anti zmat3  (Santa Cruz Biotechnology)

Journal: bioRxiv

Article Title: p53-induced RNA-binding protein ZMAT3 inhibits transcription of a hexokinase to suppress mitochondrial respiration

doi: 10.1101/2025.05.12.653341

Figure Lengend Snippet: (A) IGV snapshot shows location of the two sgRNAs used to generate ZMAT3 -KO HCT116 cells, the observed 57 bp deletion near sgRNA#2 and the p53 ChIP-seq peak in the ZMAT3 locus in response to p53 activation upon Nutlin treatment. The p53 ChIP-seq data was previously published . (B) RT-qPCR was performed from 3 biological replicates of ZMAT3 -WT and ZMAT3 -KO HCT116 cells. GAPDH served as a housekeeping gene control. (C) Colony formation assays were performed from 3 biological replicates of ZMAT3 -WT and ZMAT3 -KO HCT116 cells. (D) Notched box plot of the log 2 FC in RNA abundance of differentially expressed genes after RNA-Seq analysis of ZMAT3 -KO versus ZMAT3 -WT HCT116 cells. Median values for each group are indicated at the top, and the number of RNAs for which data were obtained for each group is indicated at the bottom. (E) Volcano plots showing differentially expressed proteins (shown in red) identified by performing quantitative proteomics from ZMAT3 -WT and ZMAT3 -KO HCT116 cells. (F) Most significantly enriched pathways in the gene set enrichment analysis of significantly upregulated genes (p<0.05) from the ZMAT3 -KO/ ZMAT3 -WT quantitative proteomics analysis. (G) TMT mass spectrometry peptide abundance of HKDC1 protein in ZMAT3 -WT and ZMAT3 -KO HCT116 cells. The values are the average of five biological replicates for ZMAT3-WT and four biological replicates for ZMAT3 -KO cells. (H) IGV snapshot for ZMAT3 and HKDC1 from RNA-seq from ZMAT3 -WT and ZMAT3 -KO HCT116 cells. ∗p < 0.05, ∗∗∗∗p < 0.0001.

Article Snippet: The following primary antibodies were used: anti-FLAG (1:1000 dilution; Sigma F1804), anti-p53 (DO-I) (1:1000 dilution; Santa Cruz Biotechnology sc-126), anti-ZMAT3 (1:500 dilution; Santa Cruz Biotechnology sc-398712), anti JUN (1:1000 dilution; Cell signalling, 9165S) and anti-HKDC1 (Proteintech, Catlog no: 25874-1-AP).

Techniques: ChIP-sequencing, Activation Assay, Quantitative RT-PCR, Control, RNA Sequencing, Quantitative Proteomics, Mass Spectrometry

Journal: bioRxiv

Article Title: p53-induced RNA-binding protein ZMAT3 inhibits transcription of a hexokinase to suppress mitochondrial respiration

doi: 10.1101/2025.05.12.653341

Figure Lengend Snippet: (A) Immunoblotting for ZMAT3, p53, p21 and GAPDH from ZMAT3 -WT and ZMAT3 -KO HCT116 cells with or without Nutlin treatment (24 hr). GAPDH served as the loading control. (B) Incucyte live cell proliferation assays were performed from ZMAT3 -WT and ZMAT3 -KO HCT116 cells. (C) Volcano plot for the differentially expressed genes identified by RNA-Seq from ZMAT3 -WT and ZMAT3 -KO HCT116 cells. Significantly expressed genes are indicated in red (p<0.05). ( D ) Most significantly enriched pathways identified by GSEA from the top 500 significantly upregulated genes (p<0.05) from ZMAT3 -KO vs ZMAT3 -WT RNA-seq.

Article Snippet: The following primary antibodies were used: anti-FLAG (1:1000 dilution; Sigma F1804), anti-p53 (DO-I) (1:1000 dilution; Santa Cruz Biotechnology sc-126), anti-ZMAT3 (1:500 dilution; Santa Cruz Biotechnology sc-398712), anti JUN (1:1000 dilution; Cell signalling, 9165S) and anti-HKDC1 (Proteintech, Catlog no: 25874-1-AP).

Techniques: Western Blot, Control, RNA Sequencing

Journal: bioRxiv

Article Title: p53-induced RNA-binding protein ZMAT3 inhibits transcription of a hexokinase to suppress mitochondrial respiration

doi: 10.1101/2025.05.12.653341

Figure Lengend Snippet: B) RT-qPCR and immunoblotting for HKDC1 from ZMAT3 -WT and ZMAT3 -KO HCT116 cells. GAPDH served as a housekeeping gene control. RT-qPCR are values are the average of three biological replicates. (C, D) RT-qPCR was performed in biological triplicates for ZMAT3 and HKDC1 mRNAs from HCT116, SW1222, HCEC-1CT and HEPG2 cells after transfection with a control (CTRL) siRNA or ZMAT3 siRNAs for 72 hr. GAPDH served as a housekeeping gene control. (E) Immunoblotting was performed for endogenous ZMAT3 and HKDC1 from HCT116 and HepG2 whole cell lysates after siRNA-mediated knockdown of ZMAT3 or HKDC1 for 72 hr. GAPDH served as housekeeping gene control. (F) Fold change for Zmat3, Trp53, Mdm4 and Hkdc1 mRNAs is shown from the RNA-seq from Zmat3 knock-out and wild-type MEFs. ( G) Analysis of HKDC1 mRNA levels from normal colon tissues or CRC patient samples from the TCGA COAD cohort. N refers to the number of samples in each group. (H) Fold change for Trp53, Zmat3, Cdkn1a and Hkdc1 mRNAs is shown from the RNA-seq from Trp53 knock-out and wild-type MEFs. N refers to the number of samples in each group. ∗p < 0.05, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001

Article Snippet: The following primary antibodies were used: anti-FLAG (1:1000 dilution; Sigma F1804), anti-p53 (DO-I) (1:1000 dilution; Santa Cruz Biotechnology sc-126), anti-ZMAT3 (1:500 dilution; Santa Cruz Biotechnology sc-398712), anti JUN (1:1000 dilution; Cell signalling, 9165S) and anti-HKDC1 (Proteintech, Catlog no: 25874-1-AP).

Techniques: Quantitative RT-PCR, Western Blot, Control, Transfection, Knockdown, RNA Sequencing, Knock-Out

Journal: bioRxiv

Article Title: p53-induced RNA-binding protein ZMAT3 inhibits transcription of a hexokinase to suppress mitochondrial respiration

doi: 10.1101/2025.05.12.653341

Figure Lengend Snippet: (A) Volcano plot for the differentially expressed genes identified by RNA-Seq performed after transfection of HCT116 cells with siCTRL or siZMAT3 for 72 hr. Significantly expressed genes are indicated in red (p<0.05). (B) GSEA analysis was performed for the top 500 significantly upregulated genes (p<0.05) in ZMAT3 knockdown HCT116 cells identified by RNA-seq. ( C, D ) Venn diagram of showing comparisons of the indicated RNA-Seq data sets. 1,023 significant upregulated (C) and 1,042 downregulated (D) differentially expressed genes were shared between the ZMAT3 -KO/ ZMAT3 -WT and siZMAT3/siCTRL comparisons. (E) Most significantly enriched pathways in the GSEA for the top 500 genes commonly upregulated genes (p<0.05) in ZMAT3-KO vs ZMAT3 -WT and siZMAT3 vs siCtrl comparisons from the RNA-seq data. (F, G) ZMAT3 and HKDC1 mRNA levels were determined in CRC patient samples in the TCGA COAD cohort from p53-WT (wild-type) and p53-Mutant CRC patient samples.

Article Snippet: The following primary antibodies were used: anti-FLAG (1:1000 dilution; Sigma F1804), anti-p53 (DO-I) (1:1000 dilution; Santa Cruz Biotechnology sc-126), anti-ZMAT3 (1:500 dilution; Santa Cruz Biotechnology sc-398712), anti JUN (1:1000 dilution; Cell signalling, 9165S) and anti-HKDC1 (Proteintech, Catlog no: 25874-1-AP).

Techniques: RNA Sequencing, Transfection, Knockdown, Mutagenesis

Journal: bioRxiv

Article Title: p53-induced RNA-binding protein ZMAT3 inhibits transcription of a hexokinase to suppress mitochondrial respiration

doi: 10.1101/2025.05.12.653341

Figure Lengend Snippet: (A) 2-Deoxyglucose analog of glucose together with luminescence-based enzymatic assay was used to assess relative glucose uptake in ZMAT3 -WT and ZMAT3 -KO HCT116 cells in presence and absence of HKDC1. For SW122 and HEPG2 cells relative glucose was measured in the presence and absence of siRNA-mediated knockdown of HKDC1 and ZMAT3 alone or in combination. (B, C) Metabolic flux assays were performed for basal glycolysis rate and basal mitochondrial respiration rate in ZMAT3 and/or HKDC1 knockdown in HCT116 cells. ( D, E ) Incucyte live cell proliferation assays and CCK8-based cell proliferation in HCT116 ZMAT3 -WT and KO cells in the presence and absence of siRNA-mediated knockdown HKDC1. ∗p < 0.05, (∗∗) p < 0.01, (∗∗∗) P<0.001. The results are the average of three independent experiments.

Article Snippet: The following primary antibodies were used: anti-FLAG (1:1000 dilution; Sigma F1804), anti-p53 (DO-I) (1:1000 dilution; Santa Cruz Biotechnology sc-126), anti-ZMAT3 (1:500 dilution; Santa Cruz Biotechnology sc-398712), anti JUN (1:1000 dilution; Cell signalling, 9165S) and anti-HKDC1 (Proteintech, Catlog no: 25874-1-AP).

Techniques: Enzymatic Assay, Knockdown

Journal: bioRxiv

Article Title: p53-induced RNA-binding protein ZMAT3 inhibits transcription of a hexokinase to suppress mitochondrial respiration

doi: 10.1101/2025.05.12.653341

Figure Lengend Snippet: Non-mitochondrial oxygen consumption in ZMAT3 and/or HKDC1 knockdown in HCT116 cells. Values are the average of four independents experiments.

Article Snippet: The following primary antibodies were used: anti-FLAG (1:1000 dilution; Sigma F1804), anti-p53 (DO-I) (1:1000 dilution; Santa Cruz Biotechnology sc-126), anti-ZMAT3 (1:500 dilution; Santa Cruz Biotechnology sc-398712), anti JUN (1:1000 dilution; Cell signalling, 9165S) and anti-HKDC1 (Proteintech, Catlog no: 25874-1-AP).

Techniques: Knockdown

Journal: bioRxiv

Article Title: p53-induced RNA-binding protein ZMAT3 inhibits transcription of a hexokinase to suppress mitochondrial respiration

doi: 10.1101/2025.05.12.653341

Figure Lengend Snippet: (A) IGV snapshots from the RNA-seq data following knockdown of p53 with p53 siRNAs. Data shows increased HKDC1 mRNA levels and decreased ZMAT3 mRNA levels upon p53 knockdown in HCT116 cells. (B) Fold change is shown for p53, p21, ZMAT3 , and HKDC1 mRNAs from the RNA-seq performed from siCTRL and sip53 transfected HCT116 cells. (C, D) HCT116 cells were transfected with CTRL siRNA or p53 siRNAs for 48 hr. The levels of ZMAT3, p53 , and HKDC1 mRNA or protein were measured by RT-qPCR (C) or immunoblotting from whole cell lysates (D). GAPDH was used as housekeeping gene control. (E) Fold change for ZMAT3, p21, HKDC1 and p53 mRNAs is shown from the RNA-seq from HCT116 cells treated with DMSO or Nutlin for 6 hr. (F) Immunoblotting was performed for HKDC1, ZMAT3 and p21 from ZMAT3 -WT and ZMAT3 -KO HCT116 cells with or without Nutlin treatment for 24 hr. GAPDH served as the loading control. ns refers to not significant. ( G,H ) Doxycycline inducible ZMAT3-FLAG-HA HCT116 cells treated with 2ug/ml doxycycline for 48 hr. The levels of ZMAT3 mRNA or protein induction were measured by RT-qPCR (G) or immunoblotting from whole cell lysates against HA antibody(H). GAPDH was used as housekeeping gene control. (I, J) Doxycycline inducible ZMAT3-FLAG-HA HCT116 cells were transfected with CTRL siRNA or p53 siRNAs for 48 hr followed by 48hr doxycycline treatment. The levels of ZMAT3, p53 , and HKDC1 mRNA or protein were measured by RT-qPCR (I) or immunoblotting from whole cell lysates (J). GAPDH was used as housekeeping gene control. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001

Article Snippet: The following primary antibodies were used: anti-FLAG (1:1000 dilution; Sigma F1804), anti-p53 (DO-I) (1:1000 dilution; Santa Cruz Biotechnology sc-126), anti-ZMAT3 (1:500 dilution; Santa Cruz Biotechnology sc-398712), anti JUN (1:1000 dilution; Cell signalling, 9165S) and anti-HKDC1 (Proteintech, Catlog no: 25874-1-AP).

Techniques: RNA Sequencing, Knockdown, Transfection, Quantitative RT-PCR, Western Blot, Control

Journal: bioRxiv

Article Title: p53-induced RNA-binding protein ZMAT3 inhibits transcription of a hexokinase to suppress mitochondrial respiration

doi: 10.1101/2025.05.12.653341

Figure Lengend Snippet: (A) Schematic of full-length ZMAT3 protein showing three zinc finger motifs. (B) Immunoblot from 10% input and FLAG immunoprecipitation from doxycycline inducible ZMAT3-FLAG-HA HCT116 cell lysates treated with/without doxycycline for 48 hr.

Article Snippet: The following primary antibodies were used: anti-FLAG (1:1000 dilution; Sigma F1804), anti-p53 (DO-I) (1:1000 dilution; Santa Cruz Biotechnology sc-126), anti-ZMAT3 (1:500 dilution; Santa Cruz Biotechnology sc-398712), anti JUN (1:1000 dilution; Cell signalling, 9165S) and anti-HKDC1 (Proteintech, Catlog no: 25874-1-AP).

Techniques: Western Blot, Immunoprecipitation

Journal: bioRxiv

Article Title: p53-induced RNA-binding protein ZMAT3 inhibits transcription of a hexokinase to suppress mitochondrial respiration

doi: 10.1101/2025.05.12.653341

Figure Lengend Snippet: (A ) Schematic for identification of ZMAT3-FLAG interacting proteins by IP mass spectrometry from HCT116 cells expressing doxycycline-induced ZMAT3-FLAG-HA. (B) Volcano plot showing significantly enriched proteins (shown in red) identified by ZMAT3-FLAG IP followed by mass spectrometry, in presence or absence of doxycycline from ZMAT3-FLAG-HA HCT116 cells. JUN was strongly enriched in the ZMAT3-FLAG IPs. ( C ) IGV snapshot showing JUN, POLR2A, H3K27Ac and H3K3Me3 peaks at the HKDC1 locus from the ENCODE project (accession from top to bottom: ENCSR000FAH, ENCSR000EDG, ENCSR000EEK, ENCSR000EUU, ENCSR661KMA and ENCSR333OPW). JUN binding motif (TGASTCA) is shown in blue (positive strand) and in red (negative strand). ( D ) Immunoblotting was performed using anti-FLAG beads and whole cell lysates from no doxy or doxycycline treated ZMAT3-FLAG-HA HCT116 cells. 10% of total cell lysate was used for input. GAPDH used as loading control. (E, F) ZMAT3 -WT and KO HCT116 cells were transfected with CTRL siRNA or JUN siRNAs for 48 hr. The levels of ZMAT3 , JUN , and HKDC1 mRNAs or the corresponding proteins were measured by RT-qPCR (E) or immunoblotting from whole cell lysates (F). GAPDH was used as housekeeping gene control. (G ) JUN ChIP-qPCR was performed in biological triplicates from HCT116 ZMAT3 -WT and KO cells to determine enrichment of JUN at the HKDC1 promoter. (H) Luciferase assays were performed in biological triplicates upon JUN or ZMAT3 knockdown alone or in combination using the HKDC1 promoter reporter constructs. ∗p < 0.05, ∗∗p < 0.01

Article Snippet: The following primary antibodies were used: anti-FLAG (1:1000 dilution; Sigma F1804), anti-p53 (DO-I) (1:1000 dilution; Santa Cruz Biotechnology sc-126), anti-ZMAT3 (1:500 dilution; Santa Cruz Biotechnology sc-398712), anti JUN (1:1000 dilution; Cell signalling, 9165S) and anti-HKDC1 (Proteintech, Catlog no: 25874-1-AP).

Techniques: Mass Spectrometry, Expressing, Binding Assay, Western Blot, Control, Transfection, Quantitative RT-PCR, ChIP-qPCR, Luciferase, Knockdown, Construct

Journal: bioRxiv

Article Title: p53-induced RNA-binding protein ZMAT3 inhibits transcription of a hexokinase to suppress mitochondrial respiration

doi: 10.1101/2025.05.12.653341

Figure Lengend Snippet: Schematic for ZMAT3-mediated regulation of HKDC1 expression and inhibition of mitochondrial respiration. In ZMAT3 -WT cells p53 activates ZMAT3 transcription, resulting in ZMAT3 protein binding to the transcription factor JUN. This inhibits the ability of JUN to bind to the HKDC1 promoter, low HKDC1 expression leading to controlled mitochondrial respiration and controlled cell proliferation. In ZMAT3 -knockout cells, JUN actively binds to the HKDC1 promoter and upregulates its expression resulting in increased mitochondrial respiration and increased cell proliferation.

Article Snippet: The following primary antibodies were used: anti-FLAG (1:1000 dilution; Sigma F1804), anti-p53 (DO-I) (1:1000 dilution; Santa Cruz Biotechnology sc-126), anti-ZMAT3 (1:500 dilution; Santa Cruz Biotechnology sc-398712), anti JUN (1:1000 dilution; Cell signalling, 9165S) and anti-HKDC1 (Proteintech, Catlog no: 25874-1-AP).

Techniques: Expressing, Inhibition, Protein Binding, Knock-Out

Journal: Cell death and differentiation

Article Title: Integrative multiomic approaches reveal ZMAT3 and p21 as conserved hubs in the p53 tumor suppression network.

doi: 10.1038/s41418-025-01513-8

Figure Lengend Snippet: Fig. 1 Genetic epistasis mapping reveals the effects of the p53-Zmat3 LUAD suppression axis in vivo. A Murine Zmat3 p53 response element (RE) with sgRNA for both sgZmat3 RE and gene-targeting sgZmat3. B Schematic of p53-mediated and ZMAT3-mediated tumor suppression, depicting ZMAT3-independent effects of p53 (i), p53-ZMAT3 effects (ii), and p53-independent effects of ZMAT3 (iii). C Design of tumor barcoding sequencing (Tuba-seqUltra) experiment. Each gene was targeted with 3 sgRNA, and inert, negative control sgRNAs (sgInerts) were included. The lentiviral vector delivers a U6-barcoded sgRNA (BC-sgRNA) and Pgk-Cre. Indicated viral titers (infectious units; ifu) of the lentiviral barcoded sgRNA library were delivered to mice of the indicated genotypes and tumorigenesis proceeded for 15 weeks before Tuba- seq analysis was carried out. D Relative tumor number for sgTrp53. E Log-normal mean tumor size using adaptive cutoff for sgTrp53. F Relative tumor number for sgZmat3 and sgZmat3 RE. G Log-normal mean tumor size using adaptive cutoff for sgZmat3 and sgZmat3 RE. Error bars indicate the 95% confidence interval, and confidence intervals and FDR-adjusted p-values were calculated using a bootstrap resampling approaches with 10,000 cycles, *p < 0.05, and sgInert data graphed are aggregate of three arbitrarily chosen sgInerts for (D–G).

Article Snippet: Extracts were run on 10% polyacrylamide SDS-PAGE gels, gels were transferred to PVDF membrane (Immobilon, Millipore), and membranes were blocked with 5% milk in TBST and probed with antibodies directed against ZMAT3 (sc-398712, 1:1000, Santa Cruz), p53 (CM5, 1:1000 Leica Novocastra), p21 (EPR18021, 1:1000, abcam), beta Actin (sc-47778, 1:1000, Santa Cruz), or GAPDH (10R-G109a, 1:20,000, Fitzgerald), followed by anti-mouse or anti-rabbit HRP-conjugated secondary antibodies (PI-2000-1 and PI-1000-1, 1:5,000, Vector Laboratories).

Techniques: In Vivo, Sequencing, Negative Control, Plasmid Preparation

Journal: Cell death and differentiation

Article Title: Integrative multiomic approaches reveal ZMAT3 and p21 as conserved hubs in the p53 tumor suppression network.

doi: 10.1038/s41418-025-01513-8

Figure Lengend Snippet: Fig. 2 An in vivo CRISPR/Cas9 screen uncovers effectors of p53-mediated tumor suppression. A (Left) Design for sgRNA library targeting direct, upregulated p53 target genes and (Right) heat map showing expression of 272 direct p53 target genes in non-targeting control sgRNA (sgNTC) versus sgTrp53 E1A;HrasG12V;H11Cas9 MEFs. B Immunoblot of p53, ZMAT3, and loading control (GAPDH) for transduced, selected, pre- injection cell populations for wild-type (WT, i), Zmat3-KO (ii), and p53-KO (iii) screens. C p53 target gene sgRNA library was lentivirally delivered to E1A;HrasG12V;H11Cas9 MEFs, which were grafted subcutaneously in immunocompromised mice to form tumors. Enrichment of sgRNAs was determined by comparing sgRNA representation in final tumors (TF) to pre-injection (T0) cells. Zmat3-KO (ii) and p53-KO (iii) screens were performed in the same way as the WT screen (i) but the sgRNA library was co-transduced with sgZmat3 or sgTrp53, respectively. D Mean final tumor weight (error bars, s.d.) of in vivo screen samples normalized to wild-type (WT) average. Ordinary one-way ANOVA, **p < 0.01, ****p < 0.0001. E Heat map of log2 fold change screen phenotype of genes across all 3 screens from MEMcrispR analysis. *MEMcrispR FDR < 0.05.

Article Snippet: Extracts were run on 10% polyacrylamide SDS-PAGE gels, gels were transferred to PVDF membrane (Immobilon, Millipore), and membranes were blocked with 5% milk in TBST and probed with antibodies directed against ZMAT3 (sc-398712, 1:1000, Santa Cruz), p53 (CM5, 1:1000 Leica Novocastra), p21 (EPR18021, 1:1000, abcam), beta Actin (sc-47778, 1:1000, Santa Cruz), or GAPDH (10R-G109a, 1:20,000, Fitzgerald), followed by anti-mouse or anti-rabbit HRP-conjugated secondary antibodies (PI-2000-1 and PI-1000-1, 1:5,000, Vector Laboratories).

Techniques: In Vivo, CRISPR, Expressing, Control, Western Blot, Injection, Transduction

Journal: Cell death and differentiation

Article Title: Integrative multiomic approaches reveal ZMAT3 and p21 as conserved hubs in the p53 tumor suppression network.

doi: 10.1038/s41418-025-01513-8

Figure Lengend Snippet: Fig. 4 ZMAT3 and CDKN1A are essential and evolutionarily conserved tumor suppressors in the p53 target gene network. A Schematic of DepMap analyses for TP53 wild-type (WT) compared to TP53-deficient (Def) cell lines for both gene effect score (ΔEffect, calculated as WT minus Def) and differential gene expression (DGE). B Table of mean gene effect score in TP53 WT versus TP53-deficient cells for key components upstream and downstream of p53 in the p53 pathway. ΔEffect equals the mean gene effect score in TP53 WT minus the mean gene effect in TP53-deficient cells. False discovery rate (FDR) was calculated for the ΔEffect. C Plot of gene effect score for ZMAT3 and CDKN1A in TP53 WT and TP53-deficient cell lines in DepMap. ****p < 0.0001. D Table of mean gene effect score in TP53 WT versus TP53-deficient cells for classical p53 target genes. ΔEffect equals the mean gene effect score in TP53 WT minus the mean gene effect in TP53-deficient cells. False discovery rate (FDR) was calculated for the ΔEffect. E Volcano plot of differentially expressed genes in TP53 WT compared to TP53-deficient DepMap pan-cancer cell lines. log2FC significance shown at ≥1.5. F Plot of mean gene expression for ZMAT3 and CDKN1A in TP53 WT and TP53- deficient cell lines in DepMap. ****p < 0.0001. A Wilcoxon rank sum test was used in (C) and (F), which show the median bounded by the 1st and 3rd quartiles; whiskers that delimit the highest data point below the third quartile +1.5× the interquartile distance and the lowest data point above the first quartile −1.5× the interquartile distance.

Article Snippet: Extracts were run on 10% polyacrylamide SDS-PAGE gels, gels were transferred to PVDF membrane (Immobilon, Millipore), and membranes were blocked with 5% milk in TBST and probed with antibodies directed against ZMAT3 (sc-398712, 1:1000, Santa Cruz), p53 (CM5, 1:1000 Leica Novocastra), p21 (EPR18021, 1:1000, abcam), beta Actin (sc-47778, 1:1000, Santa Cruz), or GAPDH (10R-G109a, 1:20,000, Fitzgerald), followed by anti-mouse or anti-rabbit HRP-conjugated secondary antibodies (PI-2000-1 and PI-1000-1, 1:5,000, Vector Laboratories).

Techniques: Gene Expression

Journal: Cell death and differentiation

Article Title: Integrative multiomic approaches reveal ZMAT3 and p21 as conserved hubs in the p53 tumor suppression network.

doi: 10.1038/s41418-025-01513-8

Figure Lengend Snippet: Fig. 6 ZMAT3 and p21 deficiency cooperatively enhance cell migration in 3D, similar to p53 loss. A Schematic of 3D migration assay. (i) Serum starved E1A;HrasG12V MEFs are stained with octadecyl rhodamine B chloride (R18), (ii) embedded in a 3D collagen-I matrix, and, after 6 hours, are then tracked by live cell confocal microscopy for 22 hours. (iv) Track length and mean track speed are then calculated for individual cells. B 80 randomly selected cell trajectories shown for 3D migration experiments in E1A;HrasG12V;H11Cas9 MEFs during a ~ 22 h period. Grid size = 20 µm. Different colors represent the tracks for different individual cells. C Track length (µm) and D mean speed (µm/min) for 3D migration experiments. (Control n = 746 cells, Cdkn1a-KO n = 701 cells, Zmat3-KO n = 659 cells, DKO n = 912 cells, p53-KO n = 833 cells). **p < 0.01, ***p < 0.001, ****p < 0.0001. Significance was calculated by Kruskal-Wallis test with Dunn’s multiple comparisons test. Violin plots show a median with 1st and 3rd quartiles.

Article Snippet: Extracts were run on 10% polyacrylamide SDS-PAGE gels, gels were transferred to PVDF membrane (Immobilon, Millipore), and membranes were blocked with 5% milk in TBST and probed with antibodies directed against ZMAT3 (sc-398712, 1:1000, Santa Cruz), p53 (CM5, 1:1000 Leica Novocastra), p21 (EPR18021, 1:1000, abcam), beta Actin (sc-47778, 1:1000, Santa Cruz), or GAPDH (10R-G109a, 1:20,000, Fitzgerald), followed by anti-mouse or anti-rabbit HRP-conjugated secondary antibodies (PI-2000-1 and PI-1000-1, 1:5,000, Vector Laboratories).

Techniques: Migration, Staining, Confocal Microscopy, Control

Journal: Cell death and differentiation

Article Title: Integrative multiomic approaches reveal ZMAT3 and p21 as conserved hubs in the p53 tumor suppression network.

doi: 10.1038/s41418-025-01513-8

Figure Lengend Snippet: Fig. 7 ZMAT3 and p21 are key effectors of p53-mediated tumor suppression. A Schematic for cooperative tumor suppression between Zmat3 and Cdkn1a downstream of p53. X denotes components that might cooperate with ZMAT3 and p21. B In p53- deficient settings, transcriptional activation of target genes, like Zmat3, Cdkn1a, and X, diminishes and causes changes to cellular behavior that promote cancer progression.

Article Snippet: Extracts were run on 10% polyacrylamide SDS-PAGE gels, gels were transferred to PVDF membrane (Immobilon, Millipore), and membranes were blocked with 5% milk in TBST and probed with antibodies directed against ZMAT3 (sc-398712, 1:1000, Santa Cruz), p53 (CM5, 1:1000 Leica Novocastra), p21 (EPR18021, 1:1000, abcam), beta Actin (sc-47778, 1:1000, Santa Cruz), or GAPDH (10R-G109a, 1:20,000, Fitzgerald), followed by anti-mouse or anti-rabbit HRP-conjugated secondary antibodies (PI-2000-1 and PI-1000-1, 1:5,000, Vector Laboratories).

Techniques: Activation Assay