Thioredoxin-1 distinctly promotes NF-κB target DNA binding and NLRP3 inflammasome activation independently of Txnip
- Institute of Molecular Health Sciences, ETH Zürich, Switzerland
Figures
Figure 1 with 3 supplements
The Trx1 system is largely dispensable for the development and homeostatic maintenance of myeloid cells.
(A–E) Txnrd1fl/fl;Rosa26-CreERT2 mice and Txnrd1fl/fl littermates were injected with TAM to delete the Txnrd1 gene and were analyzed by flow cytometry 2 weeks later. Depicted are the total numbers …
Figure 1—figure supplement 1
Efficiency of Txnrd1 gene deletion.
(A, B) Txnrd1fl/fl;Rosa26-CreERT2 mice and Txnrd1fl/fl littermates were injected with TAM to delete the Txnrd1 gene and were analyzed by flow cytometry 2 weeks later. Depicted is the analysis of …
Figure 1—figure supplement 2
Gating strategies for the analysis of distinct myeloid-cell populations in vivo.
(A–D) Shown are the gating strategies utilized for the analysis of the indicated myeloid-cell populations in the bone marrow (BM; A), blood (B), lungs (C), and spleen (D). Eo, eosinophils; Neutro, …
Figure 1—figure supplement 3
The Trx1 system is largely dispensable for the maintenance and homeostasis of myeloid cells in mixed-bone marrow chimera settings.
(A–C) Lethally irradiated WT mice (CD45.1+CD45.2+) were reconstituted with a 1:1 mixture of WT (CD45.1+) and TAM-treated Txnrd1fl/fl;Rosa26-CreERT2 (CD45.2+) bone marrows (or Txnrd1fl/fl as …
Figure 2 with 2 supplements
The GSH/Grx system sustains the development and maintenance of Txnrd1-deficient myeloid cells.
(A–I) Lethally irradiated WT mice (CD45.1+CD45.2+) were reconstituted with a 1:1 mixture of WT (CD45.1+) and Txnrd1fl/fl;Rosa26-CreERT2 (CD45.2+) bone marrows. After reconstitution, mice were …
Figure 2—figure supplement 1
The GSH/Grx system compensates for the absence of the Trx1 pathway in Txnrd1-deficient monocytes.
Lethally irradiated WT mice (CD45.1+CD45.2+) were reconstituted with a 1:1 mixture of WT (CD45.1+) and Txnrd1fl/fl;Rosa26-CreERT2 (CD45.2+) bone marrows. After reconstitution, mice were injected …
Figure 2—figure supplement 2
BSO administration does not affect the maintenance and homeostasis of WT myeloid cells.
(A, B) WT mice were treated with BSO in the drinking water and analyzed 2 weeks later (n = 4–5 mice). Shown are the total counts of the indicated myeloid populations in the bone marrow (BM; A) and …
Figure 3
The Trx1 system is required in the bone marrow during emergency hematopoiesis.
(A–E) TAM-treated Txnrd1fl/fl;Rosa26-CreERT2 mice (or Txnrd1fl/fl littermate controls) were intraperitoneally injected with 35 μg LPS in a 48 hr interval and sacrificed 24 hr later for analysis (n = …
Figure 4 with 2 supplements
Txnrd1-deficient BMDCs undergo defective transcription of pro-inflammatory cytokines.
(A–J) Txnrd1fl/fl;Rosa26-CreERT2 mice and Txnrd1fl/fl littermates were injected with TAM to delete the Txnrd1 gene. Bone marrow cells were differentiated with GM-CSF to obtain BMDCs. (A) Analysis of …
Figure 4—figure supplement 1
Txnrd1-deficient BMDCs display an impaired secretion of TNF-α.
Txnrd1fl/fl;Rosa26-CreERT2 mice and Txnrd1fl/fl littermates were injected with TAM to delete the Txnrd1 gene, and bone marrow cells were differentiated with GM-CSF to obtain BMDCs. BMDCs were then …
Figure 4—figure supplement 2
Txnrd1-deficient BMDCs display an impaired secretion of IL-1β.
Txnrd1fl/fl;Rosa26-CreERT2 mice and Txnrd1fl/fl littermates were injected with TAM to delete the Txnrd1 gene, and bone marrow cells were differentiated with GM-CSF to obtain BMDCs. BMDCs were then …
Figure 5 with 3 supplements
The Trx1 system positively regulates the binding activity of NF-κB p65 to the DNA in BMDCs.
(A–E) Txnrd1fl/fl;Rosa26-CreERT2 mice and Txnrd1fl/fl littermates were injected with TAM to delete the Txnrd1 gene, and bone marrow cells were differentiated with GM-CSF to obtain BMDCs. (A) BMDCs …
Figure 5—figure supplement 1
Txnrd1 deficiency does not affect TLR signaling.
Txnrd1fl/fl;Rosa26-CreERT2 mice and Txnrd1fl/fl littermates were injected with TAM to delete the Txnrd1 gene, and bone marrow cells were differentiated with GM-CSF to obtain BMDCs. BMDCs were then …
Figure 5—figure supplement 2
Magnification of the microscopy images of Figure 5C.
The highlighted areas in anti-NF-κB channels in Figure 5C are shown here in higher magnification.
Figure 5—figure supplement 3
Antioxidant supplementation does not rescue IL-12p40 cytokine production in Txnrd1-deficient BMDCs.
Txnrd1fl/fl;Rosa26-CreERT2 mice and Txnrd1fl/fl littermates were injected with TAM to delete the Txnrd1 gene, and bone marrow cells were differentiated with GM-CSF to obtain BMDCs. (A) …
Figure 6 with 3 supplements
The Trx1 system is critical for proper M1 polarization but dispensable for promoting NF-κB binding to target DNA in BMDMs.
(A–P) Bone marrows from Txnrd1fl/fl;Rosa26-CreERT2 mice and control Txnrd1fl/fl littermates treated with TAM were differentiated with M-CSF to obtain BMDMs. (A–H) BMDMs were polarized toward M1 and …
Figure 6—figure supplement 1
The Trx1 system promotes M1 but not M2 macrophage polarization.
(A–K) Bone marrows from Txnrd1fl/fl;Rosa26-CreERT2 mice and control Txnrd1fl/fl littermates treated with TAM were differentiated with M-CSF to obtain BMDMs. (A) Shown are the expression levels of Txn…
Figure 6—figure supplement 2
Txnrd1 deficiency does not impair TNF-α production.
Bone marrows from Txnrd1fl/fl;Rosa26-CreERT2 mice and control Txnrd1fl/fl littermates treated with TAM were differentiated with M-CSF to obtain BMDMs. BMDMs were then stimulated for 7 hr with CpG …
Figure 6—figure supplement 3
The GSH/Grx system compensates for the absence of the Trx1 pathway during IL-12p40 production by Txnrd1-deficient BMDMs.
Bone marrows from Txnrd1fl/fl;Rosa26-CreERT2 mice and control Txnrd1fl/fl littermates treated with TAM were differentiated with M-CSF to obtain BMDMs. (A) BMDMs were treated overnight with BSO (3 …
Figure 7 with 5 supplements
Trx1 but not Txnip promotes NLRP3-dependent IL-1β responses.
(A–C, F–L) Bone marrows from the indicated genotypes (from TAM-treated Txnrd1fl/fl;Rosa26-CreERT2 and control Txnrd1fl/fl littermates or from Txnipfl/fl;Vav1-Cre and Txnipfl/fl control mice) were …
Figure 7—figure supplement 1
The Trx1 system is required for NLRP3 inflammasome responses.
(A, B) Bone marrows from Txnrd1fl/fl;Rosa26-CreERT2 mice (KO) and control Txnrd1fl/fl littermates (WT) treated with TAM were differentiated with M-CSF to obtain BMDMs. (A) Txnrd1-deficient and …
Figure 7—figure supplement 2
Catalase treatment restores IL-1β production in the absence of the Trx1 system.
Bone marrows from TAM-treated Txnrd1fl/fl;Rosa26-CreERT2 and control Txnrd1fl/fl littermates were differentiated with M-CSF to obtain BMDMs. BMDMs were then treated overnight with the antioxidant …
Figure 7—figure supplement 3
Catalase treatment does not affect transcriptional IL-1β levels.
Bone marrows from Txnrd1fl/fl;Rosa26-CreERT2 mice and control Txnrd1fl/fl littermates treated with TAM were differentiated with M-CSF to obtain BMDMs. (A, B) BMDMs were treated overnight with the …
Figure 7—figure supplement 4
Supplementation with the indicated antioxidants does not restore IL-1β production in the absence of the Trx1 system in BMDMs.
Bone marrows from Txnrd1fl/fl;Rosa26-CreERT2 mice and control Txnrd1fl/fl littermates treated with TAM were differentiated with M-CSF to obtain BMDMs. (A, B) Txnrd1-deficient and control BMDMs were …
Figure 7—figure supplement 5
Txnip deficiency does not affect cytokine secretion in BMDMs.
Bone marrows from Txnipfl/fl;Vav1-Cre mice and Txnipfl/fl control littermates were differentiated with M-CSF to obtain BMDMs. (A) Depicted is the analysis of Txnip mRNA in BMDMs determined by RT-PCR …
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Genetic reagent (M. musculus) | C57BL/6J | The Jackson Laboratory | Cat#JAX:000664; RRID: IMSR_JAX:000664 | |
| Genetic reagent (M. musculus) | B6 Ptprca (CD45.1) | The Jackson Laboratory | Cat#JAX:002014, RRID:IMSR_JAX:002014 | |
| Genetic reagent (M. musculus) | Txnrd1fl/fl | PMID: 15713651 | Kindly provided by Marcus Conrad | |
| Genetic reagent (M. musculus) | Rosa26-CreERT2 | PMID: 17456738 | ||
| Genetic reagent (M. musculus) | Txnipfl/fl | The Jackson Laboratory | Cat# JAX:016847, RRID:IMSR_JAX:016847 | |
| Genetic reagent (M. musculus) | Vav1-Cre | PMID: 12548562 | ||
| Antibody | Anti-B220 PerCP-conjugated (rat, monoclonal) | Biolegend | Cat# 103234, RRID:AB_893353 | Flow cytometry, cell surface (1:400) |
| Antibody | Anti-CD103 FITC-conjugated (armenian hamster, monoclonal) | Biolegend | Cat# 121420, RRID:AB_10714791 | Flow cytometry, cell surface (1:200) |
| Antibody | Anti-CD11b BV605-conjugated (rat, monoclonal) | Biolegend | Cat# 101237, RRID:AB_11126744 | Flow cytometry, cell surface (1:2000) |
| Antibody | Anti-CD11b PerCP-Cy5.5-conjugated (rat, monoclonal) | Biolegend | Cat# 101228, RRID:AB_893232 | Flow cytometry, cell surface (1:1000) |
| Antibody | Anti-CD11c APC-conjugated (armenian hamster, monoclonal) | Thermo Fisher Scientific | Cat# 17-0114-82, RRID:AB_469346 | Flow cytometry, cell surface (1:300) |
| Antibody | Anti-CD11c BV605-conjugated (armenian hamster, monoclonal) | Biolegend | Cat# 117333, RRID:AB_11204262 | Flow cytometry, cell surface (1:1000) |
| Antibody | Anti-CD11c PE-Cy7-conjugated (armenian hamster, monoclonal) | Biolegend | Cat# 117318, RRID:AB_493568 | Flow cytometry, cell surface (1:500) |
| Antibody | Anti-CD19 BV650-conjugated (rat, monoclonal) | Biolegend | Cat# 115541, RRID:AB_11204087 | Flow cytometry, cell surface (1:500) |
| Antibody | Anti-CD206 PerCP-Cy5.5-conjugated (rat, monoclonal) | Biolegend | Cat# 141716, RRID:AB_2561992 | Flow cytometry, intracellular (1:400) |
| Antibody | Anti-PD-L2 PE-conjugated (rat, monoclonal) | Biolegend | Cat# 107206, RRID:AB_2162011 | Flow cytometry, cell surface (1:400) |
| Antibody | Anti-CD301b PE-Cy7-conjugated (rat, monoclonal) | Biolegend | Cat# 146808, RRID:AB_2563390 | Flow cytometry, cell surface (1:600) |
| Antibody | Anti-CD4 PerCP-Cy5.5-conjugated (rat, monoclonal) | Biolegend | Cat# 100540, RRID:AB_893326 | Flow cytometry, cell surface (1:500) |
| Antibody | Anti-CD45 BV785-conjugated (rat, monoclonal) | Biolegend | Cat# 103149, RRID:AB_2564590 | Flow cytometry, cell surface (1:1000) |
| Antibody | Anti-CD45.1 APC-conjugated (mouse, monoclonal) | Biolegend | Cat# 110714, RRID:AB_313503 | Flow cytometry, cell surface (1:100) |
| Antibody | Anti-CD45.1 biotin-conjugated (mouse, monoclonal) | BD Biosciences | Cat# 553774, RRID:AB_395042 | Flow cytometry, cell surface (1:200) |
| Antibody | Anti-CD45.2 APC-conjugated (mouse, monoclonal) | Thermo Fisher Scientific | Cat# 17-0454-82, RRID:AB_469400 | Flow cytometry, cell surface (1:100) |
| Antibody | Anti-CD45.2 FITC-conjugated (mouse, monoclonal) | Thermo Fisher Scientific | Cat# 11-0454-82, RRID:AB_465061 | Flow cytometry, cell surface (1:300) |
| Antibody | Anti-CD8 PE-Cy7-conjugated (rat, monoclonal) | Biolegend | Cat# 100722, RRID:AB_312761 | Flow cytometry, cell surface (1:1000) |
| Antibody | Anti-F4/80 AF488-conjugated (rat, monoclonal) | Biolegend | Cat# 123120, RRID:AB_893479 | Flow cytometry, cell surface (1:400) |
| Antibody | Anti-F4/80 BV421-conjugated (rat, monoclonal) | Biolegend | Cat# 123131, RRID:AB_10901171 | Flow cytometry, cell surface (1:300) |
| Antibody | Anti-IFN-γ APC-conjugated (rat, monoclonal) | Biolegend | Cat# 505810, RRID:AB_315404 | Flow cytometry, intracellular (1:4000) |
| Antibody | Anti-Ly-6G BV421-conjugated (rat, monoclonal) | Biolegend | Cat# 127627, RRID:AB_10897944 | Flow cytometry, cell surface (1:800) |
| Antibody | Anti-Ly-6G PerCP-Cy5.5-conjugated (rat, monoclonal) | Biolegend | Cat# 127616, RRID:AB_1877271 | Flow cytometry, cell surface (1:600) |
| Antibody | Anti-Ly-6C APC-Cy7-conjugated (rat, monoclonal) | Biolegend | Cat# 128026, RRID:AB_10640120 | Flow cytometry, cell surface (1:2000) |
| Antibody | Anti-Ly-6C PE-Cy7-conjugated (rat, monoclonal) | Biolegend | Cat# 128018, RRID:AB_1732082 | Flow cytometry, cell surface (1:2000) |
| Antibody | Anti-MHCII PE-Cy7-conjugated (rat, monoclonal) | Biolegend | Cat# 107635, RRID:AB_2561397 | Flow cytometry, cell surface (1:400) |
| Antibody | Anti-Nos2 AF647-conjugated (mouse, monoclonal) | Santa Cruz | Cat# sc-7271, RRID:AB_627810 | Flow cytometry, intracellular (1:100) |
| Antibody | Anti-Rabbit IgG(H+L) FITC-conjugated (goat, polyclonal) | SouthernBiotech | Cat# 4055–02, RRID:AB_2795979 | Flow cytometry, Microscopy (1:500) |
| Antibody | Anti-Relmα (rabbit, polyclonal) | PeproTech | Cat# 500-P214bt-50ug, RRID:AB_1268707 | Flow cytometry, intracellular (1:100) |
| Antibody | Anti-Siglec-F PE-conjugated (rat, monoclonal) | BD Biosciences | Cat# 552126, RRID:AB_394341 | Flow cytometry, cell surface (1:300) |
| Antibody | Anti-TCRβ APC-conjugated (armenian hamster, monoclonal) | Biolegend | Cat# 109212, RRID:AB_313435 | Flow cytometry, cell surface (1:500) |
| Antibody | Anti-NF-κB p65 (rabbit, polyclonal) | Biolegend | Cat# 622604, RRID:AB_2728469 | ChIP, 3 μg antibody for 40 μg chromatin |
| Antibody | CD4 MicroBeads | Miltenyi Biotec | Cat# 130-049-201, RRID:AB_2722753 | MACS enrichment |
| Antibody | CD11b MicroBeads | Miltenyi Biotec | Cat# 130-049-601, | MACS enrichment |
| Antibody | Anti-IL-1β (armenian hamster, monoclonal) | Thermo Fisher Scientific | Cat# 14-7012-81, RRID:AB_468396 | ELISA, coating antibody (1:150) |
| Antibody | Anti-IL-1β biotin-conjugated (rabbit, polyclonal) | Thermo Fisher Scientific | Cat# 13-7112-85, RRID:AB_466925 | ELISA, detection antibody (1:150) |
| Antibody | Anti-IL-12/IL-23 p40 (rat, monoclonal) | Thermo Fisher Scientific | Cat# 14-7125-81, RRID:AB_468444 | ELISA, coating antibody (1:250) |
| Antibody | Anti-IL-12/IL-23 p40 biotin-conjugated (rat, monoclonal) | Thermo Fisher Scientific | Cat# 13-7123-81, RRID:AB_466928 | ELISA, detection antibody (1:1000) |
| Antibody | Anti-IL-6 (rat, monoclonal) | Thermo Fisher Scientific | Cat# 16-7061-81, RRID:AB_469216 | ELISA, coating antibody (1:250) |
| Antibody | Anti-IL-6 biotin-conjugated (rat, monoclonal) | Thermo Fisher Scientific | Cat# 36-7062-85, RRID:AB_469761 | ELISA, detection antibody (1:500) |
| Antibody | Anti-TNF-α (rat, monoclonal) | BD Biosciences | Cat# 551225, RRID:AB_394102 | ELISA, coating antibody (1:100) |
| Antibody | Anti-TNF-α biotin-conjugated (rabbit, polyclonal) | Thermo Fisher Scientific | Cat# 13-7341-81, RRID:AB_466950 | ELISA, detection antibody (1:150) |
| Antibody | Anti-phospho-Erk1/2 (rabbit, polyclonal) | Cell Signaling Technology | Cat# 9101, RRID:AB_331646 | WB (1:1000) |
| Antibody | Anti-Erk1/2 (rabbit, polyclonal) | Cell Signaling Technology | Cat# 9102, RRID:AB_330744 | WB (1:1000) |
| Antibody | Anti-phospho-IκB-α, (mouse, monoclonal) | Cell Signaling Technology | Cat# 9246, RRID:AB_2267145 | WB (1:1000) |
| Antibody | Anti-IκB-α, (rabbit, polyclonal) | Cell Signaling Technology | Cat# 9242, RRID:AB_331623 | WB (1:1000) |
| Antibody | Anti-NF-κB p65 (rabbit, polyclonal) | Biolegend | Cat# 622602, RRID:AB_315956 | WB (1:2000) |
| Antibody | Anti-IL-1β (mouse, polyclonal) | R and D Systems | Cat# AF-401-NA, RRID:AB_416684 | WB (1:800), ‘Mouse IL-1 beta/IL-1F2 antibody’ |
| Antibody | Anti-caspase-1 p10 (rabbit, polyclonal) | Santa Cruz | Cat# sc-514, RRID:AB_2068895 | WB (1:200) |
| Antibody | Anti-β-actin peroxidase- conjugated (mouse, monoclonal) | Sigma-Aldrich | Cat# A3854, RRID:AB_262011 | WB (1:50000) |
| Antibody | Anti-NF-κB p65 (rabbit, polyclonal) | Santa Cruz | Cat# sc-7151, RRID:AB_650213 | Microscopy (1:250) |
| Peptide, recombinant protein | IFN-γ | PeproTech | Cat# 315–05 | M1 polarization (50 ng/ml) |
| Peptide, recombinant protein | IL-4 | PeproTech | Cat# 214–14 | M2 polarization (20 ng/ml) |
| Peptide, recombinant protein | M-CSF | PeproTech | Cat# 315–02 | BMDM differentiation (20 ng/ml) |
| Peptide, recombinant protein | gp61-80 peptide (LCMV) | NeoMPS | Cat# SP990990 | Sequence: GLNGPDIYKGVYQFKSVEFD |
| Chemical compound, drug | Tamoxifen (TAM) | Sigma-Aldrich | Cat# T5648-1G | To delete Txnrd1 in Txnrd1fl/fl;Rosa26-CreERT2 mice |
| Chemical compound, drug | Lipopolysaccharides (LPS) | InvivoGen | Cat# tlrl-3pelps | TLR4 agonist |
| Chemical compound, drug | Imiquimod (R837) | Tocris Bioscience | Cat# 3700 | TLR7 agonist |
| Chemical compound, drug | CpG oligodeoxynucleotides | InvivoGen | Cat# tlrl-1826–1 | TLR9 agonist |
| Chemical compound, drug | Lipoteichoic (LTA) | InvivoGen | Cat# tlrl-slta | TLR2 agonist |
| Chemical compound, drug | Zymosan A | Sigma-Aldrich | Cat# Z4250-250MG | TLR2/Dectin-1 agonist |
| Chemical compound, drug | Nigericin (Nig) | Sigma-Aldrich | Cat# N7143 | NLRP3 inflammasome inducer |
| Chemical compound, drug | Adenosine triphosphate (ATP) | Sigma-Aldrich | Cat# A7699 | NLRP3 inflammasome inducer |
| Chemical compound, drug | Alu-Gel-S (Alum) | SERVA Electrophoresis | Cat# 12261.01 | NLRP3 inflammasome inducer |
| Chemical compound, drug | Monosodium urate (MSU) crystals | InvivoGen | Cat# tlrl-msu-25 | NLRP3 inflammasome inducer |
| Chemical compound, drug | DL-Buthionine-(S,R)-sulfoximine (BSO ) | Sigma-Aldrich | Cat# B2640-500MG | Glutathione synthesis inhibitor |
| Chemical compound, drug | Catalase-polyethylene glycol (Cat) | Sigma-Aldrich | Cat# C4963 | Antioxidant |
| Chemical compound, drug | L-Ascorbic acid | Sigma-Aldrich | Cat# A92902-25G | Antioxidant |
| Chemical compound, drug | Diphenyleneiodonium chloride (DPI) | Sigma-Aldrich | Cat# D2926-10MG | Antioxidant |
| Chemical compound, drug | DL-Dithiothreitol (DTT) | Sigma-Aldrich | Cat# 43815–5G | Antioxidant |
| Chemical compound, drug | N-Acetyl-L-Cysteine (NAC) | Sigma-Aldrich | Cat# A7250-10G | Antioxidant |
| Chemical compound, drug | DNase I | Sigma-Aldrich | Cat# 4716728001 | Tissue digestion (0.02 mg/ml) |
| Chemical compound, drug | Collagenase IV | Worthington | Cat# LS004189 | Tissue digestion (2 mg/ml) |
| Chemical compound, drug | Phorbol 12-myristate 13-acetate (PMA) | Sigma-Aldrich | Cat# P-8139 | T-cell restimulation (10−7 mol/L) |
| Chemical compound, drug | Ionomycin | Sigma-Aldrich | Cat# I-0634 | T-cell restimulation (1 μg/ml) |
| Chemical compound, drug | Monensin | Sigma-Aldrich | Cat# M5273 | Used at 2 μg/ml during T-cell restimulation |
| Commercial assay or kit | NF-κB p65 Transcription Factor Assay Kit | Abcam | Cat# ab133112 | |
| Commercial assay or kit | Nuclear Extraction Kit | Abcam | Cat# ab113474 | |
| Commercial assay or kit | Glutathione Assay Kit | Sigma-Aldrich | Cat# CS0260 | |
| Commercial assay or kit | Pierce BCA Protein Assay Kit | Thermo Fisher Scientific | Cat# 23225 | |
| Commercial assay or kit | Pierce LDH Cytotoxicity Assay Kit | Thermo Fisher Scientific | Cat# 88954 | |
| Software, algorithm | FlowJo Software (version 10.4.2) | Three Star | https://www.flowjo.com/ | |
| Software, algorithm | ImageJ (for image analysis) | NIH | https://imagej.nih.gov/ij/ | |
| Software, algorithm | Prism 8 (version 8.0.0) | GraphPad Software | https://www.graphpad.com/scientific-software/prism/ | |
| Other | DAPI | Sigma-Aldrich | Cat# D9542 | Microscopy, nuclei visualization (1:5000) |
| Other | Phalloidin, AF-647-conjugated | Thermo Fisher Scientific | Cat# A22287 | Microscopy, actin staining (1:1000) |
| Other | eFluor 780 | Thermo Fisher Scientific | Cat# 65-0865-14 | Live/Dead staining (1:2000) |
| Other | Zombie Aqua | Biolegend | Cat# 423101 | Live/Dead staining (1:400) |
| Other | Zombie Red | Biolegend | Cat# 423109 | Live/Dead staining (1:800) |
| Other | CM-H2DCFDA | Thermo Fisher Scientific | Cat# C6827 | General oxidative stress indicator (1:1000) |
| Other | Annexin-V APC-conjugated | Thermo Fisher Scientific | Cat# 88-8007-74 | Apoptosis detection (1:50) |
| Other | Streptavidin BV711-conjugated | BD Biosciences | Cat# 563262 | (1:1000) |
| Other | Dynabeads Protein G | Thermo Fisher Scientific | Cat# 10004D | For the ChIP experiment |
Additional files
-
Supplementary file 1
Sequences of primers used for RT-PCR.
- https://cdn.elifesciences.org/articles/53627/elife-53627-supp1-v2.docx
-
Transparent reporting form
- https://cdn.elifesciences.org/articles/53627/elife-53627-transrepform-v2.docx
