Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: ELK4 Promotes Colorectal Cancer Progression by Activating the Neoangiogenic Factor LRG1 in a Noncanonical SP1/3-Dependent Manner.

doi: 10.1002/advs.202303378

Figure Lengend Snippet: Figure 1. ELK4 is required for CRC cell proliferation and migration in vitro and for CRC tumor growth and metastasis in vivo. A,C) CCK8 A) and Transwell assays C) of HCT116 cells with ELK4 knockdown or overexpression. B) Representative images and statistical analysis of ELK4 knockdown HCT116 xenograft tumors (n = 6). D) Representative images of liver and lung metastases and hematoxylin-eosin staining. The arrow indicates the metastatic loci. The tumor/hematoxylin-eosin-stained field was used for statistical analysis. E) Representative images of colon tumors from WT and Elk4−/−mice after AOM/DSS model induction. Tumor numbers and sizes were statistically analyzed (n = 6 mice/group), and each symbol represents an individual mouse. F) Representative images and quantification of the size of organoids formed from WT and Elk4−/−large intestine stem cells (scale bars = 20 μm). Two-way ANOVA A), One-way ANOVA B,C) and Student’s t test C–F) were performed to assess the statistical significance. The data are presented as the mean ± S.D. values. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: AOM-DSS Model of Colorectal Tumorigenesis: Elk4 knockout mice on a C57BL genetic background were purchased from Cyagen Bioscience.

Techniques: Migration, In Vitro, In Vivo, Knockdown, Over Expression, Staining

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: ELK4 Promotes Colorectal Cancer Progression by Activating the Neoangiogenic Factor LRG1 in a Noncanonical SP1/3-Dependent Manner.

doi: 10.1002/advs.202303378

Figure Lengend Snippet: Figure 2. ELK4 promotes tumor angiogenesis in CRC. A) GO enrichment analysis of differentially expressed genes identified by RNA-seq in ELK4 knock- down HCT116 cells. B,C) Representative images and statistical analysis of tube formation B) and Transwell assays C) of HUVECs in the presence of CM from HCT116 cells with ELK4 knockdown or overexpression. D) Representative images and statistical analysis of IHC staining for CD31, HIF-1𝛼, and CD105 in xenografts derived from pLKO.1- and shELK4-transduced HCT116 cells. E) Correlation data between the mRNA level of ELK4 and those of CD31 and CD34 in the TCGA CRC dataset. Pearson correlation analysis was used to evaluate the associations. One-way ANOVA B–D) and Student’s t test B) were performed to assess the statistical significance. The data are presented as the mean ± S.D. values. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: AOM-DSS Model of Colorectal Tumorigenesis: Elk4 knockout mice on a C57BL genetic background were purchased from Cyagen Bioscience.

Techniques: RNA Sequencing, Knockdown, Over Expression, Immunohistochemistry, Derivative Assay

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: ELK4 Promotes Colorectal Cancer Progression by Activating the Neoangiogenic Factor LRG1 in a Noncanonical SP1/3-Dependent Manner.

doi: 10.1002/advs.202303378

Figure Lengend Snippet: Figure 3. SP1/3, instead of SRF, are coregulators of ELK4 in CRC. A) Motif enrichment analysis of the most significant ELK4-bound genes in HCT116 cells. B) Mass spectrometry (MS) identification of ELK4-interacting proteins in HEK293T cells. C) Venn diagram displaying the overlap between ELK4, SP1, and SP3 target genes in HCT116 cells. D) Heatmaps of ChIP-seq data for ELK4, SP1, and SP3 in HCT116 cells. All peaks in each heatmap are centered ± 2.0 kb from the ELK4 peaks in ELK4-bound target genes. E) Normalized read density (per bp per peak) for ELK4, SP1, and SP3 plotted in the region ± 2.0 kb from the ELK4-bound peaks. F) Heatmaps of ChIP-seq data for ELK4 and SRF peaks in the ELK4 (left) and SRF (right) targets. G) Normalized read density (per bp per peak) for ELK4 and SRF plotted in the region ± 2.0 kb from the ELK4-bound (left) or SRF-bound (right) peaks. H) Pie charts displaying

Article Snippet: AOM-DSS Model of Colorectal Tumorigenesis: Elk4 knockout mice on a C57BL genetic background were purchased from Cyagen Bioscience.

Techniques: Mass Spectrometry, ChIP-sequencing

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: ELK4 Promotes Colorectal Cancer Progression by Activating the Neoangiogenic Factor LRG1 in a Noncanonical SP1/3-Dependent Manner.

doi: 10.1002/advs.202303378

Figure Lengend Snippet: Figure 4. Serum stimulation-mediated phosphorylation of ELK4 facilitates its interaction with SP1 and SP3. A) Coimmunoprecipitation (Co-IP) to eval- uate the interactions between exogenous FLAG-ELK4 and endogenous SP1 and SP3 in HCT116 cells. B) Proximity ligation assay (PLA) to evaluate the interactions of ELK4 with SP1 and SP3 in HCT116 cells. The red dots indicate protein interactions (scale bars = 10 μm). C) Serum stimulation en- hances the interactions between ELK4 and SP1 and SP3 in HCT116 cells. The indicated plasmids were transfected, and cells were then treated with the indicated serum concentrations for 15 min. D) U0126 suppresses the serum-induced interactions between ELK4 and SP1 and SP3 in HCT116 cells. HCT116 cells were transfected with the indicated plasmids and were then cultured in the presence or absence of U0126 (10 μm) followed by stimula- tion with 10% serum for 15 min. E) HCT116 cells were transfected with ELK4 wild-type and P329A mutant plasmids for 24 h prior to stimulation with 10% serum for 15 min. F) HCT116 cells were transfected with wild-type ELK4 and various ELK4 mutant plasmids (4MA: T361/T366/S381/S387A, 4ME: T361/T366/S381/S387E) followed by stimulation with 10% serum for 15 min. G–I) HCT116 cells overexpressing WT ELK4 or 4MA mutant ELK4 were

Article Snippet: AOM-DSS Model of Colorectal Tumorigenesis: Elk4 knockout mice on a C57BL genetic background were purchased from Cyagen Bioscience.

Techniques: Phospho-proteomics, Co-Immunoprecipitation Assay, Proximity Ligation Assay, Transfection, Cell Culture, Mutagenesis

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: ELK4 Promotes Colorectal Cancer Progression by Activating the Neoangiogenic Factor LRG1 in a Noncanonical SP1/3-Dependent Manner.

doi: 10.1002/advs.202303378

Figure Lengend Snippet: Figure 5. LRG1 is the direct target of the ELK4-SP1/3 complex in CRC. A) Scatter plot showing that a set of genes was differentially expressed by both ELK4 knockdown and SP1/3 knockdown in HCT116 cells (P < 0.05). B,C) qPCR B) and western blot C) analyses of the LRG1 mRNA level in HCT116 cells with ELK4 knockdown or SP1/3 knockdown. D–F) Detection of the expression of ELK4 and LRG1 in colorectal cancer tissues derived from WT and Elk4−/−mice by western blot D), qPCR E), and IHC F) analyses. G) Schematic depiction of the LRG1 gene locus showing the potential LRG1 enhancer and promoter, as identified by ChIP-seq data of H3K27ac (GSM2058026) and H3K4me1 (GSM2712765). The ELK4 (red), SP1 (green), SP3 (purple), H3K27ac (orange), and H3K4me1 (blue) peaks and potential binding sites are shown. H) Luciferase assay of HCT116 cells cotransfected with the LRG1 reporter containing the promoter and enhancer and empty vector or the indicated constructs of ELK4, SP1, and SP3. I) Luciferase assay of HCT116 cells cotransfected with the LRG1 reporter containing the promoter and enhancer and empty vector, the WT ELK4 plasmid, or mutant ELK4 plasmids. J,K) CM was collected from ELK4 knockdown HCT116 cells stimulated with 500 ng mL−1 rLRG1 J) or modified by LRG1 overexpression K) and was then applied to HUVECs for the tube formation assay. L) Representative images of xenograft tumors derived from ELK4 knockdown HCT116 cells overexpressing LRG1 are shown (left); xenografts were weighed for statistical quantification (right). Student’s t test E), and One-way ANOVA B, H–L) were performed to assess the statistical significance. The data are presented as the mean ± S.D. values. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: AOM-DSS Model of Colorectal Tumorigenesis: Elk4 knockout mice on a C57BL genetic background were purchased from Cyagen Bioscience.

Techniques: Knockdown, Western Blot, Expressing, Derivative Assay, ChIP-sequencing, Binding Assay, Luciferase, Plasmid Preparation, Construct, Mutagenesis, Over Expression, Tube Formation Assay

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: ELK4 Promotes Colorectal Cancer Progression by Activating the Neoangiogenic Factor LRG1 in a Noncanonical SP1/3-Dependent Manner.

doi: 10.1002/advs.202303378

Figure Lengend Snippet: Figure 7. Association of ELK4 upregulation with poor prognosis and the clinical relevance of the ELK4/SP1/3-LRG1 axis in CRC. A) Relative ELK4 mRNA levels in CRC tumor and normal tissues were analyzed based on the TCGA database (left) and GSE20196 (right). B,C) Relative mRNA B) and protein C) levels of ELK4 in 24 pairs of CRC and adjacent normal tissues. D) Representative images (left) and quantitative analysis (right) of 190 pairs of CRC tissues in the TMA cohort based on ELK4 IHC staining. E) Kaplan‒Meier plots of overall survival (up) and disease-free survival (down) for 190

Article Snippet: AOM-DSS Model of Colorectal Tumorigenesis: Elk4 knockout mice on a C57BL genetic background were purchased from Cyagen Bioscience.

Techniques: Immunohistochemistry

Journal: Journal of Personalized Medicine

Article Title: Exosomes from Microvascular Endothelial Cells under Mechanical Unloading Inhibit Osteogenic Differentiation via miR-92b-3p/ELK4 Axis

doi: 10.3390/jpm12122030

Figure Lengend Snippet: ELK4 is a direct target of miR-92b-3p and is responsible for miR-92b-3p-mediated suppression of osteogenic differentiation in MC3T3-E1 cells. ( A ) qRT–PCR analysis of ELK4 mRNA expression in MC3T3-E1 cells after transfection of mimic-92b-3p, inhibitor-92b-3p, or the corresponding control ( n = 3). ( B ) Western blotting analysis of the protein expression of ELK4 in MC3T3-E1 cells ( n = 3). ( C ) The relative luciferase activities of the ELK4 WT and MUT reporters were assessed after 293T cells were treated for 48 h with mimic-92b-3p and the equivalent controls ( n = 3). ( D ) Schematic representation of the luciferase reporters containing ELK4 3′-UTR WT or MUT sequences. ( E ) mRNA levels of ELK4 analyzed by qRT–PCR in MC3T3-E1 cells treated with Con Exos/Clino Exos (200 μg/mL) ( n = 3). ( F ) Protein levels of ELK4 analyzed by Western blotting ( n = 3). ( G ) qRT–PCR analysis of ALP, Osx, Runx2, and Ocn in MC3T3-E1 cells after the co-transfection of inhibitor-92b-3p, si-ELK4 and their negative controls in MC3T3-E1 cells ( n = 3). ( H ) Western blotting analysis of Osx, Runx2, and Ocn expression in MC3T3-E1 cells ( n = 3). ( I ) ALP activity analysis in MC3T3-E1 cells ( n = 3). ( J ) Representative images of ALP staining in MC3T3-E1 cells ( n = 3). * p < 0.05, ** p < 0.01 vs. control.

Article Snippet: After incubation with 5% skim milk (5% w / v ) for 2 h at room temperature, the membranes were co-incubated overnight at 4 °C with the following primary antibodies specific for GAPDH (1:1000; Cell Signaling Technology, USA), Runx2 (1:1000; Cell Signaling Technology, Danvers, MA, USA), Osx (1:1000; Abcam, Cambridge, UK), Ocn (1:2000; Abcam, UK), ELK4 (1:1000; Proteintech, Rosemont, IL, USA), GM130 (1:1000; Proteintech, USA), CD9 (1:1000; Proteintech, USA), and TSG101 (1:1000; Proteintech, USA).

Techniques: Quantitative RT-PCR, Expressing, Transfection, Western Blot, Luciferase, Cotransfection, Activity Assay, Staining

Journal: Journal of Personalized Medicine

Article Title: Exosomes from Microvascular Endothelial Cells under Mechanical Unloading Inhibit Osteogenic Differentiation via miR-92b-3p/ELK4 Axis

doi: 10.3390/jpm12122030

Figure Lengend Snippet: A schematic diagram illustrating the molecular mechanisms of which exosomes derived from MVECs cultured under mechanical unloading regulate osteogenic differentiation. miR-92b-3p expression was increased in MVEC-secreted exosomes after mechanical unloading, resulting in the upregulation of miR-92b-3p expression in MC3T3-E1 cells cocultured with Clino Exos. ELK4, the direct target of miR-92b-3p, is decreased in MC3T3-E1 cells treated with Clino Exos, thus inhibiting osteogenic differentiation. The blue arrow represents inhibition, and the red arrow represents promotion.

Article Snippet: After incubation with 5% skim milk (5% w / v ) for 2 h at room temperature, the membranes were co-incubated overnight at 4 °C with the following primary antibodies specific for GAPDH (1:1000; Cell Signaling Technology, USA), Runx2 (1:1000; Cell Signaling Technology, Danvers, MA, USA), Osx (1:1000; Abcam, Cambridge, UK), Ocn (1:2000; Abcam, UK), ELK4 (1:1000; Proteintech, Rosemont, IL, USA), GM130 (1:1000; Proteintech, USA), CD9 (1:1000; Proteintech, USA), and TSG101 (1:1000; Proteintech, USA).

Techniques: Derivative Assay, Cell Culture, Expressing, Inhibition

Journal: Frontiers in Immunology

Article Title: ELK4 exerts opposite roles in cytokine/chemokine production and degranulation in activated mast cells

doi: 10.3389/fimmu.2023.1171380

Figure Lengend Snippet: Elk4 is expressed and downregulated in activated mast cells. (A) Scatterplot displaying the induced (red) and downregulated genes (green) after FcϵRI-mediated stimulation in human mast cells (|Log2 fold change|≥1, adj p<0.05). The transcripts per kilobase of exon model per million mapped reads (TPM) were extracted from the GSE107316 dataset. (B) Gene ontology analysis of FcϵRI crosslinking-induced and FcϵRI crosslinking-downregulated genes by using the MSigDB hallmark gene set signatures and TCF target gene signature. Hypergeometrical tests were used to assess statistical significance. Bonferroni-adjusted p values are shown. (C) The mRNA levels of ELK1 , ELK3 , and ELK4 in unstimulated, IgE-sensitized, and IgE-DNP/HSA-stimulated human mast cells are shown. TPM data were extracted from the GSE107316 dataset. (D) qPCR analysis of Elk1 , Elk3 , and Elk4 mRNA expression in unstimulated, IgE-DNP/HSA-stimulated and Compound 48/80-stimulated BMMCs. One-way ANOVA with Dunnett’s multiple comparison test was used to assess statistical significance. Bar, mean; error bar, SD; n = 3; *p<0.05; **p<0.01. (E) Western blot analysis of ELK4 protein expression in unstimulated, IgE-DNP/HSA-stimulated and Compound 48/80-stimulated BMMCs.

Article Snippet: The following antibodies were used for endogenous co-IP and WB analysis: SIRT6 (Cell Signaling Technology, 12486), ELK4 (Atlas antibodies, HPA028863), and MITF (Cell Signaling Technology, 97800).

Techniques: Expressing, Comparison, Western Blot

Journal: Frontiers in Immunology

Article Title: ELK4 exerts opposite roles in cytokine/chemokine production and degranulation in activated mast cells

doi: 10.3389/fimmu.2023.1171380

Figure Lengend Snippet: Elk4 deficiency leads to cell cycle arrest in BMMCs. (A) The proliferation of WT BMMCs and Elk4 KO BMMCs was assessed by CCK8 assay at day 1, day 3 and day 5. Two-way ANOVA with Dunnett’s multiple comparison test was used to assess statistical significance. Bar, mean; error bar, SD; n=3; *p, < 0.05; ***, p<0.001. (B, C) Cell cycle analysis of WT BMMCs and Elk4 KO BMMCs cultured in the presence of IL3 with or without SCF. Flow cytometry data (B) and statistical data (C) are shown. Bar, mean; error bar, SD; n=3; *p, < 0.05; **p<0.01. (D) Percentage of PCMCs in CD45-positive cells from the peritoneal cavities of WT and Elk4 KO mice. Cells were stained with CD45-eFluor 450, CD117-FITC, and FcϵRI-PECY7 and analyzed by flow cytometry. (E) Toluidine blue staining of skin tissues from WT and Elk4 KO mice. The mast cell number (blue)/high-power field (HPF) was used for statistical analysis. Bar, mean; error bar, SD; n=4; ***p<0.001.

Article Snippet: The following antibodies were used for endogenous co-IP and WB analysis: SIRT6 (Cell Signaling Technology, 12486), ELK4 (Atlas antibodies, HPA028863), and MITF (Cell Signaling Technology, 97800).

Techniques: CCK-8 Assay, Comparison, Cell Cycle Assay, Cell Culture, Flow Cytometry, Staining

Journal: Frontiers in Immunology

Article Title: ELK4 exerts opposite roles in cytokine/chemokine production and degranulation in activated mast cells

doi: 10.3389/fimmu.2023.1171380

Figure Lengend Snippet: Hdc , Tnfα , Il6 , Ccl3 , and Ccl4 mRNA expression was impaired in Elk4 KO BMMCs in response to FcϵRI-mediated and Compound 48/80-induced activation. (A) qPCR analysis of Hdc , Il6 , Tnfα , Ccl3 , and Ccl4 mRNA levels in BMMCs derived from Elk4 wild-type (WT), heterozygous (HZ), and homozygous (KO) mice in response to FcϵRI-mediated activation. BMMCs were sensitized with anti-DNP-IgE(0.5 μg/ml) overnight and stimulated with DNP-HSA (100 ng/ml) for 1 hour. Bar, mean; error bar, SD; n=3; ****p<0.0001. (B) qPCR analysis of the Hdc , Ccl3 , and Ccl4 mRNA levels in BMMCs derived from Elk4 WT, HZ, and KO mice in response to Compound 48/80-induced activation. One-way ANOVA with Dunnett’s multiple comparison test was used to assess statistical significance in this figure. BMMCs were stimulated with Compound 48/80 (5 μg/ml) for 1 hour. Bar, mean; error bar, SD; n=3; *p < 0.05; **p<0.01; ***p<0.001.

Article Snippet: The following antibodies were used for endogenous co-IP and WB analysis: SIRT6 (Cell Signaling Technology, 12486), ELK4 (Atlas antibodies, HPA028863), and MITF (Cell Signaling Technology, 97800).

Techniques: Expressing, Activation Assay, Derivative Assay, Comparison

Journal: Frontiers in Immunology

Article Title: ELK4 exerts opposite roles in cytokine/chemokine production and degranulation in activated mast cells

doi: 10.3389/fimmu.2023.1171380

Figure Lengend Snippet: Elk4 deficiency promotes degranulation and histamine release. (A) Time course and dose-dependent curve detection of the beta-hexosaminidase releases. BMMCs were sensitized with anti-DNP-IgE (1 μg/ml) overnight, then stimulated with DNP-HSA (10, 100, 1000 ng/ml) for 60min (Left) or stimulated with DNP-HSA (100 ng/ml) for 5min, 15min, 30min and 60min (Right). Bar, mean; error bar, SD; n=4; **p<0.01. (B) The total contents of beta-hexosaminidase in Elk4 WT, HZ and KO BMMCs are shown. Bar, mean; error bar, SD; n=3; *p < 0.05; **p<0.01. (C, D) Degranulation of Elk4 WT, HZ and KO PCMCs was assessed by beta-hexosaminidase release assay. PCMCs were sensitized with anti-DNP-IgE (1 μg/ml) overnight and stimulated with DNP-HSA (100 ng/ml) for 1 hour. The released contents (A) and the total contents (B) of beta-hexosaminidase in Elk4 WT, HZ and KO PCMCs are shown. Bar, mean; error bar, SD; n=2; **p<0.01. (E, F) Degranulation of Elk4 WT, HZ and KO BMMCs (E) and PCMCs (F) was assessed by cell-surface LAMP1 staining using flow cytometry. Both BMMCs and PCMCs were sensitized with anti-DNP-IgE (1 μg/ml) overnight and stimulated with DNP-HSA (100 ng/ml) for 1 hour before LAMP1 staining. Bar, mean; error bar, SD; n=3; *p<0.05.

Article Snippet: The following antibodies were used for endogenous co-IP and WB analysis: SIRT6 (Cell Signaling Technology, 12486), ELK4 (Atlas antibodies, HPA028863), and MITF (Cell Signaling Technology, 97800).

Techniques: Release Assay, Staining, Flow Cytometry

Journal: Frontiers in Immunology

Article Title: ELK4 exerts opposite roles in cytokine/chemokine production and degranulation in activated mast cells

doi: 10.3389/fimmu.2023.1171380

Figure Lengend Snippet: The anaphylactic response in mice was alleviated in Elk4 -deficient mice. (A) Images showing the ear pinnae of Elk4 WT and KO mice. Evans blue dye extravasation from the ears of Elk4 WT and KO mice was examined 30 min after intravenous DNP-HSA (containing 1% Evans blue) administration. Evans blue dye extravasation was quantified by measuring the optical density at 610 nm (OD610nm)/weight. Bar, mean; error bar, SD; n=6; ***p<0.001. (B, C) WT and Elk4 KO mice were sensitized with 10 μg anti-DNP-IgE and challenged with 100 μg DNP-HSA. Changes in body temperature and serum levels of IL-6, TNFα and histamine were determined. Bar, mean; error bar, SD; n=3-5; *p<0.05; ***p<0.001. (D) HE staining, IHC analysis of MUC5AC and toluidine blue staining of lung sections from Elk4 WT and KO mice with OVA-induced asthma. The area of the bronchiole wall (Wa) and the perimeter of the bronchiole basement membrane were analyzed by ImageJ. Student’s t test was used to assess statistical significance in the figure. Bar, mean; error bar, SD; n=3; **p<0.01; ***p<0.001.

Article Snippet: The following antibodies were used for endogenous co-IP and WB analysis: SIRT6 (Cell Signaling Technology, 12486), ELK4 (Atlas antibodies, HPA028863), and MITF (Cell Signaling Technology, 97800).

Techniques: Staining, Membrane

Journal: Frontiers in Immunology

Article Title: ELK4 exerts opposite roles in cytokine/chemokine production and degranulation in activated mast cells

doi: 10.3389/fimmu.2023.1171380

Figure Lengend Snippet: Elk4 deficiency affects the FcϵRI-mediated transcriptional response in BMMCs. (A) Scatterplot displaying the induced (red) and downregulated genes (green) before and after FcϵRI-mediated stimulation in WT BMMCs (|Log2 fold change|≥1, adj p<0.05). (B) Gene ontology analysis of FcϵRI crosslinking-induced and FcϵRI crosslinking-downregulated genes in WT BMMCs by using the MSigDB hallmark gene set signatures and TCF target gene signature. Hypergeometrical tests were used to assess statistical significance. Bonferroni-adjusted p values are shown. (C) Heatmap representation of the relative mean expression levels of the 431 IgE-DNP/HSA-induced genes in Elk4 WT and KO BMMCs. (D) Identification of ELK4-dependent genes by comparison with the IgE-DNP/HSA induction ratio in Elk4 WT and KO BMMCs. ELK4-dependent genes (red, purple) exhibited a systematic relationship between their degree of induction in the two contexts, whereas the others (grey) did not. Slope and Spearman r value are indicated. (E) Gene ontology analysis of the ELK4-dependent (fold change>1.5 or <0.66, adj p<0.05 in Elk4 KO BMMCs) FcϵRI-crosslinking induced genes in BMMCs by using the MSigDB hallmark gene set signatures. Hypergeometrical tests were used to assess statistical significance. Bonferroni-adjusted p values are shown. (F) Heatmap representation of the relative mean expression levels of the differentially expressed genes between unstimulated Elk4 WT and KO BMMCs. Gene ontology analysis of the differentially expressed genes was further performed by using the MSigDB hallmark gene set signatures and MITF and TCF target gene signatures. Hypergeometrical tests were used to assess statistical significance. Bonferroni-adjusted p values are shown.

Article Snippet: The following antibodies were used for endogenous co-IP and WB analysis: SIRT6 (Cell Signaling Technology, 12486), ELK4 (Atlas antibodies, HPA028863), and MITF (Cell Signaling Technology, 97800).

Techniques: Expressing, Comparison

Journal: Frontiers in Immunology

Article Title: ELK4 exerts opposite roles in cytokine/chemokine production and degranulation in activated mast cells

doi: 10.3389/fimmu.2023.1171380

Figure Lengend Snippet: ELK4 interacts and might cooperate with MITF and SIRT6 to regulate mast cell activation and degranulation. (A) Coimmunoprecipitation analysis of endogenous MITF and ELK4 proteins in MC/9 mast cells. (B) qPCR analysis of the Hdc , Tnfα , Ccl3 , and Ccl4 mRNA levels in BMMCs treated with ML329 in response to FcϵRI-mediated stimulation. One-way ANOVA with Dunnett’s multiple comparison test was used to assess statistical significance. Bar, mean; error bar, SD; n=3; *p < 0.05; ****p<0.0001. (C) Coimmunoprecipitation analysis of endogenous SIRT6 and ELK4 proteins in MC/9 mast cells. (D) qPCR analysis of Syngr1 , Exoc3l1 , Cadm1 , and Vcl mRNA levels in Elk4 -deficient BMMCs. Student’s t test was used to assess statistical significance. Bar, mean; error bar, SD; n=3; **p<0.01. (E) qPCR analysis of Syngr1 , Exoc3l1 , Cadm1 , and Vcl mRNA levels in BMMCs treated with OSS-128167. * indicates p<0.05; ** indicates p <0.01;***indicates p<0.001. One-way ANOVA with Dunnett’s multiple comparison test was used to assess statistical significance. Bar, mean; error bar, SD; n=3; *p < 0.05; **p<0.01; ****p<0.0001. (F) Schematic diagram depicting the downregulation of Elk4 expression in activated mast cells and the effect and potential mechanism of ELK4 on mast cell activation.

Article Snippet: The following antibodies were used for endogenous co-IP and WB analysis: SIRT6 (Cell Signaling Technology, 12486), ELK4 (Atlas antibodies, HPA028863), and MITF (Cell Signaling Technology, 97800).

Techniques: Activation Assay, Comparison, Expressing

Journal: Oncotarget

Article Title: Novel transcription-induced fusion RNAs in prostate cancer

doi: 10.18632/oncotarget.17099

Figure Lengend Snippet: Nominated fusion transcripts

Article Snippet: The assays measuring the individual exons in ELK4 were as designed by AB (exon 1, Hs00360812; exon 2, Hs00360813_m1).

Techniques:

Journal: Oncotarget

Article Title: Novel transcription-induced fusion RNAs in prostate cancer

doi: 10.18632/oncotarget.17099

Figure Lengend Snippet: ( A ) and ( B ) show the expression of the 3′ partner genes, B3GNT6 and ELK4 , of the fusions ACER3-B3GNT6 and SLC45A3-ELK4 in 44 pairs of tumor and benign prostate samples. ( C ) and ( D ) show the expression of 3′ partner genes B3GNT6 and ELK4 in 50 additional prostate tumors. ( E ) and ( F ) show the correlation between the expression of ACER3-B3GNT6 and SLC45A3-ELK4 and their respective 3′ partner genes, B3GNT6 and ELK4 . X and Y axes represent log2 transformed expression values (FPKM, fragments per kilobase of transcript per million mapped reads; RPKM, reads per kilobase of transcript per million mapped reads).

Article Snippet: The assays measuring the individual exons in ELK4 were as designed by AB (exon 1, Hs00360812; exon 2, Hs00360813_m1).

Techniques: Expressing, Transformation Assay