Cellular acidosis triggers human MondoA transcriptional activity by driving mitochondrial ATP production

  1. Blake R Wilde
  2. Zhizhou Ye
  3. Tian-Yeh Lim
  4. Donald E Ayer  Is a corresponding author
  1. University of Utah, United States
8 figures, 1 table and 1 additional file

Figures

Figure 1 with 3 supplements
Acidosis drives MondoA transcriptional activity.

(A) TXNIP mRNA levels in murine embryonic fibroblasts (MEFs) following treatment with HBSS for the indicated times. (B) Glucose uptake was determined by quantifying the rate of 3H-2-deoxyglucose uptake in MEFs following a 4 hr treatment with HBSS. (C) TXNIP mRNA levels from MEFs treated for 4 hr with DMEM, HBSS and HBSS supplemented with sodium bicarbonate to the amount in DMEM (3.7 g/L). (D) CRISPR/Cas9 was used to disrupt expression of MondoA in HeLa cells. Amounts of the indicated proteins in HeLa and HeLa:MondoA-KO cells were determined using immunoblotting. Consistent with our previous findings TXNIP expression was highly dependent on MondoA. TXNIP mRNA levels from HeLa and HeLa:MondoA-KO cells following a 4 hr treatment with DMEMAcidic. (E) TXNIP mRNA levels in HEK-293T, HeLa and HepG2 cells following 4 hr treatments with DMEMAcidic. In A, C, D and E, TXNIP mRNA levels were determined by reverse transcriptase-quantitative PCR (RT-qPCR). ***p<0.001; ****p<0.0001; ns – not significant.

https://doi.org/10.7554/eLife.40199.002
Figure 1—figure supplement 1
TXNIP expression correlates with genes that regulate intracellular pH.

(A) Heatmaps showing the expression of TXNIP mRNA compared to MCT4 (breast cancer), MCT1 (lung cancer) and NHE1 (brain cancer). All expression data was collected from TCGA. Spearman and Pearson correlation statistics are reported as r and ρ, respectively. (B) An acidosis gene signature was determined for the 2016 METABRIC breast cancer dataset. These scores were compared to TXNIP expression from the dataset and correlation statistics were performed. Heatmaps depicting the expression of TXNIP mRNA compared to MCT4, MCT1 and NHE1 for normal (C) skin and (D) muscle tissues. All expression data was collected from GTEx. Spearman and Pearson correlation statistics are reported as r and ρ, respectively.

https://doi.org/10.7554/eLife.40199.003
Figure 1—figure supplement 2
Acidosis drives MondoA transcriptional activity.

(A) TXNIP and MondoA protein levels in MEFs treated with HBSS for the indicated times was determined by immunoblotting. (B) TXNIP protein levels of MEFs following 4 hr treatments with DMEM, HBSS and HBSS supplemented with sodium bicarbonate to the amount in DMEM (3.7 g/L). (C) TXNIP protein levels in MEFs following 4 hr treatments with DMEM or HBSS containing the indicated amounts of sodium bicarbonate. Cells were also treated were with the identical medias with their pH clamped to pH 7.4 with 25 mM HEPES.

https://doi.org/10.7554/eLife.40199.004
Figure 1—figure supplement 3
Acidosis drives MondoA transcriptional activity.

(A) TXNIP and MondoA protein levels were determined from MondoA-knockout MEFs complemented with empty vector, wild-type MondoA or MondoA(I766P) following 4 hr HBSS treatments. (B) Schematic showing TXNIP-promoter luciferase reporter constructs. ChoREmut contains mutations in each half of the double E-box carbohydrate-responsive element ChoRE that are predicted to reduce or eliminate MondoA:Mlx binding. Luciferase activity was measured in MEFs transfected with each reporter construct following a 4 hr HBSS treatment. Luciferase activity and transfection efficiency was normalized to β-galactosidase (β-gal) activity encoded by a co-transfected plasmid expressing LacZ (C) Chromatin-immunoprecipitation using antibodies specific for MondoA or IgG was performed on chromatin preparations from MEFs treated for 4 hr with HBSS. ***p<0.001; ns – not significant.

https://doi.org/10.7554/eLife.40199.005
Acidosis-driven MondoA transcriptional activity depends on electron transport.

(A and B) TXNIP protein levels were determine by immunoblotting following a 4 hr HBSS treatment of MEFs in the presence of the indicated inhibitors. Cont; control, Chlor; Chloroquine (25 μM), Mon; monensin (5 μM), Met; metformin (1 mM), FCCP; carbonilcyanide p-triflouromethoxyphenylhydrazone (1 µM), Oligo; oligomycin (1 μM). (C–E), TXNIP mRNA levels were determine using RT-qPCR following 4 hr treatments with DMEMAcidic: (C) 143B osteosarcoma cells, (D) 143Bρ0 cells, and (E) 143Bρ0:WT-Cybrid cells complemented with wild type mitochondria. (F) Schematic depicting nuclear- and mitochondrial-DNA encoded components of the ETC. (G) TXNIP mRNA levels following 4 hr DMEMAcidic treatments of 143Bρ0:ΔATP6/ATP8-Cybrid cells expressing empty vector or nuclear encoded, mitochondrial-targeted ATP6 and ATP8. *p<0.05; **p<0.01; ****p<0.0001; ns – not significant.

https://doi.org/10.7554/eLife.40199.006
Figure 3 with 2 supplements
Acidosis drives the synthesis of mitochondrial ATP.

(A) Intracellular pH of HeLa cells was determined using BCECF-AM staining after a 4 hr treatment with DMEMAcidic. (B) Mitochondrial membrane potential of HeLa cells was determined by JC1 staining after a 1 hr treatment with DMEMAcidic. (C) Total cellular ATP levels in a lysate prepared from HeLa cells was determined using a luciferase-based assay after a 4 hr treatment with DMEMAcidic. (D) Mit-ATEAM, which is a mitochondrial-targeted ATP-biosensor, was used to determine the relative level of mtATP in HeLa cells treated with DMEMAcidic for the indicated times. Widefield microscopy was used to capture images in the FRET and CFP channels. Images were then used to analyze FRET and CFP signals in mitochondria in individual cells. FRET signal was normalized using CFP. (E) ATP5I mRNA levels in HeLa cells that expressed scrambled (siScrm, n = 1) or ATP5I-specific siRNA (siATP5I, n = 2) following a 4 hr DMEMAcidic treatment. (F) Mit-ATEAM was used to determine relative mtATP levels in HeLa cells transfected with siScrm or siATP5I and treated with DMEMAcidic for the indicated times. (G) TXNIP mRNA levels following a 4 hr DMEMAcidic treatment of HeLa cells that had been transfected siScrm or siATP5I. In (E and G), ATP5I and TXNIP mRNA levels were measured using RT-qPCR. **p<0.01; ****p<0.0001; ns – not significant.

https://doi.org/10.7554/eLife.40199.007
Figure 3—figure supplement 1
DMEMAcidic does not inhibit mTORC1 activity Immunoblotting was used to measure the levels of the indicated proteins following DMEMAcidic treatment of HeLa cells for the indicated times.

Levels of phosphorylated S6 (p–S6), which is a mTORC1 substrate, reflect mTORC1 activity.

https://doi.org/10.7554/eLife.40199.008
Figure 3—figure supplement 2
Acidosis drives synthesis of mitochondrial ATP.

(A) Confocal images at 60X of Mit-ATEAM expressed in HeLa cells. Shown are the CFP and FRET channels as well as the ratio of FRET to CFP (indicating mtATP). (B) Widefield image at 60X of ATEAM. CFP channel only is shown. (C) Confocal images of Mit-ATEAM, Mit-ATEAM(R122K/R126K), ATEAM and ATEAM(R122K/R126K) in HeLa cells. The R122K/R126K mutations block ATP binding providing a negative control for this experimental system. HeLa cells were treated with DMEMAcidic or CCCP (1 μM) for up to 8 hr and images captured at the indicated times. Images are pseudo-colored according to the FRET/CFP ratio. Notably, the FRET/CFP ratios for Mit-ATEAM(R122K/R126K) and ATEAM(R122K/R126K) was negligible compared to non-mutated constructs. Quantification of relative mitochondrial (D) and cytosolic (E) ATP.

https://doi.org/10.7554/eLife.40199.009
MondoA is activated by G6P produced by hexokinase utilization of mtATP.

(A) Cellular fractionation of BJ-Tert cells indicating mitochondrial localization of HK2, MondoA and Mlx. Succinate dehydrogenase A (SDHA) serves as a control for the mitochondria fraction. (B) Schematic illustrating how mtATP could contribute to MondoA transcriptional activity. As mtATP is exported from the mitochondria, it is used by as a substrate by mitochondria-bound HK2 to produce G6P, resulting in MondoA activation. (C) Heatmap and log2 fold-changes of glycolytic and TCA metabolites measured using GC-MS following a 4 hr treatment of HeLa cells with DMEMAcidic. TXNIP mRNA levels, as measured by RT-qPCR, following 4 hr DMEMAcidic treatments of HeLa cells and expressing pools of four siRNAs against (D) ANT2 and (E) HK2, or (F) expressing HK2 and HK2(D657A). Immunoblots validate knockdown of the indicated proteins. **p<0.01; ***p<0.001; ****p<0.0001; ns – not significant.

https://doi.org/10.7554/eLife.40199.010
MondoA senses G6P produced by mitochondrial-bound hexokinase.

(A) TXNIP mRNA levels in BJ-Tert cells expressing mVDAC1-GFP and mVDAC1(E72Q)-GFP and treated for 4 hr with DMEMAcidic. HK2 localization was also analyzed by cellular fractionation and densitometry was used to quantify the relative amount of HK2 localized to the mitochondria. HK2 was increasingly enriched in the cytoplasmic fraction in cells expressing mVDAC1(E72Q), suggesting that it had been displaced from mitochondria. (B) Schematic depicting the use of GFP(1-10) and GFP(11) to artificially tether mVDAC1 and HK2. HK2 localization was also analyzed by cellular fractionation and densitometry was used to quantify the relative amount of HK2 present in the mitochondrial fraction. (C) TXNIP mRNA levels in BJ-Tert cells treated for 4 hr with DMEMAcidic and expressing mVDAC1-GFP, mVDAC1(E72Q)-GFP and HK2-GFP(11) in the indicated combinations. (D) MondoA nuclear localization was determined by cellular fractionation of HeLa cells treated for 4 hr with DMEMAcidic and transfected with siScrm (siRNA control), siATP5I, siANT2 or siHK2. Tubulin and LaminB1 served as controls for cytoplasmic and nuclear fractions, respectively. In (A and C), TXNIP mRNA levels were determined by RT-qPCR. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001; ns – not significant.

https://doi.org/10.7554/eLife.40199.011
The MondoA-dependent acidosis response.

RNA-sequencing was used to determine differentially regulated genes for HeLa and HeLa:MondoA-knockout cells treated with DMEMAcidic for 4 hr. Differentially regulated genes from duplicate biological samples were determined. (A) Heatmaps depicting TXNIP, ARRDC4 and the top 500 differentially regulated genes in HeLa cells following DMEMAcidic. The genotype:treatment interaction term was calculated using DESeq2 and indicates the influence of both genotype and treatment on differential expression. (B) Volcano plot of log2(fold-change) of HeLa cells treated with DMEMAcidic compared to HeLa:MondoA-KO cells treated with DMEMAcidic. Genes with an adjusted p-value≤1E-10 that are upregulated or downregulated in HeLa:MondoA-KO cells treated with DMEMAcidic are indicated in red and blue, respectively. Overrepresentation analysis was performed for upregulated and downregulated genes. Enriched pathways and their respective p-values are given in the red and blue boxes for upregulated and downregulated genes, respectively. (C) GSEA and leading edge analysis was conducted for HeLa cells treated with DMEMAcidic compared to HeLa:MondoA-KO cells treated with DMEMAcidic. Depicted are networks of gene sets with a nominal p-value≤0.001. Node colors are representative of whether the gene set was positively (red) or negatively (blue) enriched. Node size represents gene set size. Connecting line thickness represents similarity between two nodes.

https://doi.org/10.7554/eLife.40199.012
Figure 6—source data 1

Acidosis regulated genes and pathways.

https://doi.org/10.7554/eLife.40199.013
Figure 6—source data 2

Pathways enriched in acidosis treated MondoA knockout HeLa cells.

https://doi.org/10.7554/eLife.40199.014
Figure 7 with 1 supplement
MondoA preferentially occupies the promoters of TXNIP and ARRDC4.

ChIP sequencing was performed on single biological samples and used to identify MondoA’s genomic binding sites in HeLa cells grown in DMEM or treated with DMEMAcidic for 4 hr. (A) Heatmaps showing ~100 MondoA binding sites located within the promoters of potentially regulated genes. We defined the promoter region as being 2 kb upstream or downstream of the transcriptional start site (TSS). (B) Histogram of counts per million reads (CPM) values for the top 50 promoter-localized MondoA genomic binding sites following a 4 hr DMEMAcidic treatment of HeLa cells. (C) Genome browser views derived from ChIP-seq experiment of the MondoA binding sites in the promoters of the indicated genes. (D) Independent biological triplicates were grown in DMEM or treated for 4 hr with DMEMAcidic. ChIP-PCR was used to validate MondoA occupancy on the promoters of the indicated genes under the two experimental conditions. **p<0.01.

https://doi.org/10.7554/eLife.40199.015
Figure 7—figure supplement 1
DMEMAcidic increases MondoA genomic occupancy.

We analyzed the occupancy of MondoA at the 50 most highly bound promoters identified in our ChIP-seq experiment. The ratio of binding in DMEMAcidic/DMEM is presented and shows the fold increase in occupancy following DMEMAcidic treatment.

https://doi.org/10.7554/eLife.40199.016
MondoA coordinates the transcriptional response to cytoplasmic glucose and mitochondrial ATP.

Schematic depicting how acidosis drives MondoA transcriptional activity through the generation of mitochondrial ATP and utilization by mitochondria-bound hexokinase to produce G6P, which drives MondoA nuclear localization and transcriptional activity.

https://doi.org/10.7554/eLife.40199.017

Tables

Key resources table
Reagent type
(species) or resource
DesignationSource or referenceIdentifiersAdditional information
AntibodyAnti-HK2
(goat polyclonal)
Santa Cruzsc6521(1:1,000)
AntibodyAnti-MLX
(rabbit monoclonal (D8G6W)
Cell Signaling85570S(1:1,000)
AntibodyAnti-MlxIP (MondoA)
(rabbit polyclonal)
Proteintech13614–1-AP(1:2,000)
AntibodyAnti-SDHA
(2E3GC12FB2AE2)
(mouse monoclonal
AbcamAB147(1:15,000)
AntibodyAnti-Tubulin
(mouse monoclonal)
Molecular Probes236–10501(1:50,000)
AntibodyAnti-TXNIP
(rabbit monoclonal)
Abcamab188865(1:2,000)
AntibodyAnti-LaminB1
(rabbit polyclonal)
Abcamab16048(1:1,000)
Antibodyanti-goat IgG-HRP
(donkey polyclonal)
Santa Cruzsc-2056(1:2,000)
AntibodyMouse IgG, HRP-linked
whole Ab
(sheep polyclonal)
GE Life ScienceNA-931(1:5,000)
AntibodyRabbit IgG, HRP-linked
whole Ab
(donkey polyclonal)
GE Life ScienceNA-934(1:15,000)
Chemical
compound, drug
BCECF-AMThermo FisherB1170
Chemical
compound, drug
Blotting Grade
Blocker Non-fat Dry Milk
Bio-Rad1706404XTU
Chemical
compound, drug
CCCPSigma AldrichC2759
Chemical
compound, drug
ChloroquineSigma Aldrich415480
Chemical
compound, drug
Deoxy-D-Glucose,
2-[1,2-3H(N)]
American Radiolabeled
Chemicals, Inc.
0103–250
Chemical
compound, drug
DMEMGibco11995–065
Chemical
compound, drug
DMEM Powder without
sodium bicarbonate,
glucose, L-glutamine,
sodium pyruvate and
phenol red
Cellgro90–113-PB
Chemical
compound, drug
DMSOFisherBP231
Chemical
compound, drug
FCCPSigma AldrichC2920
Chemical
compound, drug
Fetal bovine serum (FBS)Gibco26140–079
Chemical
compound, drug
GlucoseFisherD16-1
Chemical
compound, drug
GlutamineCellgro25–005 Cl
Chemical
compound, drug
HBSSGibco24020–117
Chemical
compound, drug
HEPESSigma AldrichH3375
Chemical
compound, drug
JC1Thermo FisherT3168
Chemical
compound, drug
MetforminSigma AldrichD150959
Chemical
compound, drug
MonensinSigma AldrichM5273
Chemical
compound, drug
Non-essential
amino acids
Gibco11140–050
Chemical
compound, drug
Oligomycin ASigma Aldrich75351
Chemical
compound, drug
Pennicillin/StreptomycinGibco15140–112
Chemical
compound, drug
Phenol RedSigma AldrichP-0290
Chemical
compound, drug
Sodium bicarbonateFisherL-23200
Chemical
compound, drug
Sodium pyruvateGibco11360–070
Chemical
compound, drug
Trypsin-EDTA (0.25%)Gibco25200–056
Chemical
compound, drug
Tween-20FisherBP-337
Commercial
assay, or kit
Quick RNA miniprep kitGenesee ScientificR1055
Commercial
assay, or kit
ATP determination kitThermo FisherA22066
Commercial
assay, or kit
Mitochondria isolation
kit for cultured cells
Thermo Fisher89874
Commercial
assay, or kit
Stranded mRNA-Seq
kit with mRNA capture
beads
Kapa BiosystemsKK8421
Commercial
assay, or kit
Galacto-Light Reaction
Buffer Diluent with
Galacton-Plus
Thermo FisherT1055
Commercial
assay, or kit
Luciferase Assay SystemPromegaE4550
Commercial
assay, or kit
ProSignal Pico ECLGenesee Scientific20-300B
Commercial
assay, or kit
Reporter 5X Lysis BufferPromegaE4030
Commercial
assay, or kit
SuperSignal West FemtoThermo Fisher34094
Cell line
(M. musculus)
MondoA +/+mouse
embryonic fibroblasts
Peterson et al. (2010),
PMID: 20385767
Cell line
(M. musculus)
MondoA Δ/Δmouse
embryonic fibroblasts
Peterson et al. (2010),
PMID: 20385767
Cell line
(H. sapiens)
143BWeinberg et al. 2010,
PMID: 20421486
Cell line
(H. sapiens)
143Bρ0Weinberg et al. 2010,
PMID: 20421486
Cell line
(H. sapiens)
143Bρ0:Wild type cybridWeinberg et al. 2010,
PMID: 20421486
Cell line
(H. sapiens)
143Bρ0:ΔATP6/ΔATP8
cybrid
Boominathan et al. (2016),
PMID: 27596602
Cell line
(H. sapiens)
143Bρ0:ΔATP6/ΔATP8
cybrid + ATP6nuc+ATP8nuc
Boominathan et al. (2016),
PMID: 27596602
Cell line
(H. sapiens)
HeLaATCCCCL-2
Cell line
(H. sapiens)
BJ-TertATCCCRL-4001
Sequence-based
reagent
TXNIP_forward (human):
TGACTTTGGCCTACAGTGGG
Peterson et al. (2010),
PMID: 20385767
Sequence-based
reagent
TXNIP_reverse (human):
TTGCGCTTCTCCAGATACTGC
Peterson et al. (2010),
PMID: 20385767
Sequence-based
reagent
TXNIP_forward (mouse):
CCTGACCTAATGGCACC
Peterson et al. (2010),
PMID: 20385767
Sequence-based
reagent
TXNIP_reverse (mouse):
GAGATGTCATCACCTTCAC
Peterson et al. (2010),
PMID: 20385767
Sequence-based
reagent
ATP5I_forward: CAGGTCTCTCCGCTCATCAAGThis paper
Sequence-based
reagent
ATP5I_reverse: GCCCGAGGTTTTAGGTAATTGTThis paper
Sequence-based
reagent
Actin_forward: TCCATCATGAAGTGTGACGTPeterson et al. (2010),
PMID: 20385767
Sequence-based
reagent
Actin_reverse: TACTCCTGCTTGCTGATCCACPeterson et al. (2010),
PMID: 20385767
Sequence-based
reagent
TXNIP_forward ChIP primer:
CAGCGATCTCACTGATTG
This paper
Sequence-based
reagent
TXNIP_reverse ChIP primer:
AGTTTCAAGCAGGAGGCG
This paper
Sequence-based
reagent
ARRDC4_forward ChIP primer:
TGCTTTAGCGAGAACCCAGT
This paper
Sequence-based
reagent
ARRDC4_reverse ChIP primer:
TGGACAGACAGTGGGAAACA
This paper
Sequence-based
reagent
TMEM97_forward ChIP primer:
CTTACTGCAGAAGGCCCAAG
This paper
Sequence-based
reagent
TMEM97_reverse ChIP primer:
TGTAGATTGCGGTTGTGAGC
This paper
Sequence-based
reagent
KLF10_forward ChIP primer:
AATCAACGGCAAAGGTGTGT
This paper
Sequence-based
reagent
KLF10_reverse ChIP primer:
CACTCAATCAGGTGGCCTCT
This paper
Sequence-based
reagent
siRNA: Dharmacon
ON-TARGETplus control
siRNA
GE Life SciencesD00-1810-10-20
Sequence-based
reagent
siRNA: siATP5I SmartPoolGE Life SciencesM-019688–01
Sequence-based
reagent
siRNA: siSLC25A5
SmartPool (siANT2)
GE Life SciencesM-007486
Sequence-based
reagent
siRNA: siHK2 SmartPoolGE Life SciencesL-006735-00-0005
recombinant DNA
reagent
LXSH (plasmid)Stoltzman et al. (2008),
PMID: 18458340
Recombinant DNA
reagent
LXSH-MondoA
(plasmid)
Stoltzman et al. (2008),
PMID: 18458340
Recombinant DNA
reagent
LXSH-MondoA(I766P)
(plasmid)
Stoltzman et al. (2008),
PMID: 18458340
Recombinant DNA
reagent
pcDNA3-AT1.03 (ATEAM)
(plasmid)
Imamura et al. (2009),
PMCID: PMC2735558
Recombinant DNA
reagent
pcDNA3-mitAT1.03
(Mit-ATEAM)
(plasmid)
Imamura et al. (2009),
PMID: 19720993
Recombinant DNA
reagent
pcDNA3-AT1.03
R122K/R126K
(plasmid)
Imamura et al. (2009),
PMID: 19720993
Recombinant DNA
reagent
pcDNA3-mitAT1.03
R122K/R126K
(plasmid)
Imamura et al. (2009),
PMID: 19720993
Recombinant DNA
reagent
pEGFP-N1-mVDAC1
(plasmid)
Zaid et al. (2005),
PMID: 15818409
Recombinant DNA
reagent
pEGFP-N1-mVDAC1
(E72Q)
(plasmid)
Zaid et al. (2005),
PMID: 15818409
Recombinant DNA
reagent
pCDV-SPORT6-HK2
(plasmid)
Stoltzman et al. (2008),
PMID: 18458340
Recombinant DNA
reagent
pCDV-SPORT6-HK2
(D657A)
(plasmid)
Stoltzman et al. (2008),
PMID: 18458340
Recombinant DNA
reagent
pcDNA3.1-mVDAC1-
GFP(1-10)
(plasmid)
This paperProgenitors:
PCR, pcDNAGFP(1-10)
Recombinant DNA
reagent
pcDNA3.1-mVDAC1
(E72Q)-GFP(1-10)
(plasmid)
This paperProgenitors:
PCR, pcDNAGFP(1-10)
Recombinant DNA
reagent
pcDNA3.1-HK2-GFP(11)
(plasmid)
This paperProgenitors:
PCR, pcDNAGFP(11)
Recombinant DNA
reagent
pGL3Basic-TXNIP_Promoter
(plasmid)
Peterson et al. (2010),
PMID: 20385767
Recombinant DNA
reagent
pGL3Basic-TXNIP_Promoter(ChoREmut)
(plasmid)
Peterson et al. (2010),
PMID: 20385767
Recombinant DNA
reagent
pcDNAGFP(1-10)Kamiyama et al. (2016),
PMID: 26988139
Recombinant DNA
reagent
pcDNAGFP(11)Kamiyama et al. (2016),
PMID: 26988139
Software, algorithmPrismGraphpad Software
Software, algorithmImageJhttps://imagej.nih.gov/ij/
Software, algorithmCFX Manager 3.1Bio-Rad
Software, algorithmRhttps://www.r-project.org
Software, algorithmjavaGSEABroad Institute
Software, algorithmCytoscape 3.6.1https://cytoscape.org
Software, algorithmNIS ElementsNikon
OtherNunc Lab-Tek II
Chambered Coverglass,
8-well
Thermo Fisher155409PK
Other3.5 mm glass bottom
culture dishes
MatTek CorporationP35G-.15–14 C
OtherHybond P PVDF
Membrane; 0.45 μm
Genesee Scientific83–646R
Other2 ml PTFE tissue grinderVWR89026–398
OtherBioruptor Plus
sonication devise
DiagenodeB01020001

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  1. Blake R Wilde
  2. Zhizhou Ye
  3. Tian-Yeh Lim
  4. Donald E Ayer
(2019)
Cellular acidosis triggers human MondoA transcriptional activity by driving mitochondrial ATP production
eLife 8:e40199.
https://doi.org/10.7554/eLife.40199