MA1-250

antibody from Invitrogen Antibodies

Targeting: DDIT3 CHOP, CHOP10, GADD153

Provider product page for MA1-250

Western blot

Immunocytochemistry

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Antibody data

Antibody Data
Antigen structure
References [84]
Comments [0]
Validations
Western blot [4]
Immunocytochemistry [1]
Other assay [40]

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Product number: MA1-250 - Provider product page
Provider: Invitrogen Antibodies
Product name: CHOP Monoclonal Antibody (9C8)
Antibody type: Monoclonal
Antigen: Other
Description: MA1-250 detects CHOP protein in human and mouse samples.
Antibody clone number: 9C8
Concentration: 1 mg/mL

DDIT3 protein structure - MA1-250 shown in red.

Supportive validation

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Experimental details: Western blot analysis of CHOP was performed by loading 30 µg of whole cell lysates from undifferentiated 3T3-L1 fibroblasts (Fib) and 3T3-L1 fibroblasts differentiated into adipocytes (Day 2 and Day 9 post-differentiation) per well onto an SDS-PAGE gel. 3T3-L1 fibroblasts were cultured in differentiation medium for 3 days to generate adipocytes. Differentiated adipocytes were maintained in maturation media containing either 5mM glucose (normal glucose) or 30mM glucose (high glucose). Whole cell lysates were generated from cells collected on day 2 and day 9 post-differentiation (early maturation and late maturation, respectively) and duplicate lanes of each differentiated sample were loaded. Proteins were transferred to a PVDF membrane and blocked with 5% milk. The membrane was probed with a CHOP monoclonal antibody (Product # MA1-250) at a dilution of 1:1000, washed, and probed with an HRP-conjugated anti-mouse IgG at a dilution of 1:10,000. Detection was performed using ECL Plus (Product # 32132). Data courtesy of the Innovators Program.

Supportive validation

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Experimental details: Western blot analysis of HeLa cell lysates using CHOP Monoclonal Antibody (Product # MA1-250). HeLa cells were left untreated (-) or treated with tunicamycin (5 µg/mL) for 11 hours (+). Membranes containing whole cell extracts were blocked with 5% milk in TBST (blocking buffer) and probed overnight at 4°C with MA1-250 (diluted 1:2000 in blocking buffer), followed by a goat anti-mouse IgG-HRP secondary antibody (diluted in blocking buffer) and a chemiluminescent substrate. Membranes were also probed for actin as a loading control. Data courtesy of the Innovators Program.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of CHOP was performed by loading 30 µg of whole cell lysates from undifferentiated 3T3-L1 fibroblasts (Fib) and 3T3-L1 fibroblasts differentiated into adipocytes (Day 2 and Day 9 post-differentiation) per well onto an SDS-PAGE gel. 3T3-L1 fibroblasts were cultured in differentiation medium for 3 days to generate adipocytes. Differentiated adipocytes were maintained in maturation media containing either 5mM glucose (normal glucose) or 30mM glucose (high glucose). Whole cell lysates were generated from cells collected on day 2 and day 9 post-differentiation (early maturation and late maturation, respectively) and duplicate lanes of each differentiated sample were loaded. Proteins were transferred to a PVDF membrane and blocked with 5% milk. The membrane was probed with a CHOP monoclonal antibody (Product # MA1-250) at a dilution of 1:1000, washed, and probed with an HRP-conjugated anti-mouse IgG at a dilution of 1:10,000. Detection was performed using ECL Plus (Product # 32132). Data courtesy of the Innovators Program.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockout of CHOP was achieved by CRISPR-Cas9 genome editing using LentiArray™ Lentiviral sgRNA (Product # A32042, Assay ID CRISPR830773_LV) and LentiArray Cas9 Lentivirus (Product # A32064). Western blot analysis of CHOP was performed by loading 30 µg of HeLa Wild Type untreated (Lane 1), HeLa Wild Type treated with 5 µg/mL Tunicamycin for 4hrs (Lane 2), HeLa Cas9 untreated (Lane 3), HeLa Cas9 treated with 5 µg/mL Tunicamycin for 4hrs (Lane 4), HeLa CHOP KO untreated (Lane 5) and HeLa CHOP KO Treated with 5 µg/mL Tunicamycin for 4hrs (Lane 6) whole cell extracts. The samples were electrophoresed using NuPAGE™ Novex™ 4-12% Bis-Tris Protein Gel (Product # NP0322BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with Anti-CHOP Monoclonal Antibody (9C8) (Product # MA1250, 1 µg/mL dilution) and Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using SuperSignal™ West Atto Ultimate Sensitivity Substrate (Product # A38556). Loss of signal upon CRISPR mediated knockout (KO) using the LentiArray™ CRISPR product line confirms that antibody is specific to CHOP.

Supportive validation

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Experimental details: Figure 1 Selective downregulation of BiP and activation of the UPR signaling pathways by HKH40A. ( a ) Chemical structure of HKH40A; ( b ) Representative protein bands from western blot analysis. HCT-116 and HT-29 cells were cultured for the indicated time in the presence of 100 nM HKH40A. Total cell lysates were subjected to western blot analysis with specific antibodies as indicated, beta -actin was used as a loading control. ATF6 was probed with two different antibodies. The top panel was probed by a rabbit polyclonal that only detects total ATF6 (p90). The bottom panel was probed by the mouse monoclonal antibody from Abcam that detects both full-length ATF6 (p90) and cleaved products ATF6 (p50). ( c ) Analysis of XBP1 splicing by RT-PCR using XBP1-specific primers. Tunicamycin (TM) (2.5 mu M) treatment was used as positive control and GAPDH was internal control. ( d ) Western blot analysis of other ER chaperons in HCT-116 and HT-29 cells treated with 100 nM HKH40A

Supportive validation

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Experimental details: Figure 1 Expression of two critical components of the UPR, BiP and CHOP are blocked by L. pneumophila ( a ) Caspase 1/11 (-/-) bone marrow derived macrophages or ( b ) HEK-293 FCgamma cells-expressing Fc gamma (HEK-293 FCgamma) were either left uninfected or infected with WT or DeltadotA strains of L. pneumophila at an MOI of 150. Cells were then left untreated or treated with thapsigargin (Tg; 1 mug mul -1 ) for 6 h. BiP, CHOP and PDI protein expression were monitored via immunoblot. GAPDH was used as a loading control. Full blots are provided for Fig. 1a,b in Supplementary Fig. 5 . ( c , d ) BiP and CHOP levels were quantified respectively from three biological replicates using the same experimental conditions in HEK-293 FCgamma cells. Data are depicted as the fold change normalized to uninfected levels. Data represent three independent experiments. Values in all graphs are means+-s.e.m. * P

Supportive validation

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Experimental details: Figure 5 Cd 2+ -induced reactive oxygen species (ROS) formation in kidney PTC induces up-regulation of CHOP, but has no effect on Wnt signaling . (A) When used, the ROS scavenger alpha-tocopherol was pre-incubated for 1 h. Cells were loaded with 20 muM carboxy-H 2 DCFDA. Cell suspensions were incubated with Cd 2+ +- alpha-tocopherol and fluorescent signals were measured immediately at 500/535 nm. Graph shows means +- SEM of 4-7 experiments. (B) beta-catenin immunoblot of WKPT-0293 Cl.2 cells +- alpha-tocopherol (TOCO) (100 muM) +- 25 muM Cd 2+ for 6 h. (C) Expression of CHOP mRNA in WKPT-0293 Cl.2 cells without (ctl) or with 25 muM Cd 2+ by RT-PCR. (D) Prevention of Cd 2+ -mediated increase in CHOP mRNA expression by alpha-tocopherol. (E) Association of CHOP with TCF4 is increased in WKPT-0293 Cl.2 cells incubated with 25 muM Cd 2+ for 6 h when compared to controls (ctl). (F) Effect of CHOP overexpression on Cd 2+ -induced expression of c-Myc by RT-PCR. PTC transiently transfected with full length human CHOP (CHOP) or empty vector (vector) were exposed to 25 muM Cd 2+ .

Supportive validation

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Experimental details: Figure 3 TUSC3 loss leads to increased viability, N-glycosylation and Akt signaling. (a) shTUSC3 and control cells were serum starved for 36 hours (-) and serum was added for 30 minutes before lysis (+). PI3K/Akt and MAPK signaling pathway were evaluated by immunoblotting. Increased downstream activation of Akt can be observed in serum starved TUSC3 silenced PC3 cells as well as in DU145 cells after stimulation with serum. ER stress and CHOP are induced by prolonged serum starvation (lane 5 and 6) in PC3 cells. Loss of TUSC3 decreases CHOP levels in PC3 cells (lane 7 and 8). (b) Increased N-glycosylation in shTUSC3 cells. Lectin blotting using Concanavalin A and Phytohaemagglutinin-L lectins on isolated cell surface proteins was performed in cell membrane fractions of PC3 and DU145 cells following 72 h serum starvation. Control for protein loading was performed by amido black staining ( Supplementary Figure S2C ). (c) DU145 and PC3 prostate cancer cell lines were serum starved for 72 hours before lysis. Silencing of TUSC3 (sh) leads to sustained phosphorylation of Akt and decreased expression of CHOP in both cell lines. (d) Viability of TUSC3 silenced (shTUSC3) and control (scrambled shRNA) prostate cancer cells was assessed with the CellTiter-Blue(r) Assay after treatment with 5 muM tunicamycin or DMSO for 72 hours in full medium. Experiments were performed in triplicates and results are representative of several independent experiments. * p = 0.01.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 4 TUSC3 alleviates ER stress in prostate cancer cell lines. (a) CHOP and ENTPD5 expression was measured after 24 hours of 5 muM tunicamycin (TM) or DMSO treatment. Decreased CHOP induction in shTUSC3 PC3 cells indicates reduced ER stress response (lane 7 vs. 8). (b) Cells were treated with 5 muM tunicamycin (TM) or DMSO for 24 hours. TUSC3 mRNA expression was assessed using qRT-PCR. (c) UPR was assessed by expression levels of BiP, IRE-1 and XBP-1s in serum starved (72 h) prostate cancer cells. XBP-1 mRNA splicing was also measured by semiquantitative RT-PCR. Intensity of the bands was quantified and ratio of spliced to unspliced isoform was calculated. Increased ratio indicates elevated levels of ER stress. Cropped blots are depicted for clarity reasons and full-length blots will be provided upon request. (d) qRT-PCR was used to evaluate the mRNA expression of BiP and spliced XBP-1 isoform. Silencing of TUSC3 leads to an increase in BiP expression in PC3 cells and conversely to an increase in spliced XBP-1 in DU145 cells, suggesting opposing effects on UPR signaling.* p = 0.03 and ** p = 0.04.

Supportive validation

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Experimental details: Figure 5 Guanabenz treatment alters T cell distribution and upregulates ISR activity while protecting oligodendrocytes in chronic EAE. All analyses conducted using lumbar spinal cords of PID15 mice immunized with adjuvant only (CFA) or adjuvant with MOG 35-55 (EAE), then treated with vehicle or guanabenz from PID7 to 15. ( a ) Haematoxylin and eosine (H&E) staining. Note only the vehicle-treated EAE sample displays cellular infiltrates (arrowhead). ( b ) LFB staining. Note the demyelinated focal areas (dotted areas) in vehicle-treated EAE mice only. ( c ) Higher magnification of sections stained with toluidine blue. Note the demyelination (arrows) in areas of cellular infiltration in vehicle-treated EAE mice. ( d ) Quantification of cells positive for CD3, a T cell marker. ( e ) Isolated pockets of these CD3+ areas were considered 'lesion' areas and quantified. ( f ) Quantification of cells positive for ASPA, a mature oligodendrocyte marker, in 'lesion' areas. ( g ) Quantification of cells positive for both p-eIF2alpha and tubulin polymerization promoting protein (TPPP), a mature oligodendrocyte marker. Unpaired t- test, * P

Supportive validation

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Experimental details: Figure 5 Global protein translation inhibition does not the affect the UPR in the same manner as L. pneumophila infection. ( a ) RAW 264.7 cells were left untreated or treated with either thapsigargin (Tg; 1 mug mul -1 ), CHX (1 mug mul -1 ), or a combination of both for 6 h. Protein levels of UPR targets BiP, CHOP and PDI were monitored via immunoblot. ( b ) RAW267.4 macrophage cells were treated with CHX or left untreated and BiP and CHOP mRNA levels were monitored via qRT-PCR. Three internal replicates were performed, values in all graphs are means+-s.e.m. * P

Supportive validation

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Experimental details: Figure 2. Cur inhibits UVA-induced ER Stress in human dermal fibroblasts. (A) Morphological alterations in the ER were observed by transmission electron microscopy in control, Cur, UVA and Cur + UVA groups. Scale bars: Control, Cur and UVA groups, 500 nm; Cur + UVA group, 1 uM (a different magnification is presented for the last group as a clear image could not be attained at the higher magnification). The areas of increased magnification within each image were taken directly from the larger image therefore the precise magnification cannot be stated. (B) Western blot analysis was performed to investigate the protein expression of GRP78 and CHOP using specific antibodies. (C) Relative expression of GRP78 and CHOP proteins was quantified by densitometry. All data are presented as the mean +- standard deviation, n=3. **P

Supportive validation

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Experimental details: Fig 5 Impact of 2062 on macrophage ER stress response. (A) RNA-Seq heatmap of BMDM genes regulated by 2062 (left) or inactive congener (right) over time. Fold change over DMSO control is indicated by numbers. (B) 2062 treatment induces a signature response of an ER stress. BMDM were treated for indicated times with 3 muM 2062. Cytosolic fractions separated on 10% SDS-PAGE were probed with antibodies to p-PERK, p-eIF2alpha and CHOP.

Supportive validation

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Experimental details: Figure 1 The Transcriptional and Translational UPR Landscape In Vivo (A) Overview of the experimental design. Mice (n = 3) were injected intraperitoneally with 0.1 mg/kg tunicamycin (Tm), and livers were collected at 0, 2, 4, 6, and 10 h following injection. (B) Chop , Ppp1r15a , Atf4 , Bip , and Grp94 mRNAs expression in mouse livers at the indicated times following Tm injection quantified by qPCR and normalized to cyclophilin. Data are mean +- SEM (n = 3). * p < 0.05, ** p < 0.01, and *** p < 0.001, as determined by two-way ANOVA. (C) FPKM values from RNA-seq showing the changes in transcript abundance in mouse liver 2 h following Tm injection. Downregulated (purple) and upregulated (orange) mRNAs that passed a 2-fold cut-off. (D) Same as (C), but 4 h following Tm injection. (E) Representative polysome profiles (optical density of the fractions at 260 nm) on 7%-47% sucrose gradients of mouse liver extracts collected after Tm injection for the indicated time. (F) Quantification of polysome profiles such as shown in (E) and represented as a pentasomes (five ribosomes) to disomes (two ribosomes) ratio. Data are mean +- SEM (n = 3). (G) Autoradiogram of [35S]-methionine-labeled proteins in cell lysates resolved by SDS-PAGE after a 10-min labeling pulse of HeLa cells exposed to Tm (2.5 mug/mL) for the indicated times. Lower panel is an image of the InstantBlue-stained gel. (H) Representative immunoblots with indicated antibodies with lysates from mouse livers treated with Tm for

Supportive validation

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Experimental details: Figure 4. Memantine and clemastine induce ER stress. (A) Effect of memantine, clemastine, and flunarizine on the expression of ER stress markers. NGF-differentiated PC12D cells were treated with 2 uM tunicamycin (Tm), 10 mM 2-deoxyglucose (2-DG), 100 uM memantine (Mem), 5 uM clemastine (Cle), or 20 uM flunarizine (Flu). After 12 h (for detection of EIF2S1 phosphorylation) or 24 h (for detection of HSPA5 and DDIT3 expression), the cells were collected and subjected to western blotting analysis with the indicated antibodies. (B) Memantine and clemastine induce EIF2AK3 phosphorylation. NGF-differentiated PC12D cells were treated with the indicated compounds at the same concentrations as described in (A). After 12 h, the cells were collected and subjected to western blotting analysis with the indicated antibodies. (C) Memantine and clemastine induce alternative Xbp1 mRNA splicing. NGF-differentiated PC12D cells were treated with the indicated compounds for 12 h at the same concentrations as described in (A). Unspliced (U) and spliced (S) Xbp1 were detected by RT-PCR. Data are shown as mean +- SD (n = 3). n.s., non-significant, *p < 0.05, **p < 0.01 (two-tailed Student's t test)

Supportive validation

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Experimental details: Figure 4 The effect of Ezetimibe on unfolded protein response (UPR) gene expression in THP-1 cells exposed to ischemia-reperfusion (IR). ( a ) The mRNA expression of activating transcription factor 6 (ATF6) and CCAAT-enhancer-binding protein homologous protein (CHOP). ( b ) Representative Western blot analyses for the indicated proteins. ( c,d ) The average quantification of ATF6 and CHOP obtained by the densitometric analysis of three independent experiments. mRNA was analyzed by quantitative real-time PCR; normalized gene expression levels are given as the ratio between the mean value for the target gene and that for beta-actin in each sample. Data are expressed as mean +- SD. * p < 0.01 vs. control (up-regulation); ** p < 0.01 vs. IR; SS p < 0.01 vs. control (down-regulation).

Supportive validation

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Experimental details: Figure 5 Pgrmc1 KO mice shows lipotoxicity in HFD condition. ( A ) Gross image of hearts of HFD WT and Pgrmc1 KO mice. Heart weight (HW), HW per body weight (BW) and HW per tibia length were measured. ( B ) H&E staining of hearts of HFD WT and Pgrmc1 KO mice (scale bar 50 um). Nucleus per area was measured by Image J program. ( C ) Western blot analysis and quantification of ER stress genes in hearts of HFD WT and Pgrmc1 KO mice. beta-Actin was used for an internal control. ( D ) Cardiac levels of reduced (GSH) and oxidized (GSSG) glutathione (uM), and ratio of GSSG:GSH in HFD WT and Pgrmc1 KO mice. ( E ) mRNA expression of pro-inflammatory genes in hearts of HFD WT and Pgrmc1 KO mice. Rplp0 was used for an internal control. ( F ) Plasma CPK (U/I) level of HFD WT and Pgrmc1 KO mice. ( G ) Masson trichrome staining of hearts of HFD WT and Pgrmc1 KO mice (scale bar 30 um). Positive area was measured by Image J program. Blue fibroblasts were set as positive. ( H ) Western blot analysis and quantification of ANP in hearts of HFD WT and Pgrmc1 KO mice. beta-Actin was used for an internal control. Values represent means +- SD. *p < 0.05. Student's t test was performed. Total numbers of mice used for experiment were 10 (HFD WT) and 7 (HFD Pgrmc1 KO).

Supportive validation

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Experimental details: Figure 4 Aging preferentially activates UPR-mediated proapoptotic signaling in the aortae of ApoE -/- mice. A through C , Aortae derived from aged ApoE -/- mice displayed a significant increase in the levels of the apoptosis executor CHOP (protein and mRNA), phosphorylated JNK, and cleaved Casp-3 relative to WD-fed mice. D through G , Immunofluorescence analysis of the aortic expression of CHOP ( D and E ), and cleaved Casp-3 ( F and G ) in each group. Fresh and frozen cross-sections of aortae without evident lesions were immunostained for CHOP ( D ) or cleaved Casp-3 ( F ), and their fluorescence signals (red) were quantified using ImageJ ( E and G , respectively). Nuclei were counterstained with DAPI (blue), and elastin was visualized via its autofluorescence (green). The yellow boxes indicate the areas shown in higher magnification below the corresponding images. From each aortic cross-section, the red fluorescence intensity was quantified in 3 to 5 randomly selected image fields, averaged, and expressed as arbitrary units (a.u. ; n=3-5). Data are presented as means+-SE. * P

Supportive validation

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Experimental details: Figure 5. CRISPR/Cas9-mediated knockdown of ELOVL2 promotes the loss of mitochondrial transmembrane potential and apoptosis in ACHN cells. (A) Analysis of cell apoptosis by caspase 3/7 assay in ACHN/sgControl or ACHN/sgELOVL2 cells. (B) Representative images of Annexin V-FITC staining in ACHN/sgControl or ACHN/sgELOVL2 cells. Scale bar, 100 um. (C) Aggregates/monomers relative ratio evaluation in ACHN/sgControl or ACHN/sgELOVL2 cells by using JC-1 assay. (D) Apoptosis-related gene expression levels in ACHN/sgControl or ACHN/sgELOVL2 cells. (E) Representative images of ER tracker staining in ACHN/sgControl or ACHN/sgELOVL2 cells. Scale bar, 50 um. (F) Representative flow cytometry dot plots of ER tracker staining in ACHN/sgControl or ACHN/sgELOVL2 cells. (G) ER tracker fluorescence quantification in ACHN/sgControl or ACHN/sgELOVL2 cells. (H) Western blot analysis of pIRE1alpha, IRE1alpha, and CHOP in ACHN/sgControl or ACHN/sgELOVL2 cells. P-values were assessed using ANOVA with post hoc comparisons using Dunnett's test (A, C, D and G). **P

Supportive validation

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Experimental details: FIG 8 Mpt64 inhibits the unfolded protein response. (A) RAW267.4 cells stably transduced with empty lentivirus or lentivirus expressing Mpt64DeltaSP under the control of the CMV promoter were treated with DMSO or thapsigargin (Tg) (50 nM) for 4 h, and CHOP protein accumulation was detected by Western blotting. (B) Quantitative densitometry analysis of the Western blot in panel A. The results of one experiment representative of >3 experiments are shown.

Supportive validation

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Experimental details: Figure 4 SLC35A1 loss promotes cell death and apoptosis upon VSV infection. ( a ) Percentage of early apoptotic cells (AnnexinV + ) and cells that lost membrane integrity due to a late apoptotic/necrotic state (AnnexinV + /PI + ) in HAP1 cell lines expressing sgRNAs against SLC35A1 and Renilla luciferase upon infection with VSV at MOI 2. ( b ) Immunoblot analysis of Golgi stress signalling pathway activation in the VSV-infected (MOI 2) and Brefeldin A-stimulated (3mug/mL) A549 cells carrying sgRNAs against SLC35A1 and Renilla luciferase . Cropped images are shown for conciseness. Full-length blots are presented in Supplementary Fig. 5 . ( c , d ) VSV-GFP virus replication in SLC35A1 knockout HAP1 cells ( c ) and HAP1 cells overexpressing SLC35A1 cDNA ( d ) infected with MOI ranging from 0.001 to 0.01 for 14 hours. ( e ) Quantification (flow cytometry) of virus titers produced by HAP1wt and two SLC35A 1 single cell knockout clones infected with MOI of 2 of VSV-GFP for 8 hours. Statistical significance was assessed using one-way ANOVA with Tukey's ( a , d ) and Dunnett's ( c , e ) tests. Unless otherwise indicated, adjusted P-values in relation to sgRen control or HAP1wt are shown. Data are represented as mean +- SD of one representative experiment out of at least two independent replicates. **p

Supportive validation

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Experimental details: Fig. 3 Loss of SLC39A7 mediates resistance to TNFalpha-induced cell death by diminishing TNFR1 surface expression. a Growth curve for KBM7 FADD - sg Ren or SLC39A7 - cells. Equal cell numbers were seeded and cell growth monitored over 7 days. Data represent mean value +- s.d. of five independent experiments; statistical analysis by t -test, * P < 0.01. b Cell viability in KBM7 FADD - sg Ren or SLC39A7 - cells treated overnight (16 h) with TNFalpha (10 ng/mL), SMAC mimetic (0.5 uM), and Nec-1s (50 µM) as indicated. Cell viability was assessed using a luminescence-based readout for ATP (CellTiter Glo). Data represent mean value +- s.d. of two independent experiments performed in triplicates. c KBM7 FADD - SpCas9 (empty, sg Ren or SLC39A7 - ) cells were stimulated for the time indicated with TNFalpha (10 ng/mL). Cells were then lysed and subjected to immunoblotting with the indicated antibodies. d Flow cytometry analysis for TNFR1 surface expression. KBM7 TNFRSF1A - cells serve as negative control for background staining. Data shown are representative of two independent experiments. e KBM7 FADD - SpCas9 (empty, sg Ren or SLC39A7 - ) cells were treated for 7 h with Brefeldin A (0.5 uM), Tunicamycin (2 µM), Thapsigargin (0.5 uM), MG-132 (10 µM) or DMSO as control. Cells were then lysed and subjected to immunoblotting with the indicated antibodies. f KBM7 TNFRSF1A - , KBM7 FADD - and KBM7 FADD - SLC39A7 - cells were lysed and subjected to immunoblotting with the i