rabbit anti map2k3 antibody  (Boster Bio)

Journal: Cancer Management and Research

Article Title: Comprehensive Analysis of 5-Methylcytosine Profiles of Messenger RNA in Human High-Grade Serous Ovarian Cancer by MeRIP Sequencing

doi: 10.2147/CMAR.S319312

Figure Lengend Snippet: Relationship between MAP2K3 and HGSOC clinical prognosis. ( A ) The expression of MAP2K3 in three pairs of HGSOC tissue specimens (×200). ( B ) Low and high expression of MAP2K3 was detected by IHC in 80 clinical HGSOC samples (×200). ( C ) Association of overall survival with MAP2K3 expression.

Article Snippet: Anti-rabbit MAP2K3 (1:500, Bioss Antibodies, USA) was used as the primary antibody.

Techniques: Expressing

Journal: Cancer Management and Research

Article Title: Comprehensive Analysis of 5-Methylcytosine Profiles of Messenger RNA in Human High-Grade Serous Ovarian Cancer by MeRIP Sequencing

doi: 10.2147/CMAR.S319312

Figure Lengend Snippet: Clinicopathological Variables and the Expression Status of MAP2K3 in HGSOC Patients

Article Snippet: Anti-rabbit MAP2K3 (1:500, Bioss Antibodies, USA) was used as the primary antibody.

Techniques: Expressing

Journal: International Journal of Molecular Sciences

Article Title: Quadruple and Truncated MEK3 Mutants Identified from Acute Lymphoblastic Leukemia Promote Degradation and Enhance Proliferation

doi: 10.3390/ijms222212210

Figure Lengend Snippet: Linear representation of MEK3 indicating location of ALL mutations. Schematic diagram of MEK3 showing mutations identified from the ALL patient library. Amino acids are shown located within structural domains of MEK3.

Article Snippet: Samples were separated in 4–20% SDS-PAGE, transferred to a PVDF membrane, and immunoblotted as previously described [ ] using the following antibodies: MEK3 pT222 (OriGene, Rockville, MD, USA) or p38 ppT180/Y182 (cell signaling 4115S).

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: Quadruple and Truncated MEK3 Mutants Identified from Acute Lymphoblastic Leukemia Promote Degradation and Enhance Proliferation

doi: 10.3390/ijms222212210

Figure Lengend Snippet: Quadruple and truncated MEK3 mutants decrease protein stability. Cells were not transfected (NT) or transiently transfected concurrently with WT-cMYC or MEK3-cMYC mutant constructs. ( A ) Total cell lysates (TCL) from HEK293 ( B ) and HEPG2 cells transfected with either WT, quadruple (4M), or truncated (Δ) MEK3 constructs, separated by SDS-PAGE (4–20%) and immunoblotted (WB) as indicated. MEK3 protein expression band intensities were normalized to GAPDH using densitometric analysis. ( C ) Quantification of MEK3 mutant protein expression relative to WT in HEK293 and ( D ) in HEPG2 cells. Data are presented as mean ± SEM ( n = 3). One-way ANOVA with post hoc Tukey’s test for multiple comparisons was used to determine statistical significance * p < 0.01. ( E ) Immunofluorescence of transfected HEK293 cells. Subcellular localization of MEK3 was determined by staining for cMYC (middle panel), and nuclei using DAPI (left panel). Right panel shows merged images. Scale bar 20 μM.

Article Snippet: Samples were separated in 4–20% SDS-PAGE, transferred to a PVDF membrane, and immunoblotted as previously described [ ] using the following antibodies: MEK3 pT222 (OriGene, Rockville, MD, USA) or p38 ppT180/Y182 (cell signaling 4115S).

Techniques: Transfection, Mutagenesis, Construct, SDS Page, Expressing, Immunofluorescence, Staining

Journal: International Journal of Molecular Sciences

Article Title: Quadruple and Truncated MEK3 Mutants Identified from Acute Lymphoblastic Leukemia Promote Degradation and Enhance Proliferation

doi: 10.3390/ijms222212210

Figure Lengend Snippet: Quadruple MEK3 mutant was degraded at an accelerated rate compared to wild type. HEK293 cells were not transfected (−) or transiently transfected with ( A ) WT-cMYC or ( B ) 4M-MEK3-cMYC mutant construct. Twenty-four hours post-transfection, cells were left untreated (−) or treated with CHX (+) for the indicated time points. Cell lysates were collected and separated by SDS-PAGE and immunoblotted (WB), as indicated. ( C ) Quantified protein expression of WT-MEK3 or 4M-MEK3 is shown relative to time 0. Band intensities were normalized to GAPDH using densitometric analysis and presented as a ratio of cMYC to GAPDH. Data are presented as mean ± SEM ( n = 3). A t-test (unpaired) was used to determine statistical significance. *, p < 0.05.

Article Snippet: Samples were separated in 4–20% SDS-PAGE, transferred to a PVDF membrane, and immunoblotted as previously described [ ] using the following antibodies: MEK3 pT222 (OriGene, Rockville, MD, USA) or p38 ppT180/Y182 (cell signaling 4115S).

Techniques: Mutagenesis, Transfection, Construct, SDS Page, Expressing

Journal: International Journal of Molecular Sciences

Article Title: Quadruple and Truncated MEK3 Mutants Identified from Acute Lymphoblastic Leukemia Promote Degradation and Enhance Proliferation

doi: 10.3390/ijms222212210

Figure Lengend Snippet: Cysteine/serine protease inhibitors rescue quadruple MEK3 mutant protein expression. HEK293 cells were not transfected (−) or transiently transfected with WT-cMYC or 4M-MEK3-cMYC mutant constructs. At 24 hours post-transfection, cells were left untreated (−) or treated with CHX or a combination with proteolytic inhibitors for the indicated time points. ( A , B ) WT-MEK3 was treated with MG132. 4M-MEK3 was treated with either ( C , D ) CQ, ( E , F ) MG132, or ( G , H ) cysteine/serine protease inhibitors. Cell lysates were separated by SDS-PAGE and immunoblotted (WB) as indicated. Quantified protein expression of WT-MEK3 or 4M-MEK3 is shown relative to time 0 (Lower panels B , D , F and H ). Band intensities were normalized to GAPDH using densitometric analysis and presented as a ratio of cMYC to GAPDH. Data are presented as mean ± SEM ( n = 3 and n = 2 for Panel H). A t-test (unpaired) was used to determine statistical significance. *, p < 0.05.

Article Snippet: Samples were separated in 4–20% SDS-PAGE, transferred to a PVDF membrane, and immunoblotted as previously described [ ] using the following antibodies: MEK3 pT222 (OriGene, Rockville, MD, USA) or p38 ppT180/Y182 (cell signaling 4115S).

Techniques: Mutagenesis, Expressing, Transfection, Construct, SDS Page

Journal: International Journal of Molecular Sciences

Article Title: Quadruple and Truncated MEK3 Mutants Identified from Acute Lymphoblastic Leukemia Promote Degradation and Enhance Proliferation

doi: 10.3390/ijms222212210

Figure Lengend Snippet: Degradation of truncated MEK3 mutant by the proteasome is ubiquitin-independent . HEK293 cells were not transfected (NT) or transiently transfected concurrently with WT-cMYC or MEK3-cMYC constructs (4M-MEK3 and Δ-MEK3). After 24 hours of transfection, cells were left untreated (−) or treated with MG132 (+) as per the indicated time points. ( A ) Cell lysates were separated by SDS-PAGE and immunoblotted as indicated. ( B ) Transfected cells were treated with DMSO (−) or MG132 (+) for 8 h and immunoprecipitated (IP) for cMYC, and ( C ) probed using cell lysate. Samples were separated by SDS-PAGE and immunoblotted (WB) as indicated. ( D ) Quantified protein expression of WT-MEK3 or MEK3 constructs is shown relative to time 0. Band intensities were normalized to GAPDH using densitometric analysis and presented as a ratio of cMYC to GAPDH. Data are presented as mean ± SEM ( n = 3). T-test (unpaired) was used to determine statistical significance. *, p < 0.05.

Article Snippet: Samples were separated in 4–20% SDS-PAGE, transferred to a PVDF membrane, and immunoblotted as previously described [ ] using the following antibodies: MEK3 pT222 (OriGene, Rockville, MD, USA) or p38 ppT180/Y182 (cell signaling 4115S).

Techniques: Mutagenesis, Ubiquitin Proteomics, Transfection, Construct, SDS Page, Immunoprecipitation, Expressing

Journal: International Journal of Molecular Sciences

Article Title: Quadruple and Truncated MEK3 Mutants Identified from Acute Lymphoblastic Leukemia Promote Degradation and Enhance Proliferation

doi: 10.3390/ijms222212210

Figure Lengend Snippet: MEK3 mutants disrupt auto-phosphorylation at T222 and reduce T180/Y182 phosphorylation of p38 MAPK. HEK293 cells were not transfected (NT) or transiently transfected concurrently with WT-MEK3 or MEK3-cMYC constructs (4M-MEK3 and Δ-MEK3). ( A ) Immunoprecipitated (IP) cMYC or cell lysate was separated by SDS-PAGE and immunoblotted (WB) as indicated. ( B ) Phospho-p38 band intensities were normalized to total p38 by densitometric analysis. Data are presented as a ratio of cMYC to GAPDH. Quantification of phospho-p38 is shown relative to WT. Representative data for n = 3 are presented as mean ± SEM. One-way ANOVA with post hoc Tukey’s test for multiple comparisons was used to determine statistical significance. *, p < 0.05.

Article Snippet: Samples were separated in 4–20% SDS-PAGE, transferred to a PVDF membrane, and immunoblotted as previously described [ ] using the following antibodies: MEK3 pT222 (OriGene, Rockville, MD, USA) or p38 ppT180/Y182 (cell signaling 4115S).

Techniques: Phospho-proteomics, Transfection, Construct, Immunoprecipitation, SDS Page

Journal: International Journal of Molecular Sciences

Article Title: Quadruple and Truncated MEK3 Mutants Identified from Acute Lymphoblastic Leukemia Promote Degradation and Enhance Proliferation

doi: 10.3390/ijms222212210

Figure Lengend Snippet: Quadruple and truncated MEK3 mutants exhibited less suppressive activity on the viability of HEK293 cells. HEK293 cells were non-transfected (NT) or transiently transfected concurrently with WT-cMYC or MEK3-cMYC mutant constructs (4M-MEK3 and Δ-MEK3). Cell viability was measured at 24 h using MTS. Values from two independent experiments are presented as percent viability (mean ± SEM). One-way ANOVA with post hoc Tukey’s test for multiple comparisons was used to determine statistical significance *, p < 0.01.

Article Snippet: Samples were separated in 4–20% SDS-PAGE, transferred to a PVDF membrane, and immunoblotted as previously described [ ] using the following antibodies: MEK3 pT222 (OriGene, Rockville, MD, USA) or p38 ppT180/Y182 (cell signaling 4115S).

Techniques: Activity Assay, Transfection, Mutagenesis, Construct

Journal: Circulation

Article Title: Cardiac Fibroblast–Specific Activating Transcription Factor 3 Protects Against Heart Failure by Suppressing MAP2K3-p38 Signaling

doi: 10.1161/circulationaha.116.024599

Figure Lengend Snippet: Figure 6: CHIP analysis identifies Map2K3 as an ATF3 target gene

Article Snippet: The following antibodies were used: ATF3 (SC-188) and Col 1a2 (SC-393573) from Santa Cruz Biotechnology; α-SMA (A5228) and α-actinin (A5044) from Sigma; HDAC1(AB7028), GFP (Ab13970), p-smad3 (Ab52903), Vimentin (Ab8978), CD31 (Ab393573) purchased from Abcam; H4AC (8647), Map2k3 (8535S), p-p38 (4511), Tp38 (9212S), p-MK2 (3007), T-MK2 (3042S), p-JNK1/2 (4668S); T-JNK1/2 (9258S), p-ERK1/2 (4370), T-ERK1/2 (4695), IgG (14708) from Cell Signaling Technology.

Techniques:

Journal: Circulation

Article Title: Cardiac Fibroblast–Specific Activating Transcription Factor 3 Protects Against Heart Failure by Suppressing MAP2K3-p38 Signaling

doi: 10.1161/circulationaha.116.024599

Figure Lengend Snippet: Figure 7: ATF3 deficiency-induced ventricular remodeling is dependent upon Map2K3

Article Snippet: The following antibodies were used: ATF3 (SC-188) and Col 1a2 (SC-393573) from Santa Cruz Biotechnology; α-SMA (A5228) and α-actinin (A5044) from Sigma; HDAC1(AB7028), GFP (Ab13970), p-smad3 (Ab52903), Vimentin (Ab8978), CD31 (Ab393573) purchased from Abcam; H4AC (8647), Map2k3 (8535S), p-p38 (4511), Tp38 (9212S), p-MK2 (3007), T-MK2 (3042S), p-JNK1/2 (4668S); T-JNK1/2 (9258S), p-ERK1/2 (4370), T-ERK1/2 (4695), IgG (14708) from Cell Signaling Technology.

Techniques:

Journal: Journal of Virology

Article Title: A Kinome-Wide Small Interfering RNA Screen Identifies Proviral and Antiviral Host Factors in Severe Acute Respiratory Syndrome Coronavirus Replication, Including Double-Stranded RNA-Activated Protein Kinase and Early Secretory Pathway Proteins

doi: 10.1128/JVI.01029-15

Figure Lengend Snippet: Evaluation of the antiviral hits CDK6, MAP2K1, and MAP2K3. 293/ACE2 cells were transfected with four individual siRNAs targeting, MAP2K1 (A and B), or MAP2K3 (C and D) or three CDK6-specific siRNAs (E and F). A nontargeting scrambled siRNA was used as a control. At 48 h p.t. cells were infected with SARS-CoV-GFP at an MOI of 10 (A, C, and E) and fixed 24 h later, and GFP fluorescence (black bars) was quantified and normalized to the value measured in infected, scrambled siRNA-transfected cells (100%). The effect of siRNA transfection on cell viability was analyzed in parallel (gray bars), and values were normalized to those of scrambled siRNA-transfected control cells (100%). Each experiment was repeated at least three times (averages ± standard deviations [SD]). In parallel, siRNA-transfected cells were infected with wt SARS-CoV (MOI, 5), and at 8 h p.i., SARS-CoV N expression was monitored by Western blotting (B, D, and F). TfR was used as a loading control. Knockdown levels of the host proteins were analyzed by Western blotting (B, D, and F). The amount of SARS-CoV N protein and remaining quantity of host protein compared to that of scrambled siRNA-transfected cells (100%) is shown below each lane. All experiments were repeated at least twice.

Article Snippet: The following antisera against cellular proteins were used: rabbit anti-PKR (no. 610764; BD Biosciences), goat anti-COPB2 (sc-13332; Santa-Cruz), rabbit anti-CDK6 (sc-177; Santa Cruz), rabbit anti-MAP2K1 (710446; Life Technologies), rabbit anti-MAP2K3 (sc-961; Santa Cruz), and mouse monoclonal antibodies against β-actin (A5316; Sigma) and the transferrin receptor (TfR; no. 13-6890; Invitrogen).

Techniques: Transfection, Infection, Fluorescence, Expressing, Western Blot

Journal: BMC cancer

Article Title: MicroRNA-21 promotes hepatocellular carcinoma HepG2 cell proliferation through repression of mitogen-activated protein kinase-kinase 3.

doi: 10.1186/1471-2407-13-469

Figure Lengend Snippet: Figure 1 Immunohistochemistry (IHC) staining determined MAP2K3 expression in human HCC tumor and matched adjacent tissues. A-D: Representative images of MAP2K3 protein expression determined by IHC staining. A: An image represented a negative (-) expression of MAP2K3 expression; B: An image represented a low level (+) expression of MAP2K3, which showed a weak immunoreactive staining in cytoplasm; C: An image represented a negative (++) expression of MAP2K3 expression; D: An image represented a high level (+++) expression of MAP2K3, which exhibited a strong IHC staining in cytoplasm and perinuclear localization (arrowhead). E: Semi-quantitative analysis of MAP2K3 protein expression using integrated absorbance (IA) in human HCC tissues. Value was expressed as the average values from each individual sample of HCC tumor tissues or its matched adjacent tissue. The total average value of IA in the HCC tumor tissues was significantly greater as compared with the matched adjacent tissues (p<0.05, n = 14). Data was expressed as mean ± SD for 14 sets of samples.

Article Snippet: The membranes were probed with rabbit anti-MAP2K3 antibody and anti-GAPDH antibody (Boster, Wuhan, China) or (1:200, Boster, Wuhan, China) were for the interested protein MAP2K3 and endogenous GAPDH for loading control, respectively.

Techniques: Immunohistochemistry, Expressing, Staining

Journal: BMC cancer

Article Title: MicroRNA-21 promotes hepatocellular carcinoma HepG2 cell proliferation through repression of mitogen-activated protein kinase-kinase 3.

doi: 10.1186/1471-2407-13-469

Figure Lengend Snippet: Figure 2 Validation of MAP2K3 mRNA as a target of miR-21. (A): Sequence of potential binding site of miR-21 in the 3’UTR of MAP2K3 mRNA (top panel), mutations were introduced into the binding site for generation of mutated MAP2K3 3’TUR (bottom panel). (B and C): Validation of miR-21 target using MAP2K3 3’UTR luciferase reporter. Cells co-transfected with pMIR-Report/MAP2K3 3’UTR (WT) or pMIR-Report/ Mut-MAP2K3 3’UTR (Mut) and pAd/pri-miR-21 (B), pAd/miR-21/inhibitor (C), and pAd/con plasmids showed a decreased luciferase activity in pAd/pri-miR-21 cells (B). Luciferase activity after site directed mutagenesis of the 3’UTR of MAP2K3 mRNA in the miR-21 seed sequence (pMIR-Report/Mut-MAP2K3) was significantly higher with respect to the pMIR-Report/MAP2K3 vector transfected cells (B and C). Results represented the mean ± SD from three independent triplicated experiments (N=9).

Article Snippet: The membranes were probed with rabbit anti-MAP2K3 antibody and anti-GAPDH antibody (Boster, Wuhan, China) or (1:200, Boster, Wuhan, China) were for the interested protein MAP2K3 and endogenous GAPDH for loading control, respectively.

Techniques: Biomarker Discovery, Sequencing, Binding Assay, Luciferase, Transfection, Activity Assay, Mutagenesis, Plasmid Preparation

Journal: BMC cancer

Article Title: MicroRNA-21 promotes hepatocellular carcinoma HepG2 cell proliferation through repression of mitogen-activated protein kinase-kinase 3.

doi: 10.1186/1471-2407-13-469

Figure Lengend Snippet: Figure 4 miR-21 targets MAP2K3 mRNA. The HepG2 cells were infected with Ad/pri-miR-21, Ad/miR-21/inhibitor or Ad/con adenoviral vector. The expression of MAP2K3 was detected by immunoblotting analysis against anti-MAP2K3 antibody. Compared with Ad/con group, *: p<0.05. Data in A represented the mean ± SD from three independent triplicated experiments (N=9).

Article Snippet: The membranes were probed with rabbit anti-MAP2K3 antibody and anti-GAPDH antibody (Boster, Wuhan, China) or (1:200, Boster, Wuhan, China) were for the interested protein MAP2K3 and endogenous GAPDH for loading control, respectively.

Techniques: Infection, Plasmid Preparation, Expressing, Western Blot

Journal: Molecular and Cellular Biology

Article Title: SCO2 Induces p53-Mediated Apoptosis by Thr 845 Phosphorylation of ASK-1 and Dissociation of the ASK-1-Trx Complex

doi: 10.1128/MCB.06798-11

Figure Lengend Snippet: (A) The percent change in the ratios of phosphorylated (P) to total (T) MAP2K3 (M2K3), M2K6, M2K4, M2K7, JNK, and p38 was obtained for HCT p53+/+ cells (error bars represent standard deviations determined by analysis of variance; n = 7). (B) The change in the ratio of the above-mentioned proteins in HCT p53+/+ tumors was determined (error bars represent standard deviations determined by analysis of variance; n = 5).

Article Snippet: Anti-rabbit IgG antibodies for MAP2K3 (catalog number SAB4300391), phospho-MAP2K3 (catalog number SAB4300119), MAP2K6 (catalog number SAB4300422), and phospho-MAP2K6 (catalog number SAB4300120) were procured from Sigma-Aldrich (St. Louis, MO).

Techniques: