncogenic signals attenuate mitochondrial function and promote the switch to glycolysis remains unclear. We previously reported that tyrosine phosphorylation activates and inhibits mitochondrial pyruvate dehydrogenase kinase (PDK) and phosphatase (PDP), respectively, leading to enhanced inhibitory serine phosphorylation of pyruvate dehydrogenase (PDH) and consequently inhibition of pyruvate dehydrogenase complex (PDC) in cancer cells. In particular, Tyr-381 phosphorylation of PDP1 dissociates deacetylase SIRT3 and recruits acetyltransferase ACAT1 to PDC, resulting in increased inhibitory lysine acetylation of PDHA1 and PDP1. Here we report that phosphorylation at another tyrosine residue, Tyr-94, inhibits ...
Although the oxidative pentose phosphate pathway is important for tumor growth, how 6-phosphogluc... more Although the oxidative pentose phosphate pathway is important for tumor growth, how 6-phosphogluconate dehydrogenase (6PGD) in this pathway is upregulated in human cancers is unknown. We found that 6PGD is commonly activated in EGF-stimulated cells and human cancer cells by lysine acetylation. Acetylation at K76 and K294 of 6PGD promotes NADP + binding to 6PGD and formation of active 6PGD dimers, respectively. Moreover, we identified DLAT and ACAT2 as upstream acetyltransferases of K76 and K294, respectively, and HDAC4 as the deacetylase of both sites. Expressing acetyl-deficient mutants of 6PGD in cancer cells significantly attenuated cell proliferation and tumor growth. This is due in part to reduced levels of 6PGD products ribulose-5phosphate and NADPH, which led to reduced RNA and lipid biosynthesis as well as elevated ROS. Furthermore, 6PGD activity is upregulated with increased lysine acetylation in primary leukemia cells from human patients, providing mechanistic insights into 6PGD upregulation in cancer cells.
SUMMARY Immune cells can metabolize glucose, amino acids, and fatty acids (FAs) to generate energ... more SUMMARY Immune cells can metabolize glucose, amino acids, and fatty acids (FAs) to generate energy. The roles of different FA species and their impacts on humoral immunity remain poorly understood. Here, we report that proliferating B cells require monounsaturated FAs (MUFAs) to maintain mitochondrial metabolism and mTOR activity and to prevent excessive autophagy and endoplasmic reticulum (ER) stress. Furthermore, B cell-extrinsic stearoyl-CoA desaturase (SCD) activity generates MUFA to support early B cell development and germinal center (GC) formation in vivo during immunization and influenza infection. Thus, SCD-mediated MUFA production is critical for humoral immunity.
bioRxiv (Cold Spring Harbor Laboratory), Jun 18, 2024
Mitochondrial creatine kinases are key players in maintaining energy homeostasis in cells by work... more Mitochondrial creatine kinases are key players in maintaining energy homeostasis in cells by working in conjunction with cytosolic creatine kinases for energy transport from mitochondria to cytoplasm. High levels of MtCK observed in Her2+ breast cancer and inhibition of breast cancer cell growth by substrate analog, cyclocreatine, indicate dependence of cancer cells on the 'energy shuttle' for cell growth and survival. Hence, understanding the key mechanistic features of creatine kinases and their inhibition plays an important role in the development of cancer therapeutics. Herein, we present the mutational and structural investigation on understudied ubiquitous mitochondrial creatine kinase (uMtCK). Our cryo-EM structures and biochemical data on uMtCK showed closure of the loop comprising residue His61 is specific to and relies on creatine binding and the reaction mechanism of phosphoryl transfer depends on electrostatics in the active site. In addition, the previously identified covalent inhibitor CKi showed inhibition in breast cancer BT474 cells, however our biochemical and structural data indicated that CKi is not a potent inhibitor for breast cancer due to strong dependency on the covalent link formation and inability to induce conformational changes upon binding.
Alveolar macrophages (AMs) are critical for normal lung homeostasis, surfactant metabolism, and h... more Alveolar macrophages (AMs) are critical for normal lung homeostasis, surfactant metabolism, and host defense against various respiratory pathogens. Despite being terminally differentiated cells, AMs are able to proliferate and self-renew to maintain their compartment without the input of the hematopoietic system in the adulthood during homeostasis. However, the molecular and metabolic mechanisms modulating AM proliferative responses are still incompletely understood. Here we have investigated the metabolic regulation of AM proliferation and self-renewal. Inhibition of glucose uptake or fatty acid oxidation did not significantly impact AM proliferation. Rather, inhibition of the glutamine uptake and/or glutaminase activity impaired AM mitochondrial respiration and cellular proliferation in vitro and in vivo in response to growth factor stimulation. Furthermore, mice with a genetic deletion of glutaminase in macrophages showed decreased proliferation. Our data indicate that glutamine is a critical substrate for fueling mitochondrial metabolism that is required for AM proliferation. Overall, our study is expected to shed light on the AM maintenance and repopulation by glutamine during homeostasis and following acute respiratory viral infection.
BackgroundOverexpression of c-Myc is required for the progression of pre-malignant plasma cells i... more BackgroundOverexpression of c-Myc is required for the progression of pre-malignant plasma cells in monoclonal gammopathy of undetermined significance (MGUS) to malignant plasma cells in multiple myeloma (MM). c-Myc also increases glutamine anaplerosis into the tricarboxylic acid (TCA) cycle within cancer cells. Whether increased glutamine anaplerosis is associated with the progression of pre-malignant to malignant plasma cells is unknown.MethodsHuman volunteers (N = 7) and patients with MGUS (N = 11) and MM (N = 12) were prospectively recruited to undergo an intravenous infusion of 13C-labelled glutamine followed by a bone marrow aspiration to obtain bone marrow cells and plasma.ResultsDespite notable heterogeneity, stable isotope resolved metabolomics (SIRM) revealed that the mean 13C-labelled glutamine anaplerosis into the TCA cycle was higher in malignant compared to pre-malignant bone marrow plasma cells relative to the remainder of their paired bone marrow mononuclear cells. RN...
BRCA1 plays a key role in homologous recombination (HR) DNA repair. Accordingly, changes that dow... more BRCA1 plays a key role in homologous recombination (HR) DNA repair. Accordingly, changes that downregulate BRCA1, including BRCA1 mutations and reduced BRCA1 transcription, due to promoter hypermethylation or loss of the BRCA1 transcriptional regulator CDK12, disrupt HR in multiple cancers. In addition, BRCA1 has also been implicated in the regulation of metabolism. Here, we show that reducing BRCA1 expression, either by CDK12 or BRCA1 depletion, led to metabolic reprogramming of ovarian cancer cells, causing decreased mitochondrial respiration and reduced ATP levels. BRCA1 depletion drove this reprogramming by upregulating nicotinamide N-methyltransferase (NNMT). Notably, the metabolic alterations caused by BRCA1 depletion and NNMT upregulation sensitized ovarian cancer cells to agents that inhibit mitochondrial metabolism (VLX600 and tigecycline) and to agents that inhibit glucose import (WZB117). These observations suggest that inhibition of energy metabolism may be a potential s...
The Warburg effect: a phenomenon of cancer cell metabolism yet poorly understood • The Warburg ef... more The Warburg effect: a phenomenon of cancer cell metabolism yet poorly understood • The Warburg effect in cancer cells consists of an increase in aerobic glycolysis and enhanced lactate production.
MYC gene amplification and somatic mutations are frequent in both adult and pediatric AML althoug... more MYC gene amplification and somatic mutations are frequent in both adult and pediatric AML although how MYC drives and contributes to the development and maintenance of AML has not been resolved. Transcription factor EB (TFEB) is a master regulator of genes that control autophagy and lysosome biogenesis, a central catabolic recycling pathway that regulates cell survival. Given the oncogenic effects of MYC in AML and that the induction of autophagy compromises AML cell growth and survival, we tested if the oncogenic effect of MYC depends on its suppression of TFEB transcription programs in AML. In support of this hypothesis, inducible MYC expression in K562 and THP-1 leukemia cells was sufficient to suppress expression of TFEB and its target genes. Further and conversely, MYC knockdown in NB4 AML cells provoked increased expression of TFEB mRNA and protein, as well as increased expression of TFEB target genes. Notably, dose response studies demonstrated that expression of TFEBS211A, c...
Lysine succinylation (Ksucc), defined as a transfer of a succinyl group to a lysine residue of a ... more Lysine succinylation (Ksucc), defined as a transfer of a succinyl group to a lysine residue of a protein, is a newly identified protein post-translational modification 1e3. This chemical modification is reversible, dynamic, and evolutionarily conserved 4 where it has been comprehensively studied in both bacterial and mammalian cells 5e7. Numerous proteins involved in the regulation of various cellular and biological processes have been shown to be heavily succinylated 5e7. Emerging clinical data provides evidence that dysregulation of Ksucc is correlated with the development of several diseases, including cardiovascular diseases and cancer 7e9. Therefore, an in-depth understanding of Ksucc and its regulation is important not only for understanding its physiological function but also for developing drug therapies and targeted agents for these diseases. In this review, we highlight some of the recent advances in understanding the role of Ksucc and desuccinylation under physiological and pathological conditions.
Highlights d Bhlhe40 is required for Trm cell and TIL fitness and function d Bhlhe40 is critical ... more Highlights d Bhlhe40 is required for Trm cell and TIL fitness and function d Bhlhe40 is critical for TIL reinvigoration following anti-PD-L1 blockade d Bhlhe40 programs Trm cell and TIL mitochondrial metabolism and active chromatin state d Epigenetic targeting Trm cell and TIL functional program promotes tumor control
Lysine succinylation was recently identified as a post-translational modification in cells. Howev... more Lysine succinylation was recently identified as a post-translational modification in cells. However, the molecular mechanism underlying lysine succinylation remains unclear. Here, we show that carnitine palmitoyltransferase 1A (CPT1A) has lysine succinyl-transferase (LSTase) activity in vivo and in vitro. Using a stable isotope labeling by amino acid in cell culture (SILAC)-based proteomics approach, we found that 101 proteins were more succinylated in cells expressing wildtype (WT) CPT1A compared with vector control cells. One of the most heavily succinylated proteins in this analysis was enolase 1. We found that CPT1A WT succinylated enolase 1 and reduced enolase enzymatic activity in cells and in vitro. Importantly, mutation of CPT1A Gly710 (G710E) selectively inactivated carnitine palmitoyltransferase (CPTase) activity but not the LSTase activity that decreased enolase activity in cells and promoted cell proliferation under glutamine depletion. These findings suggest that CPT1A acts as an LSTase that can regulate
The Warburg Effect, or aerobic glycolysis, is one of the major metabolic alterations observed in ... more The Warburg Effect, or aerobic glycolysis, is one of the major metabolic alterations observed in cancer. Hypothesized to increase a cell's proliferative capacity via regenerating NAD + , increasing the pool of glycolytic biosynthetic intermediates, and increasing lactate production that affects the tumor microenvironment, the Warburg Effect is important for the growth and proliferation of tumor cells. The mechanisms by which a cell acquires the Warburg Effect phenotype are regulated by the expression of numerous oncogenes, including oncogenic tyrosine kinases. Oncogenic tyrosine kinases play a significant role in phosphorylating and regulating the activity of numerous metabolic enzymes. Tyrosine phosphorylation of glycolytic enzymes increases the activities of a majority of glycolytic enzymes, thus promoting increased glycolytic rate and tumor cell proliferation. Paradoxically however, tyrosine phosphorylation of pyruvate kinase M2 isoform (PKM2) results in decreased PKM2 activity, and this decrease in PKM2 activity promotes the Warburg Effect. Furthermore, recent studies have shown that PKM2 is also able to act as a protein kinase using phosphoenolpyruvate (PEP) as a substrate to promote tumorigenesis. Therefore, numerous recent studies have investigated both the role of the classical and non-canonical activity of PKM2 in promoting the Warburg Effect and tumor growth, which raise further interesting questions. In this review, we will summarize these recent advances revealing the importance of tyrosine kinases in the regulation of the Warburg Effect as well as the role of PKM2 in the promotion of tumor growth.
How mitochondrial metabolism is altered by oncogenic tyrosine kinases to promote tumor growth is ... more How mitochondrial metabolism is altered by oncogenic tyrosine kinases to promote tumor growth is incompletely understood. Here, we show that oncogenic HER2 tyrosine kinase signaling induces phosphorylation of mitochondrial creatine kinase 1 (MtCK1) on tyrosine 153 (Y153) in an ABL-dependent manner in breast cancer cells. Y153 phosphorylation, which is commonly upregulated in HER2 breast cancers, stabilizes MtCK1 to increase the phosphocreatine energy shuttle and promote proliferation. Inhibition of the phosphocreatine energy shuttle by MtCK1 knockdown or with the creatine analog cyclocreatine decreases proliferation of trastuzumab-sensitive and -resistant HER2 cell lines in culture and in xenografts. Finally, we show that cyclocreatine in combination with the HER2 kinase inhibitor lapatinib reduces the growth of a trastuzumab-resistant HER2 patient-derived xenograft. These findings suggest that activation of the phosphocreatine energy shuttle by MtCK1 Y153 phosphorylation creates a ...
The production of the oncometabolite 2-hydroxyglutarate (2-HG) has been associated with c-MYC ove... more The production of the oncometabolite 2-hydroxyglutarate (2-HG) has been associated with c-MYC overexpression. c-MYC also regulates glutamine metabolism and drives progression of asymptomatic precursor plasma cell (PC) malignancies to symptomatic multiple myeloma (MM). However, the presence of 2-HG and its clinical significance in PC malignancies is unknown. By performing 13C stable isotope resolved metabolomics (SIRM) using U[13C6]Glucose and U[13C5]Glutamine in human myeloma cell lines (HMCLs), we show that 2-HG is produced in clonal PCs and is derived predominantly from glutamine anaplerosis into the TCA cycle. Furthermore, the 13C SIRM studies in HMCLs also demonstrate that glutamine is preferentially utilized by the TCA cycle compared with glucose. Finally, measuring the levels of 2-HG in the BM supernatant and peripheral blood plasma from patients with precursor PC malignancies such as smoldering MM (SMM) demonstrates that relatively elevated levels of 2-HG are associated with ...
Tumor tissues are chronically exposed to hypoxia owing to aberrant vascularity. Lipid droplet (LD... more Tumor tissues are chronically exposed to hypoxia owing to aberrant vascularity. Lipid droplet (LD) accumulation is a hallmark of hypoxic cancer cells, yet how LDs form and function during hypoxia remains poorly understood. Herein, we report that upon oxygen deprivation HIF-1 activation down-modulates LD catabolism mediated by adipose triglyceride lipase (ATGL), the key enzyme for intracellular lipolysis, in various cancer cells. Proteomics and functional analyses identified hypoxia-inducible gene 2 (HIG2), a HIF-1 target, as a new inhibitor of ATGL. Knockout of HIG2 enhanced LD breakdown and fatty acid (FA) oxidation, leading to increased ROS production and apoptosis in hypoxic cancer cells as well as impaired growth of tumor xenografts. All of these effects were reversed by co-ablation of ATGL. Thus, by inhibiting ATGL, HIG2 acts downstream of HIF-1 to sequester FAs in LDs away from the mitochondrial pathways for oxidation and ROS generation, thereby sustaining cancer cell survival...
The oxidative pentose phosphate pathway (PPP) contributes to tumour growth, but the precise contr... more The oxidative pentose phosphate pathway (PPP) contributes to tumour growth, but the precise contribution of 6-phosphogluconate dehydrogenase (6PGD), the third enzyme in this pathway, to tumorigenesis remains unclear. We found that suppression of 6PGD decreased lipogenesis and RNA biosynthesis and elevated ROS levels in cancer cells, attenuating cell proliferation and tumour growth. 6PGD-mediated production of ribulose-5-phosphate (Ru-5-P) inhibits AMPK activation by disrupting the active LKB1 complex, thereby activating acetyl-CoA carboxylase 1 and lipogenesis. Ru-5-P and NADPH are thought to be precursors in RNA biosynthesis and lipogenesis, respectively; thus, our findings provide an additional link between the oxidative PPP and lipogenesis through Ru-5-P-dependent inhibition of LKB1-AMPK signalling. Moreover, we identified and developed 6PGD inhibitors, physcion and its derivative S3, that effectively inhibited 6PGD, cancer cell proliferation and tumour growth in nude mice xenogr...
Many cancer cells rely more on aerobic glycolysis (the Warburg effect) than mitochondrial oxidati... more Many cancer cells rely more on aerobic glycolysis (the Warburg effect) than mitochondrial oxidative phosphorylation and catabolize glucose at a high rate. Such a metabolic switch is suggested to be due in part to functional attenuation of mitochondria in cancer cells. However, how oncogenic signals attenuate mitochondrial function and promote the switch to glycolysis remains unclear. We previously reported that tyrosine phosphorylation activates and inhibits mitochondrial pyruvate dehydrogenase kinase (PDK) and phosphatase (PDP), respectively, leading to enhanced inhibitory serine phosphorylation of pyruvate dehydrogenase (PDH) and consequently inhibition of pyruvate dehydrogenase complex (PDC) in cancer cells. In particular, Tyr-381 phosphorylation of PDP1 dissociates deacetylase SIRT3 and recruits acetyltransferase ACAT1 to PDC, resulting in increased inhibitory lysine acetylation of PDHA1 and PDP1. Here we report that phosphorylation at another tyrosine residue, Tyr-94, inhibits ...
Although the oxidative pentose phosphate pathway is important for tumor growth, how 6-phosphogluc... more Although the oxidative pentose phosphate pathway is important for tumor growth, how 6-phosphogluconate dehydrogenase (6PGD) in this pathway is upregulated in human cancers is unknown. We found that 6PGD is commonly activated in EGF-stimulated cells and human cancer cells by lysine acetylation. Acetylation at K76 and K294 of 6PGD promotes NADP + binding to 6PGD and formation of active 6PGD dimers, respectively. Moreover, we identified DLAT and ACAT2 as upstream acetyltransferases of K76 and K294, respectively, and HDAC4 as the deacetylase of both sites. Expressing acetyl-deficient mutants of 6PGD in cancer cells significantly attenuated cell proliferation and tumor growth. This is due in part to reduced levels of 6PGD products ribulose-5phosphate and NADPH, which led to reduced RNA and lipid biosynthesis as well as elevated ROS. Furthermore, 6PGD activity is upregulated with increased lysine acetylation in primary leukemia cells from human patients, providing mechanistic insights into 6PGD upregulation in cancer cells.
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Papers by Taro Hitosugi