Aurora kinase A promotes trained immunity via regulation of endogenous S-adenosylmethionine metabolism
- MOE Key Laboratory of Gene Function and Regulation, Guangdong Province Key Laboratory of Pharmaceutical Functional Genes, State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, China
- State Key Laboratory of Oncology in South China, Guangdong Provincial Clinical Research Center for Cancer, Sun Yat-sen University Cancer Center, China
- Guangzhou National Laboratory, Guangzhou International Bio-Island, China
- Department of Pathology, School of Basic Medical Sciences, Southern Medical University, China
- Department of Pathology, Nanfang Hospital, Southern Medical University, China
- Guangdong Provincial Key Laboratory of Molecular Tumor Pathology, China
- Metabolic Center, Zhongshan School of Medicine, Sun Yat-sen University, China
- Center for Stem Cell Biology and Tissue Engineering, Key Laboratory for Stem Cells and Tissue Engineering, Ministry of Education, Sun Yat-sen University, China
- Hainan Academy of Medical Sciences, Hainan Medical University, China
Figures
Figure 1 with 1 supplement
Inhibition of Aurora kinase A suppresses trained immunity in macrophages.
(A) Bone marrow-derived macrophages (BMDMs) were trained with β-glucan at a dosage of 50 μg/mL in the presence of different concentrations of alisertib for 24 hr. The viability of BMDMs was measured by CCK8 assay. (B) Supernatant levels of IL-6 (left) and TNF (right) in trained BMDMs with alisertib (0.5 μM or 1 μM), followed by restimulation with LPS (100 ng/mL) for 24 hr. (C) qPCR analysis of relative mRNA expression of Il6 and Tnf in trained BMDMs with alisertib (0.5 μM or 1 μM), followed by restimulation with LPS (100 ng/mL) for 6 hr. Actb served as a reference gene. (D) Immunoblotting analysis of Aurora kinase A (AurA) phosphorylation after the treatment of β-glucan (50 μg/mL) with or without alisertib (1 μM) for 90 min. (E) Immunoblotting analysis of AurA in BMDMs transfected with siRNA targeting AurA for 48 hr. (F) The BMDMs were firstly transfected with siRNAs for 48 hr and then stimulated with β-glucan (50 μg/mL). Supernatant levels of IL-6 and TNF were detected by ELISA after 3 days rest and restimulation with LPS (100 ng/mL) for 24 hr. (G) The BMDMs was stimulated with β-glucan (50 μg/mL) together with AurA knockdown or alisertib, followed by a rest for 3 days and restimulation with MC38 cell culture supernatant for 48 hr. (H) Graphical outline of in vivo training model (3 mice per group). Intraperitoneal injection, i.p; intragastric adminstration, i.g. (I) Supernatant levels of IL-6 (left) and TNF (right) in trained BMDMs as shown in (H). Data are presented as the mean ± SEM (except mean ± SD for in vivo training in I). P values were derived from one-way ANOVA with Dunnett’s multiple-comparison test (A, F, G, I), compared with only β-glucan stimulation group; or two-way ANOVA with Tukey’s multiple-comparison test (B, C), compared with every other group. In D, similar results were obtained from three independent experiments. Related to Figure 1—figure supplement 1, Figure 1—source data 1–2.
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Figure 1—source data 1
Uncropped and labeled blots for Figure 1D and E.
- https://cdn.elifesciences.org/articles/104138/elife-104138-fig1-data1-v1.zip
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Figure 1—source data 2
Raw unedited blots for Figure 1D and E.
- https://cdn.elifesciences.org/articles/104138/elife-104138-fig1-data2-v1.zip
Figure 1—figure supplement 1
Targeting aurora A inhibits β-glucan-induced trained immunity.
(A) Schematic of the assay protocol for the drug screening in bone marrow-derived macrophages (BMDMs). (B) Fold change of IL-6 production showing the outcomes of the entire drug screening, including inhibitors targeting Aurora kinase, JAK/STAT pathway, PI3K/AKT/mTOR pathway, histone methyltransferases (HMT), lysine-specific demethylases (KDM), histone deacetylases (HDAC), Sirtuin (SIRT), Bromodomain and PARP families. (C) Fold change of IL-6 production inhibited by AurA inhibitors compared to that of β-glucan only group. (D) BMDMs were trained with β-glucan (50 μg/mL) in the presence of Tozasertib (1 μM), alisertib (1 μM), or MLN8054 (1 μM) for 24 hr, followed by LPS (100 ng/mL) stimulation for 6 hr after a rest for 3 days. (E) Supernatant levels of IL-6 in trained J774A.1 cells and THP-1 cells. J774A.1 cells were transfected with AurA-specific siRNAs followed by β-glucan (50 μg/mL) stimulation. After resting for 3 days, trained J774A.1 cells was counted and seeded into cell culture plate with MC38 supernatant rechallenge for 48 hr. THP-1 cells were trained with β-glucan (50 μg/mL) for 24 hr, and were centrifuged and washed once to remove medium. The THP-1 cells were then cultured with fresh medium containing 10 ng/mL PMA for 48 hr and rest 1 day, followed by LPS rechallenge (100 ng/mL). Data in D and E are representative of three independent experiments and presented as the mean ± SEM. P values were derived from one-way ANOVA with Dunnett’s multiple-comparison test, compared with β-glucan stimulation only group. Related to Figure 1.
Figure 2 with 1 supplement
Aurora kinase A inhibition remodels chromatin landscape of inflammatory genes.
(A) Principal component analysis (PCA) of gene peaks in ATAC-seq (n=2 mice per group). (B) Gene ontology (GO) enrichment analysis of erased peaks by alisertib in trained bone marrow-derived macrophages (BMDMs). (C) Representative motifs in the erased (n=15,431) and written (n=19,733) peaks, respectively. (D, E) Genome browser views of ATAC-seq signal of representative genes inhibited by alisertib, including Cxcl2, Il1a, Tnf, and Il6 (D) and representative genes enhanced by alisertib, including Mrc1 and Chil3 (E). (F, G) KEGG enrichment of differentially expressed genes in trained BMDMs rechallenged with LPS; alisertib downregulated genes (F) and upregulated genes (G) were mapped into KEGG, respectively. Related to Figure 2—figure supplement 1.
Figure 2—figure supplement 1
Aurora kinase A inhibition suppresses the expression of transcription factors involved in inflammation activation.
(A) In vivo training model in C57BL/6 J mice with intraperitoneal injection of β-glucan (2 mg per mice) and daily administration of alisertib (30 mg/kg/d) for 7 days (n=2 mice per group). (B) Heatmap from RNA-seq analysis showing the differentially expressed transcription factors (DETFs) from mice treated as described in (A). (C) Gene ontology (GO) enrichment analysis of differentially expressed transcription factors (DETFs). (D) Multiplex immunoassay measuring 18 cytokines/chemokines in supernatant from trained BMDMs as described in (A), which were rechallenged with LPS (100 ng/mL) for 6 hr. Related to Figure 2.
Figure 3 with 1 supplement
Aurora kinase A inhibition decreases glycolysis and S-adenosylmethionine (SAM) level.
(A, B) Extracellular acidification rate (ECAR) in bone marrow-derived macrophages (BMDMs) with different treatments after a glycolysis stress test upon sequential addition of glucose (Gluc, 10 mM), oligomycin (Oligo, 1 μM), and 2-deoxyglucose (2-DG, 50 mM), as indicated (A); basal glycolysis and maximal glycolysis (B). (C, F, G) LC–MS/MS measurements of fumarate (C), serine and SAM (F), S-adenosylhomocysteine (SAH) and HCY (G) in trained BMDMs treated with vehicle or alisertib for 24 hr. (D) BMDMs were trained with β-glucan (50 μg/mL) with or without alisertib (1 μM) for 24 hr. The BMDMs were collected for RNA extraction and followed by RNA-seq (n=2 mice per group). The TOP 10 enriched pathways identified by KEGG enrichment analysis of differentially expressed genes (Fold change >1.2, FDR <0.05) by comparing trained BMDMs with or without alisertib. (E) Intracellular levels of glutathione in trained BMDMs with or without alisertib for 24 hr. The level was normalized to untrained BMDMs. (H) Western blot analysis of GNMT in trained BMDMs treated with vehicle or alisertib for 24 hr. β-actin was used as a loading control; * showed the position of GNMT protein. (I) Western blot showing GNMT protein levels in wild-type BMDMs that were transfected with siRNA targeting GNMT. (J) LC–MS/MS measurements of SAM and SAH in trained BMDMs treated by alisertib together with or without knockdown of GNMT. The SAM/SAH ratio is calculated by SAH normalization. P values were derived from two-tailed t-tests. (K) Supernatant levels of IL-6 and TNF in trained BMDMs with AurA inhibition by alisertib or by siRNAs targeting AurA or GNMT. N=3 per group. Data are presented as the mean ± SEM. P values were derived from one-way ANOVA with Tukey’s multiple-comparison test (B, K) compared with every other group or with Dunnett’s multiple-comparison test (C, E–G) compared with only β-glucan stimulation group. In H and I, similar results were obtained for three independent experiments. Related to Figure 3—figure supplement 1, Figure 3—source data 1–2.
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Figure 3—source data 1
Uncropped and labeled blots for Figure 3H and I.
- https://cdn.elifesciences.org/articles/104138/elife-104138-fig3-data1-v1.zip
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Figure 3—source data 2
Raw unedited blots for Figure 3H and I.
- https://cdn.elifesciences.org/articles/104138/elife-104138-fig3-data2-v1.zip
Figure 3—figure supplement 1
Alisertib inhibits glucose incorporation into glycolysis and tricarboxylic acid (TCA) cycle.
(A) Oxygen consumption rate (OCR) in bone marrow-derived macrophages (BMDMs) with different treatments after a mito stress test upon sequential addition of oligomycin (oligo, 1 μM), FCCP (1 mM), and rotenone/antimycin A (0.5 μM), as indicated; (B) Mass labeling of trained BMDMs with U-13C-glucose in the absence or presence of alisertib for 24 hr. Training by β-glucan increased incorporation of 13C-glucose into glycolysis and TCA cycle intermediates; this was reversed by alisertib. (C) Peak area of tyrosine and comparing of the sum of peak areas for unlabeled and labeled tyrosine between different treatment groups. (D) Diagram illustrating the cross-link between glycolysis, TCA cycle, glutathione, and S-adenosylmethionine (SAM). N=3 per group. Data are presented as the mean ± SEM and p-values were derived from one-way ANOVA (B) or two-way ANOVA (C) with Dunnett’s multiple-comparison test. Related to Figure 3.
Figure 4 with 1 supplement
Inhibition of Aurora kinase A impairs histone trimethylation at H3K4 and H3K36.
(A) Western blot analysis of histone methylation levels in trained BMDM treated with vehicles or alisertib. Histone 3 (H3) was used as a loading control. bone marrow-derived macrophages (BMDMs) were trained with β-glucan (50 μg/mL) with or without alisertib (1 μM) for 24 hr, then BMDMs were washed and cultured in fresh medium for 3 days, followed by protein extraction and Western blot analysis. (B) ChIP-qPCR analysis of H3K4me3 and H3K36me3 enrichment in Il6 and Tnf promoter regions in trained BMDMs treated with vehicles or alisertib for 24 h and rest for 3 days. N=3 per group. (C) Western blot analysis of total H3K4me3 and H3K36me3 levels upon GNMT knockdown in BMDMs. The BMDMs were transfected with siRNA targeting GNMT for 48 hr, followed by β-glucan (50 μg/mL) with or without alisertib (1 μM) treatment for 24 hr. Then the BMDMs were washed and cultured in fresh medium for 3 days and the protein was extracted for Western blot analysis. Data are presented as the mean ± SEM. P values were derived from one-way ANOVA with Tukey’s multiple-comparison test compared with each other. In A and C, similar results were obtained for three independent experiments. Related to Figure 4—figure supplement 1, Figure 4—source data 1–2.
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Figure 4—source data 1
Uncropped and labeled blots for Figure 4A and C.
- https://cdn.elifesciences.org/articles/104138/elife-104138-fig4-data1-v1.zip
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Figure 4—source data 2
Raw unedited blots for Figure 4A and C.
- https://cdn.elifesciences.org/articles/104138/elife-104138-fig4-data2-v1.zip
Figure 4—figure supplement 1
Inhibition of Aurora kinase A impairs histone trimethylation at H3K4 and H3K36.
Quantification of H3K4me1, H3K9me3, H3K36me3, H3K4me3, and H3K27me3 protein levels was determined by Image Lab software; data are represented as mean ± SEM of three independent experiments. P values were derived from one-way ANOVA with Dunnett’s multiple-comparison test compared with β-glucan stimulation only group. Related to Figure 4.
Figure 5
Aurora kinase A regulates glycine N-methyltransferase (GNMT) through transcription factor FOXO3.
(A) Protein level of GNMT in bone marrow-derived macrophages (BMDMs) with FOXO3 knockdown was detected by Western blot; * showed the position of FOXO3 band. (B, C) Western blot analysis of GNMT downregulation by siFoxo3 in trained BMDMs with AurA inhibition. BMDMs were transfected with siRNA targeting FOXO3 for 48 hr, followed by β-glucan (50 μg/mL) and alisertib (1 μM) for 24 hr (B); BMDMs were co-transfected with siRNAs targeting FOXO3 and AurA for 48 hr, followed by β-glucan (50 μg/mL) stimulation for another 24 hr (C). (D) Supernatant levels of IL-6 in BMDMs. The cells were treated with β-glucan (50 μg/mL) and alisertib (1 μM) after transfection of siRNAs targeting FOXO3. (E) Western blot analysis of phosphorylation level of FOXO3 at ser 315 in BMDMs treated with β-glucan (50 μg/mL) with or without alisertib (1 μM) for 12 hr. (F) Immunofluorescence staining of FOXO3 in BMDMs after 12 hr β-glucan stimulation with or without alisertib. Scale bars: 10 μm (left). The nuclear localization of FOXO3 was compared by calculating the ratio of mean nuclear intensity to cytoplasmic intensity, and the representative data (right) showed the mean intensity of counted macrophages. (G) Western blot analysis of AKT-mTOR-S6 pathway in β-glucan-trained BMDM. BMDMs were transfected with siRNA targeting AurA for 48 hr, followed by β-glucan stimulation for 6 hr (left); BMDM was trained with β-glucan in the absence or presence of alisertib for 6 hr (right). (H) Supernatant levels of IL-6 and TNF in BMDMs. The trained cells were treated with siRNA targeting AurA or alisertib in combination with an mTOR agonist, MHY1485 (2 μM), and restimulated with MC38 culture supernatant for 48 hr. Data in D and H are representative of three independent experiments and presented as the mean ± SEM. P values were derived from one-way ANOVA with Tukey’s multiple-comparison test compared with each other in D and H, or with Dunnett’s multiple-comparison test in F compared with β-glucan only group. In A-C and E-G, similar results were obtained for three independent experiments. Related to Figure 5—source data 1–3.
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Figure 5—source data 1
Uncropped and labeled blots for Figure 5A–C, E and G.
- https://cdn.elifesciences.org/articles/104138/elife-104138-fig5-data1-v1.zip
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Figure 5—source data 2
Raw unedited blots for Figure 5A–C, E and G.
- https://cdn.elifesciences.org/articles/104138/elife-104138-fig5-data2-v1.zip
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Figure 5—source data 3
Original images for Figure 5F.
- https://cdn.elifesciences.org/articles/104138/elife-104138-fig5-data3-v1.zip
Figure 6 with 1 supplement
Alisertib abrogates the anti-tumor effect induced by trained immunity.
(A) Experimental scheme of tumor model. 6~8-weeks-old mice was injected with β-glucan together with vehicle or alisertib, followed by subcutaneous inoculation of MC38 cells (1×106 cells/mouse, n=5 per group). (B) The MC38 tumor growth curves of mice in A. (C) Experimental scheme of bone marrow transplantation model. CD45.1+ mice (n=3 per group) as donor were treated as indicated, and CD45.2+ mice (n=7 per group) as recipient received 4×106 bone marrow (BM) cells from CD45.1+ mice at day 8 post irradiation with 9 Gy. After 4 weeks, CD45.2+ mice were inoculated with MC38 cells. (D) The MC38 tumor growth curves of mice in C. (E) Flow cytometric analysis of myeloid cells (CD45+CD11b+) and macrophages (CD45+CD11b+F4/80+) in MC38 tumors in A. Gating strategy was shown in Figure 6—figure supplement 1D. (F) Co-immunofluorescence staining of DAPI, F4/80, and glycine N-methyltransferase (GNMT) in tumor section from A; Scale bars: 20 μm. (G) Flow cytometry analysis for intracellular phospho-S6 in macrophages from tumors in A. (H) Tumor tissue was lysed and the lysates were collected for detection of IL-6 by ELISA. (I) FACS analysis of intracellular IL-6 in tumor-infiltrated myeloid cells and macrophages from tumors in A. Data are represented as the mean ± SD. P values were derived from one-way ANOVA (E, F, G, H, I) or two-way ANOVA (B, D) with Dunnett’s multiple-comparison test with data from mice treated with β-glucan only as control. Related to Figure 6—figure supplement 1, Figure 6—source data 1.
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Figure 6—source data 1
Intensity analysis for Figure 6F.
- https://cdn.elifesciences.org/articles/104138/elife-104138-fig6-data1-v1.zip
Figure 6—figure supplement 1
Alisertib abrogates the anti-tumor effect induced by trained immunity.
(A) Individual tumor growth curves (n=5 per group). (B) Representative FACS analysis for CD45.1+ and CD45.2+ cells in the spleen of a chimeric mouse at the end of experiment (day 19 after tumor inoculation). (C) Individual tumor growth curves of chimeric mice (n=7 per group). (D) The gating strategy used for analyzing tumor-infiltrated myeloid cells and macrophages. (E) Tumor tissue from Figure 6A was lysed and the lysates were collected for detection of IL-1β and IL-12p70 (n=5). Data are represented as mean ± SD. P values were derived from one-way ANOVA with Dunnett’s multiple-comparison test with data from mice treated with β-glucan only as control. Related to Figure 6.
Tables
Appendix 1—key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Chemical compound, drug | β-glucan | Sigma-Aldrich | Cat# G6513 | Inducer of Trained immunity |
| Chemical compound, drug | MHY1485 | MedChemExpress | Cat# HY-B0795 | |
| Chemical compound, drug | (2-Hydroxypropyl)-beta-cyclodextrin | Beyotime | Cat# ST2114 | Solubilizer for oral gavage in animal studies |
| Chemical compound, drug | Lipofectamine RNAiMAX | Invitrogen | Cat# 13778030 | |
| Chemical compound, drug | Tozasertib | TargetMol | Cat# T2509 | |
| Chemical compound, drug | NaHCO3 | Sigma-Aldrich | Cat# S5761 | Solubilizer for oral gavage in animal studies |
| Chemical compound, drug | Alisertib | Apexbio | Cat# A4110 | Aurora kinase A inhibitor; for in vivo administration |
| Chemical compound, drug | Alisertib | TargetMol | Cat# T2241 | Aurora kinase A inhibitor; for in vitro cell culture |
| Cell line (Homo sapiens) | THP-1 | Cell Bank of the Chinese Academy of Sciences (Shanghai, China) | CSTR:19375.09.3101 HUMSCSP567 | |
| Cell line (M. musculus) | MC38 | Cell Bank of the Chinese Academy of Sciences (Shanghai, China) | CSTR:19375.09.3101 MOUSCSP5431 | |
| Cell line (M. musculus) | J774A.1 | Cell Bank of the Chinese Academy of Sciences (Shanghai, China) | CSTR:19375.09.3101 MOUSCSP5224 | |
| Sequence-based reagent (M. musculus) | siRNA: nontargeting control | This paper, synthesis by RuiBiotech | 5′-UUCUCCGAACGUGUCACGUTT-3′ | |
| Sequence-based reagent (M. musculus) | siRNA to AurA#1 | This paper, synthesis by RuiBiotech | 5′-CGAGCAGAGAACAGCUACUUATT-3′ | |
| Sequence-based reagent (M. musculus) | siRNA to AurA#2 | This paper, synthesis by RuiBiotech | 5′-GCACCCUUGGAUCAAAGCUAATT-3′ | |
| Sequence-based reagent (M. musculus) | siRNA to GNMT#1 | This paper, synthesis by RuiBiotech | 5′-GGACAAAGAUGUGCUUUCATT-3′ | |
| Sequence-based reagent (M. musculus) | siRNA to GNMT#2 | This paper, synthesis by RuiBiotech | 5′-CGUCAGUACUGACAGUCAATT-3′ | |
| Sequence-based reagent (M. musculus) | siRNA to FOXO3#1 | This paper, synthesis by RuiBiotech | 5′-CGGCAACCAGACACUCCAATT-3′ | |
| Sequence-based reagent (M. musculus) | siRNA to FOXO3#2 | This paper, synthesis by RuiBiotech | 5′-CUGUAUUCAGCUAGUGCAATT-3′ | |
| Sequence-based reagent (M. musculus) | Il6-F | This paper, synthesis by RuiBiotech | PCR primers | 5′-TAGTCCTTCCTACCCCAATTTCC-3′ |
| Sequence-based reagent (M. musculus) | Il6-R | This paper, synthesis by RuiBiotech | PCR primers | 5′-TTGGTCCTTAGCCACTCCTTC-3′ |
| Sequence-based reagent (M. musculus) | Tnf-F | This paper, synthesis by RuiBiotech | PCR primers | 5′- CCCTCACACTCAGATCATCTTCT-3′ |
| Sequence-based reagent (M. musculus) | Tnf-R | This paper, synthesis by RuiBiotech | PCR primers | 5′- GCTACGACGTGGGCTACAG-3′ |
| Sequence-based reagent (M. musculus) | Actb-F | This paper, synthesis by RuiBiotech | PCR primers | 5′-AGAGGGAAATCGTGCGTGAC-3′ |
| Sequence-based reagent (M. musculus) | Actb-R | This paper, synthesis by RuiBiotech | PCR primers | 5′-CAATAGTGATGACCTGGCCGT-3′ |
| Sequence-based reagent (M. musculus) | Il6-F | This paper, synthesis by RuiBiotech | Chip-PCR primers | 5′-TCGATGCTAAACGACGTCACA-3′ |
| Sequence-based reagent (M. musculus) | Il6-R | This paper, synthesis by RuiBiotech | Chip-PCR primers | 5′-CGTCTTTCAGTCACTATTAGGAGTC-3′ |
| Sequence-based reagent (M. musculus) | Tnf-F | This paper, synthesis by RuiBiotech | Chip-PCR primers | 5′-TGGCTAGACATCCACAGGGA-3′ |
| Sequence-based reagent (M. musculus) | Tnf-R | This paper, synthesis by RuiBiotech | Chip-PCR primers | 5′-AAGTTTCTCCCCCAACGCAA-3′ |
| Commercial assay or kit | CCK8 | Yeasen | Cat# 40203ES60 | |
| Commercial assay or kit | Protease Inhibitor Cocktail | Selleck | Cat# B14001 | |
| Antibody | Rabbit monoclonal anti-Aurora A | Abcam | Cat# ab108353, RRID:AB_10865712 | IB (1:1000) |
| Antibody | Mouse monoclonal anti-β-actin | Sigma-Aldrich | Cat# A1978, RRID:AB_476692 | IB (1:10000) |
| Antibody | Rabbit polyclonal anti-Phospho Aurora A (Thr288) | Invitrogen | Cat# 44–1210 G | IB (1:500) |
| Antibody | Rabbit polyclonal anti-FOXO3A | Abcam | Cat# ab23683, RRID:AB_732424 | IB (1:500), IF (1:200) |
| Antibody | Rabbit polyclonal anti-Phospho FOXO3 (Ser315) | Proteintech | Cat# 28755–1-AP, RRID:AB_2881210 | IB (1:500) |
| Antibody | Rabbit polyclonal anti-GNMT | Invitrogen | Cat# PA5-100018, RRID:AB_2815548 | IB (1:500), IF (1:200) |
| Antibody | Rabbit monoclonal anti-H3K4me3 | Abcam | Cat# ab213224, RRID:AB_2923013 | IB (1:3000), IP (1:500) |
| Antibody | Rabbit monoclonal anti-H3K4me1 | Abcam | Cat# ab8895, RRID:AB_306847 | IB (1:3000) |
| Antibody | Rabbit polyclonal anti-H3K9me3 | Abcam | Cat# ab8898, RRID:AB_306848 | IB (1:3000) |
| Antibody | Rabbit monoclonal anti-H3K36me3 | Abcam | Cat# ab282572, RRID:AB_3095544 | IB (1:3000), IP (1:500) |
| Antibody | Rabbit monoclonal anti-H3K27me3 | Cell Signaling Technology | Cat# 9733, RRID:AB_2616029 | IB (1:3000) |
| Antibody | Mouse monoclonal anti-H3 | Cell Signaling Technology | Cat# 14269, RRID:AB_2756816 | IB (1:5000) |
| Antibody | Mouse monoclonal anti-AKT | Cell Signaling Technology | Cat# 2920, RRID:AB_1147620 | IB (1:1000) |
| Antibody | Rabbit monoclonal anti- Phospho-Akt (Ser473) | Cell Signaling Technology | Cat# 4060, RRID:AB_2315049 | IB (1:1000) |
| Antibody | Rabbit polyclonal anti-mTOR | Cell Signaling Technology | Cat# 2972, RRID:AB_330978 | IB (1:1000) |
| Antibody | Rabbit monoclonal anti-Phospho-mTOR (Ser2448) | Cell Signaling Technology | Cat# 5536, RRID:AB_1069155 | IB (1:1000) |
| Antibody | Rabbit polyclonal anti-p70 S6 Kinase | Cell Signaling Technology | Cat# 9202, RRID:AB_331676 | IB (1:1000) |
| Antibody | Rabbit polyclonal anti-Phospho-p70 S6 Kinase (Thr389) | Cell Signaling Technology | Cat# 9205, RRID:AB_330944 | IB (1:1000) |
| Antibody | Rabbit monoclonal anti-S6 | Cell Signaling Technology | Cat# 2217, RRID:AB_331355 | IB (1:1000) |
| Antibody | Rabbit monoclonal anti-Phospho-S6 (Ser235/236) | Cell Signaling Technology | Cat# 4858, RRID:AB_916156 | IB (1:1000), FACS (1:200) |
| Antibody | Rat monoclonal anti-F4/80 | Abcam | Cat# ab90247, RRID:AB_10712189 | IF (1:200) |
| Antibody | Rat monoclonal anti-mouse CD45-BV421 (Brilliant Violet 421) | BD Biosciences | Cat# 563890, RRID:AB_2651151 | FACS (1:100) |
| Antibody | Rat monoclonal anti-mouse CD11b (PerCP-Cyanine5.5) | Thermo Fisher Scientific | Cat# 45-0112-82, RRID:AB_953558 | FACS (1:100) |
| Antibody | Rat monoclonal anti-mouse F4/80 (FITC) | Thermo Fisher Scientific | Cat# 11-4801-82, RRID:AB_2637191 | FACS (1:100) |
| Antibody | Goat anti-Rabbit, Alexa Fluor 488 | Thermo Fisher Scientific | Cat# A-11008, RRID:AB_143165 | IF (1:200) |
| Antibody | Anti-rabbit IgG, Alexa Fluor 555 | Cell Signaling Technology | Cat# 4413, RRID:AB_10694110 | FACS (1:200) |
| Antibody | Anti-rat IgG, Alexa Fluor 555 | Cell Signaling Technology | Cat# 4417, RRID:AB_10696896 | IF (1:200) |
| Commercial assay or kit | IL-6 Mouse Uncoated ELISA Kit | Invitrogen | Cat# 88-7064-88 | |
| Commercial assay or kit | TNF-α Mouse Uncoated ELISA Kit | Invitrogen | Cat# 88-7324-88 | |
| Commercial assay or kit | IL-6 Human Uncoated ELISA Kit | Invitrogen | Cat# 88-7066-88 | |
| Commercial assay or kit | 12p70 Mouse Uncoated ELISA Kit | Invitrogen | Cat# 88-7121-88 | |
| Commercial assay or kit | TGF Mouse Uncoated ELISA Kit | Invitrogen | Cat# 88-8350-88 | |
| Commercial assay or kit | IL-1β Mouse Uncoated ELISA Kit | Invitrogen | Cat# 88-7013-88 | |
| Commercial assay or kit | Glutathione Assay Kit | BioAssay Systems | Cat# DIGT-250 | |
| Commercial assay or kit | Cell Counting Kit (CCK-8) | Yeasen | Cat# 40203ES60 | |
| Commercial assay or kit | Phosflow Lyse/Fix Buffer | BD Biosciences | Cat# 558049 | |
| Commercial assay or kit | BD Perm/Wash | BD Biosciences | Cat# 554723 | |
| Commercial assay or kit | Agilent Seahorse XF Glycolysis Stress Test Kit | Agilent | Cat# 103020–100 | |
| Commercial assay or kit | High-Sensitivity Open Chromatin Profile Kit 2.0 (for Illumina) | Novoprotein | Cat# N248 | |
| Software, algorithm | Fiji | National Institutes of Health | RRID:SCR_002285 | Image analysis |
| Software, algorithm | GraphPad Prism v8.0 | GraphPad Software | RRID:SCR_002798 | Statistical analysis |
| Software, algorithm | FlowJo software v10.8.1 | Tree Star | RRID:SCR_008520 | |
| Other | C57BL/6 Mice | Vital River Laboratory Animal Technology | ||
| Other | CD45.1 B6/SJL Mice | Shanghai Model Organisms |
Additional files
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Supplementary file 1
List of compounds in the epigenetic drug library.
- https://cdn.elifesciences.org/articles/104138/elife-104138-supp1-v1.xlsx
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MDAR checklist
- https://cdn.elifesciences.org/articles/104138/elife-104138-mdarchecklist1-v1.pdf
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Source data 1
Statistical sources data used to generate main figures and figure supplements.
- https://cdn.elifesciences.org/articles/104138/elife-104138-data1-v1.xlsx
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Aurora kinase A promotes trained immunity via regulation of endogenous S-adenosylmethionine metabolism
eLife 14:RP104138.
https://doi.org/10.7554/eLife.104138.3
