SUCLA2-coupled regulation of GLS succinylation and activity counteracts oxidative stress in tumor cells.

Glutaminase regulates glutaminolysis to promote cancer cell proliferation. However, the mechanism underlying glutaminase activity regulation is largely unknown. Here, we demonstrate that kidney-type glutaminase (GLS) is highly expressed in human pancreatic ductal adenocarcinoma (PDAC) specimens with correspondingly upregulated glutamine dependence for PDAC cell proliferation. Upon oxidative stress, the succinyl-coenzyme A (CoA) synthetase ADP-forming subunit ß (SUCLA2) phosphorylated by p38 mitogen-activated protein kinase (MAPK) at S79 dissociates from GLS, resulting in enhanced GLS K311 succinylation, oligomerization, and activity. Activated GLS increases glutaminolysis and the production of nicotinamide adenine dinucleotide phosphate (NADPH) and glutathione, thereby counteracting oxidative stress and promoting tumor cell survival and tumor growth in mice. In addition, the levels of SUCLA2 pS79 and GLS K311 succinylation, which were mutually correlated, were positively associated with advanced stages of PDAC and poor prognosis for patients. Our findings reveal critical regulation of GLS by SUCLA2-coupled GLS succinylation regulation and underscore the regulatory role of metabolites in glutaminolysis and PDAC development.

Induction of MTHFD2 in Macrophages Inhibits Reactive Oxygen Species-mediated NF-κB Activation and Protects against Inflammatory Responses.

The one-carbon metabolism enzyme methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) is critical for cancer cell proliferation and immune cell phenotypes, but whether it can contribute to macrophage inflammatory responses remains unclear. In this study, we show that MTHFD2 was upregulated by LPS in murine macrophages upon activation of the TLR4-MyD88-IKKα/ß-NF-κB signaling pathway. MTHFD2 significantly attenuated LPS-induced macrophage proinflammatory cytokine production through its enzymatic activity. Notably, ablation of myeloid MTHFD2 rendered mice more sensitive to septic shock and CCl4-induced acute hepatitis. Mechanistically, MTHFD2 restrained IKKα/ß-NF-κB activation and macrophage inflammatory phenotype by scavenging reactive oxygen species through the generation of NADPH. Our study reveals MTHFD2 as a "self-control" mechanism in macrophage-mediated inflammatory responses.

Phosphoglycerate Kinase 1 Phosphorylates Beclin1 to Induce Autophagy.

Autophagy is crucial for maintaining cell homeostasis. However, the precise mechanism underlying autophagy initiation remains to be defined. Here, we demonstrate that glutamine deprivation and hypoxia result in inhibition of mTOR-mediated acetyl-transferase ARD1 S228 phosphorylation, leading to ARD1-dependent phosphoglycerate kinase 1 (PGK1) K388 acetylation and subsequent PGK1-mediated Beclin1 S30 phosphorylation. This phosphorylation enhances ATG14L-associated class III phosphatidylinositol 3-kinase VPS34 activity by increasing the binding of phosphatidylinositol to VPS34. ARD1-dependent PGK1 acetylation and PGK1-mediated Beclin1 S30 phosphorylation are required for glutamine deprivation- and hypoxia-induced autophagy and brain tumorigenesis. Furthermore, PGK1 K388 acetylation levels correlate with Beclin1 S30 phosphorylation levels and poor prognosis in glioblastoma patients. Our study unearths an important mechanism underlying cellular-stress-induced autophagy initiation in which the protein kinase activity of the metabolic enzyme PGK1 plays an instrumental role and reveals the significance of the mutual regulation of autophagy and cell metabolism in maintaining cell homeostasis.

Type I interferons mediate pancreatic toxicities of PERK inhibition.

The great preclinical promise of the pancreatic endoplasmic reticulum kinase (PERK) inhibitors in neurodegenerative disorders and cancers is marred by pancreatic injury and diabetic syndrome observed in PERK knockout mice and humans lacking PERK function and suffering from Wolcott-Rallison syndrome. PERK mediates many of the unfolded protein response (UPR)-induced events, including degradation of the type 1 interferon (IFN) receptor IFNAR1 in vitro. Here we report that whole-body or pancreas-specific Perk ablation in mice leads to an increase in IFNAR1 protein levels and signaling in pancreatic tissues. Concurrent IFNAR1 deletion attenuated the loss of PERK-deficient exocrine and endocrine pancreatic tissues and prevented the development of diabetes. Experiments using pancreas-specific Perk knockouts, bone marrow transplantation, and cultured pancreatic islets demonstrated that stabilization of IFNAR1 and the ensuing increased IFN signaling in pancreatic tissues represents a major driver of injury triggered by Perk loss. Neutralization of IFNAR1 prevented pancreatic toxicity of PERK inhibitor, indicating that blocking the IFN pathway can mitigate human genetic disorders associated with PERK deficiency and help the clinical use of PERK inhibitors.

Type I interferon controls propagation of long interspersed element-1.

Type I interferons (IFN) including IFNα and IFNß are critical for the cellular defense against viruses. Here we report that increased levels of IFNß were found in testes from mice deficient in MOV10L1, a germ cell-specific RNA helicase that plays a key role in limiting the propagation of retrotransposons including Long Interspersed Element-1 (LINE-1). Additional experiments revealed that activation of LINE-1 retrotransposons increases the expression of IFNß and of IFN-stimulated genes. Conversely, pretreatment of cells with IFN suppressed the replication of LINE-1. Furthermore, the efficacy of LINE-1 replication was increased in isogenic cell lines harboring inactivating mutations in diverse elements of the IFN signaling pathway. Knockdown of the IFN receptor chain IFNAR1 also stimulated LINE-1 propagation in vitro. Finally, a greater accumulation of LINE-1 was found in mice that lack IFNAR1 compared with wild type mice. We propose that LINE-1-induced IFN plays an important role in restricting LINE-1 propagation and discuss the putative role of IFN in preserving the genome stability.

Fam72a functions as a cell-cycle-controlled gene during proliferation and antagonizes apoptosis through reprogramming PP2A substrates.

The cell cycle is key to life. After decades of research, it is unclear whether any parts of this process have yet to be identified. Fam72a is a poorly characterized gene and is evolutionarily conserved across multicellular organisms. Here, we have found that Fam72a is a cell-cycle-regulated gene that is transcriptionally and post-transcriptionally regulated by FoxM1 and APC/C, respectively. Functionally, Fam72a directly binds to tubulin and both the Aα and B56 subunits of PP2A-B56 to modulate tubulin and Mcl1 phosphorylation, which in turn affects the progression of the cell cycle and signaling of apoptosis. Moreover, Fam72a is involved in early responses to chemotherapy, and it efficiently antagonizes various anticancer compounds such as CDK and Bcl2 inhibitors. Thus, Fam72a switches the tumor-suppressive PP2A to be oncogenic by reprogramming its substrates. These findings identify a regulatory axis of PP2A and a protein member in the cell cycle and tumorigenesis regulatory network in human cells.

Nuclear Fructose-1,6-Bisphosphate Inhibits Tumor Growth and Sensitizes Chemotherapy by Targeting HMGB1.

Metabolites are important for cell fate determination. Fructose-1,6-bisphosphate (F1,6P) is the rate-limiting product in glycolysis and the rate-limiting substrate in gluconeogenesis. Here, it is discovered that the nuclear-accumulated F1,6P impairs cancer cell viability by directly binding to high mobility group box 1 (HMGB1), the most abundant non-histone chromosome structural protein with paradoxical roles in tumor development. F1,6P disrupts the association between the HMGB1 A-box and C-tail by targeting K43/K44 residues, inhibits HMGB1 oligomerization, and stabilizes P53 protein by increasing P53-HMGB1 interaction. Moreover, F1,6P lowers the affinity of HMGB1 for DNA and DNA adducts, which sensitizes cancer cells to chemotherapeutic drug(s)-induced DNA replication stress and DNA damage. Concordantly, F1,6P resensitizes cancer cells with chemotherapy resistance, impairs tumor growth and enhances chemosensitivity in mice, and impedes the growth of human tumor organoids. These findings reveal a novel role for nuclear-accumulated F1,6P and underscore the potential utility of F1,6P as a drug for cancer therapy.

MTHFD2 reprograms macrophage polarization by inhibiting PTEN.

The one-carbon metabolism enzyme methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) is involved in the regulation of tumor oncogenesis and immune cell functions, but whether it can contribute to macrophage polarization remains elusive. Here, we show that MTHFD2 suppresses polarization of interferon-γ-activated macrophages (M(IFN-γ)) but enhances that of interleukin-4-activated macrophages (M(IL-4)) both in vitro and in vivo. Mechanistically, MTHFD2 interacts with phosphatase and tensin homolog (PTEN) to suppress PTEN's phosphatidylinositol 3,4,5-trisphosphate (PIP3) phosphatase activity and enhance downstream Akt activation, independent of the N-terminal mitochondria-targeting signal of MTHFD2. MTHFD2-PTEN interaction is promoted by IL-4 but not IFN-γ. Furthermore, amino acid residues (aa 215-225) of MTHFD2 directly target PTEN catalytic center (aa 118-141). Residue D168 of MTHFD2 is also critical for regulating PTEN's PIP3 phosphatase activity by affecting MTHFD2-PTEN interaction. Our study suggests a non-metabolic function of MTHFD2 by which MTHFD2 inhibits PTEN activity, orchestrates macrophage polarization, and alters macrophage-mediated immune responses.

Aerobic glycolysis promotes tumor immune evasion by hexokinase2-mediated phosphorylation of IκBα.

High expression of PD-L1 in tumor cells contributes to tumor immune evasion. However, whether PD-L1 expression in tumor cells is regulated by the availability of nutrients is unknown. Here, we show that in human glioblastoma cells, high glucose promotes hexokinase (HK) 2 dissociation from mitochondria and its subsequent binding and phosphorylation of IκBα at T291. This leads to increased interaction between IκBα and µ-calpain protease and subsequent µ-calpain-mediated IκBα degradation and NF-κB activation-dependent transcriptional upregulation of PD-L1 expression. Expression of IκBα T291A in glioblastoma cells blocked high glucose-induced PD-L1 expression and promoted CD8+ T cell activation and infiltration into the tumor tissue, reducing brain tumor growth. Combined treatment with an HK inhibitor and an anti-PD-1 antibody eliminates tumor immune evasion and remarkably enhances the anti-tumor effect of immune checkpoint blockade. These findings elucidate a novel mechanism underlying the upregulation of PD-L1 expression mediated by aerobic glycolysis and underscore the roles of HK2 as a glucose sensor and a protein kinase in regulation of tumor immune evasion.

Serine metabolism orchestrates macrophage polarization by regulating the IGF1-p38 axis.

Serine metabolism is reportedly involved in immune cell functions, but whether and how serine metabolism regulates macrophage polarization remain largely unknown. Here, we show that suppressing serine metabolism, either by inhibiting the activity of the key enzyme phosphoglycerate dehydrogenase in the serine biosynthesis pathway or by exogenous serine and glycine restriction, robustly enhances the polarization of interferon-γ-activated macrophages (M(IFN-γ)) but suppresses that of interleukin-4-activated macrophages (M(IL-4)) both in vitro and in vivo. Mechanistically, serine metabolism deficiency increases the expression of IGF1 by reducing the promoter abundance of S-adenosyl methionine-dependent histone H3 lysine 27 trimethylation. IGF1 then activates the p38-dependent JAK-STAT1 axis to promote M(IFN-γ) polarization and suppress STAT6-mediated M(IL-4) activation. This study reveals a new mechanism by which serine metabolism orchestrates macrophage polarization and suggests the manipulation of serine metabolism as a therapeutic strategy for macrophage-mediated immune diseases.