Journal: International journal of oncology

Article Title: Death associated protein‑3 (DAP3) and DAP3 binding cell death enhancer‑1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance.

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Figure 1. IHC staining for the DAP3 and DELE1 in colon cancer tissues (n=52) and the adjacent tissues (n=62). (A) DAP3 expression and (B) DELE1 expres‑ sion in adjacent normal tissue, T3 and T4 colon cancer tissue with the magnification of x10 and x40, respectively. (C and D) Box plot presenting the differential (C) DAP3 and (D) DELE1 expression in adjacent tissue, early stage, and the advanced stage of colon cancer. *P<0.05, **P<0.01 and ***P<0.001. DAP3, death associated protein‑3; DELE1, DAP3 binding cell death enhancer‑1.

Article Snippet: Anti‐human DELE1 siRNA was purchased from Santa Cruz Biotechnologies, Inc. An anti‐human DAP3 ribozyme transgene that was specifically used to target and knockdown human DAP3 was prepared as previously reported (12).

Techniques: Immunohistochemistry, Expressing, Binding Assay

Journal: International journal of oncology

Article Title: Death associated protein‑3 (DAP3) and DAP3 binding cell death enhancer‑1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance.

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Figure 2. Kaplan‑Meier survival analyses showed correlations between overall survival, disease‑free survival, distant metastasis‑free survival, recurrence‑free survival and altered expression of DAP3, DELE1 and DAP3/DELE1 ratio. P‑values shown are from the log ranked test. DAP3, death associated protein‑3; DELE1, DAP3 binding cell death enhancer‑1.

Article Snippet: Anti‐human DELE1 siRNA was purchased from Santa Cruz Biotechnologies, Inc. An anti‐human DAP3 ribozyme transgene that was specifically used to target and knockdown human DAP3 was prepared as previously reported (12).

Techniques: Expressing, Binding Assay

Journal: International journal of oncology

Article Title: Death associated protein‑3 (DAP3) and DAP3 binding cell death enhancer‑1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance.

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Figure 3. Response to chemotherapy with differential transcript DELE1 and DAP3 respectively. This was based on the public database (www.rocplot.com). Shown in the figure is the median of the respective group. *P<0.05 by Mann‑Whitney U test. DAP3, death associated protein‑3; DELE1, DAP3 binding cell death enhancer‑1.

Article Snippet: Anti‐human DELE1 siRNA was purchased from Santa Cruz Biotechnologies, Inc. An anti‐human DAP3 ribozyme transgene that was specifically used to target and knockdown human DAP3 was prepared as previously reported (12).

Techniques: Mann-Whitney U-Test, Binding Assay

Journal: International journal of oncology

Article Title: Death associated protein‑3 (DAP3) and DAP3 binding cell death enhancer‑1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance.

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Figure 4. Correlation between DAP3 and DELE1 with angiogenesis biomarkers. (A and B) Correlation efficiency between DAP3 and angiogenesis markers in The Cancer Genome Atlas‑COAD cohort was presented in (A) heatmap (A) and (B) scatter plots. (C and D) Association between DELE1 and angiogenesis biomarkers was revealed in (C) heatmap and (D) scatter plots. DAP3, death associated protein‑3; DELE1, DAP3 binding cell death enhancer‑1. **P<0.01.

Article Snippet: Anti‐human DELE1 siRNA was purchased from Santa Cruz Biotechnologies, Inc. An anti‐human DAP3 ribozyme transgene that was specifically used to target and knockdown human DAP3 was prepared as previously reported (12).

Techniques: Binding Assay

Journal: International journal of oncology

Article Title: Death associated protein‑3 (DAP3) and DAP3 binding cell death enhancer‑1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance.

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Figure 5. Validation of the DAP3 and DELE1 knockdown in RKO cells by quantitative PCR gene transcript analysis. Left panel: Expression of DAP3 in the cell models. The levels of expression in wild type, control transfection (control plasmid and control siRNA), DAP3 knockdown, DELE1 knockdown and DAP3/DELE1 dual knockdown cells were compared. Control transfections resulted in minor and statistically insignificant changes of DAP3 when compared with wild type cells (P>0.05) Right panel: Expression of DELE1 in the cell models. The levels of expression in wild type, control transfection (control plasmid and control siRNA), DELE1 knockdown and DAP3/DELE1 dual knockdown cells were compared. Control transfections resulted in minor and statisti‑ cally insignificant changes of DELE1 when compared with wild type cells (P>0.05). Values presented in the figure are mean and standard deviation. Error bars represent standard deviation. **P<0.01 vs. RKO wild type cells, ΔΔP<0.001 vs. RKO‑control‑plasmid cells and ##P<0.01 VS RKO‑control‑siRNA cells. DAP3, death associated protein‑3; DELE1, DAP3 binding cell death enhancer‑1; siRNA, small interfering RNA.

Article Snippet: Anti‐human DELE1 siRNA was purchased from Santa Cruz Biotechnologies, Inc. An anti‐human DAP3 ribozyme transgene that was specifically used to target and knockdown human DAP3 was prepared as previously reported (12).

Techniques: Biomarker Discovery, Knockdown, Real-time Polymerase Chain Reaction, Expressing, Control, Transfection, Plasmid Preparation, Standard Deviation, Binding Assay, Small Interfering RNA

Journal: International journal of oncology

Article Title: Death associated protein‑3 (DAP3) and DAP3 binding cell death enhancer‑1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance.

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Figure 7. Metabolic of colorectal cancer cells following DAP3 and DELE1 knockdown. (A) Concentrations of extracellular glucose in different RKO cells were determined by luminescence measurement. (B) NAD(P)H measurements quantified by luminescence in different RKO cells. (C) Differential extracellular lactate expression and the ratio in different RKO cells. (D) NO2 Standard curve and measurements of NO2 in different RKO cells. Error bar stands standard error of means. *P<0.05 and **P<0.01 by Student's t‑test. DAP3, death associated protein‑3; DELE1, DAP3 binding cell death enhancer‑1.

Article Snippet: Anti‐human DELE1 siRNA was purchased from Santa Cruz Biotechnologies, Inc. An anti‐human DAP3 ribozyme transgene that was specifically used to target and knockdown human DAP3 was prepared as previously reported (12).

Techniques: Knockdown, Expressing, Binding Assay

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Primer sequences using in the qPCR analyses.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Sequencing

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: IHC staining for the DAP3 and DELE1 in colon cancer tissues (n=52) and the adjacent tissues (n=62). (A) DAP3 expression and (B) DELE1 expression in adjacent normal tissue, T3 and T4 colon cancer tissue with the magnification of ×10 and ×40, respectively. (C and D) Box plot presenting the differential (C) DAP3 and (D) DELE1 expression in adjacent tissue, early stage, and the advanced stage of colon cancer. * P<0.05, ** P<0.01 and *** P<0.001. DAP3, death associated protein-3; DELE1, DAP3 binding cell death enhancer-1.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Immunohistochemistry, Expressing, Binding Assay

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Transcript levels of DAP3 and DELE1 in colorectal cancers.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques:

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Kaplan-Meier survival analyses showed correlations between overall survival, disease-free survival, distant metastasis-free survival, recurrence-free survival and altered expression of DAP3, DELE1 and DAP3/DELE1 ratio. P-values shown are from the log ranked test. DAP3, death associated protein-3; DELE1, DAP3 binding cell death enhancer-1.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Expressing, Binding Assay

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: The value of DAP3, DELE1, and clinical factors in predicting the overall, disease free, metastasis free, and recurrence free survivals of the patients.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques:

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Response to chemotherapy with differential transcript DELE1 and DAP3 respectively. This was based on the public database ( www.rocplot.com ). Shown in the figure is the median of the respective group. * P<0.05 by Mann-Whitney U test. DAP3, death associated protein-3; DELE1, DAP3 binding cell death enhancer-1.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: MANN-WHITNEY, Binding Assay

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Response to the chemotherapeutic drugs with differential DAP3 and DELE1 expression.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Expressing

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Correlation between DAP3 and DELE1 with angiogenesis biomarkers. (A and B) Correlation efficiency between DAP3 and angiogenesis markers in The Cancer Genome Atlas-COAD cohort was presented in (A) heatmap (A) and (B) scatter plots. (C and D) Association between DELE1 and angiogenesis biomarkers was revealed in (C) heatmap and (D) scatter plots. DAP3, death associated protein-3; DELE1, DAP3 binding cell death enhancer-1. ** P<0.01.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Binding Assay

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Correlation between DAP3 and angiogenic factors in the Beijing CRC cohort a .

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques:

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Validation of the DAP3 and DELE1 knockdown in RKO cells by quantitative PCR gene transcript analysis. Left panel: Expression of DAP3 in the cell models. The levels of expression in wild type, control transfection (control plasmid and control siRNA), DAP3 knockdown, DELE1 knockdown and DAP3/DELE1 dual knockdown cells were compared. Control transfections resulted in minor and statistically insignificant changes of DAP3 when compared with wild type cells (P>0.05) Right panel: Expression of DELE1 in the cell models. The levels of expression in wild type, control transfection (control plasmid and control siRNA), DELE1 knockdown and DAP3/DELE1 dual knockdown cells were compared. Control transfections resulted in minor and statistically insignificant changes of DELE1 when compared with wild type cells (P>0.05). Values presented in the figure are mean and standard deviation. Error bars represent standard deviation. ** P<0.01 vs. RKO wild type cells, ΔΔ P<0.001 vs. RKO-control-plasmid cells and ## P<0.01 VS RKO-control-siRNA cells. DAP3, death associated protein-3; DELE1, DAP3 binding cell death enhancer-1; siRNA, small interfering RNA.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Biomarker Discovery, Knockdown, Real-time Polymerase Chain Reaction, Expressing, Control, Transfection, Plasmid Preparation, Standard Deviation, Binding Assay, Small Interfering RNA

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Metabolic of colorectal cancer cells following DAP3 and DELE1 knockdown. (A) Concentrations of extracellular glucose in different RKO cells were determined by luminescence measurement. (B) NAD(P)H measurements quantified by luminescence in different RKO cells. (C) Differential extracellular lactate expression and the ratio in different RKO cells. (D) NO 2 Standard curve and measurements of NO 2 in different RKO cells. Error bar stands standard error of means. * P<0.05 and ** P<0.01 by Student's t-test. DAP3, death associated protein-3; DELE1, DAP3 binding cell death enhancer-1.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Knockdown, Expressing, Binding Assay

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Primer sequences using in the qPCR analyses.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Sequencing

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: IHC staining for the DAP3 and DELE1 in colon cancer tissues (n=52) and the adjacent tissues (n=62). (A) DAP3 expression and (B) DELE1 expression in adjacent normal tissue, T3 and T4 colon cancer tissue with the magnification of ×10 and ×40, respectively. (C and D) Box plot presenting the differential (C) DAP3 and (D) DELE1 expression in adjacent tissue, early stage, and the advanced stage of colon cancer. * P<0.05, ** P<0.01 and *** P<0.001. DAP3, death associated protein-3; DELE1, DAP3 binding cell death enhancer-1.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Immunohistochemistry, Expressing, Binding Assay

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Transcript levels of DAP3 and DELE1 in colorectal cancers.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques:

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Kaplan-Meier survival analyses showed correlations between overall survival, disease-free survival, distant metastasis-free survival, recurrence-free survival and altered expression of DAP3, DELE1 and DAP3/DELE1 ratio. P-values shown are from the log ranked test. DAP3, death associated protein-3; DELE1, DAP3 binding cell death enhancer-1.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Expressing, Binding Assay

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: The value of DAP3, DELE1, and clinical factors in predicting the overall, disease free, metastasis free, and recurrence free survivals of the patients.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques:

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Response to chemotherapy with differential transcript DELE1 and DAP3 respectively. This was based on the public database ( www.rocplot.com ). Shown in the figure is the median of the respective group. * P<0.05 by Mann-Whitney U test. DAP3, death associated protein-3; DELE1, DAP3 binding cell death enhancer-1.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: MANN-WHITNEY, Binding Assay

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Response to the chemotherapeutic drugs with differential DAP3 and DELE1 expression.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Expressing

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Correlation between DAP3 and DELE1 with angiogenesis biomarkers. (A and B) Correlation efficiency between DAP3 and angiogenesis markers in The Cancer Genome Atlas-COAD cohort was presented in (A) heatmap (A) and (B) scatter plots. (C and D) Association between DELE1 and angiogenesis biomarkers was revealed in (C) heatmap and (D) scatter plots. DAP3, death associated protein-3; DELE1, DAP3 binding cell death enhancer-1. ** P<0.01.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Binding Assay

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Validation of the DAP3 and DELE1 knockdown in RKO cells by quantitative PCR gene transcript analysis. Left panel: Expression of DAP3 in the cell models. The levels of expression in wild type, control transfection (control plasmid and control siRNA), DAP3 knockdown, DELE1 knockdown and DAP3/DELE1 dual knockdown cells were compared. Control transfections resulted in minor and statistically insignificant changes of DAP3 when compared with wild type cells (P>0.05) Right panel: Expression of DELE1 in the cell models. The levels of expression in wild type, control transfection (control plasmid and control siRNA), DELE1 knockdown and DAP3/DELE1 dual knockdown cells were compared. Control transfections resulted in minor and statistically insignificant changes of DELE1 when compared with wild type cells (P>0.05). Values presented in the figure are mean and standard deviation. Error bars represent standard deviation. ** P<0.01 vs. RKO wild type cells, ΔΔ P<0.001 vs. RKO-control-plasmid cells and ## P<0.01 VS RKO-control-siRNA cells. DAP3, death associated protein-3; DELE1, DAP3 binding cell death enhancer-1; siRNA, small interfering RNA.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Biomarker Discovery, Knockdown, Real-time Polymerase Chain Reaction, Expressing, Control, Transfection, Plasmid Preparation, Standard Deviation, Binding Assay, Small Interfering RNA

Journal: International Journal of Oncology

Article Title: Death associated protein-3 (DAP3) and DAP3 binding cell death enhancer-1 (DELE1) in human colorectal cancer, and their impacts on clinical outcome and chemoresistance

doi: 10.3892/ijo.2022.5455

Figure Lengend Snippet: Metabolic of colorectal cancer cells following DAP3 and DELE1 knockdown. (A) Concentrations of extracellular glucose in different RKO cells were determined by luminescence measurement. (B) NAD(P)H measurements quantified by luminescence in different RKO cells. (C) Differential extracellular lactate expression and the ratio in different RKO cells. (D) NO 2 Standard curve and measurements of NO 2 in different RKO cells. Error bar stands standard error of means. * P<0.05 and ** P<0.01 by Student's t-test. DAP3, death associated protein-3; DELE1, DAP3 binding cell death enhancer-1.

Article Snippet: Primary antibodies used in the current study included anti-DAP3 antibody (cat. no. sc-373911) and anti-DELE1 antibody (cat. no. sc-515080; both from Santa Cruz Biotechnologies Inc.).

Techniques: Knockdown, Expressing, Binding Assay

Journal: Frontiers in Physiology

Article Title: Identification and characterization of CHCHD1, AURKAIP1, and CRIF1 as new members of the mammalian mitochondrial ribosome

doi: 10.3389/fphys.2013.00183

Figure Lengend Snippet: Peptide sequences of new mitochondrial ribosomal proteins identified from LC-MS/MS analyses of in-gel tryptic digestions of 28S, 39S, and 55S samples prepared at high salt conditions .

Article Snippet: Proteins were transferred to PVDF membranes, which were probed with rabbit polyclonal anti-CHCHD1 antibody at a 1:1000 dilution (Abcam), rabbit anti-AURKAIP1 antibody at a 1:1000 dilution (Sigma-Aldrich), goat anti-CRIF1 antibody at a 1:500 dilution (Santa Cruz Biotechnology), mouse monoclonal anti-MRPS29 (DAP3) and anti-HSP60 antibodies at 1:5000 dilutions (BD Transduction Laboratories), mouse anti-OXPHOS (MITOPROFILE®) at a 1:5000 dilution (Abcam Inc.), or rabbit polyclonal human anti-MRPL47 and mouse polyclonal human anti-MRPS18-2 antibodies at 1:5000 dilutions (produced in-house) for 16 h at 4°C.

Techniques: Sequencing

Journal: Frontiers in Physiology

Article Title: Identification and characterization of CHCHD1, AURKAIP1, and CRIF1 as new members of the mammalian mitochondrial ribosome

doi: 10.3389/fphys.2013.00183

Figure Lengend Snippet: Identification and detection of mitochondrial ribosomal proteins and their subunit association by capLC-MS/MS and immunoblotting analyses. (A) Purified mitochondrial (55S) ribosomes (~10 A 260 ) prepared in high salt conditions were dissociated into subunits by sedimentation through a second 10–30% linear sucrose gradient in the presence of 2 mM Mg 2+ as described in Figure . After the sedimentation of the purified subunits, the same A 260 units of 28S and 39S subunits and 55S ribosomes (~0.5) were separated on SDS-PAGE. The gel was cut into 30 pieces for in-gel digestion of protein bands with trypsin and the peptides were analyzed by capLC-MS/MS analysis to identify the proteins present. Gel fractions containing the peptides detected from newly identified MRPs are marked on the image. Proteins identified in these gel pieces are given in Tables S1, S2. (B) Purified 28S and 39S subunits (~0.2 A 260 ) were obtained from the dissociation of purified 55S ribosomes as described in Materials and Methods at 2 mM Mg 2+ . Proteins present in these preparations were separated on 12% SDS-PAGE and analyzed by immunoblotting using CHCHD1, AURKAIP1, CRIF1, MRPS29, and MRPL47 antibodies to confirm subunit distribution of new MRPs. (C) To confirm the ribosome association of newly identified proteins, crude mitochondrial ribosomes prepared at high salt and detergent conditions were separated on a 10–30% linear sucrose gradient in the presence of 20 mM Mg 2+ as described in Figure . Distribution of new proteins, CHCHD1, AURKAIP1, and CRIF1, were detected by immunoblotting analyses of sucrose gradient fractions containing dissociated ribosomal subunits (28S and 39S) and 55S ribosomes. Antibodies against two 28S subunit proteins, MRPS18B and MRPS29, and a 39S protein, MRPL47, were used to indicate the locations of the subunits and the 55S ribosome.

Article Snippet: Proteins were transferred to PVDF membranes, which were probed with rabbit polyclonal anti-CHCHD1 antibody at a 1:1000 dilution (Abcam), rabbit anti-AURKAIP1 antibody at a 1:1000 dilution (Sigma-Aldrich), goat anti-CRIF1 antibody at a 1:500 dilution (Santa Cruz Biotechnology), mouse monoclonal anti-MRPS29 (DAP3) and anti-HSP60 antibodies at 1:5000 dilutions (BD Transduction Laboratories), mouse anti-OXPHOS (MITOPROFILE®) at a 1:5000 dilution (Abcam Inc.), or rabbit polyclonal human anti-MRPL47 and mouse polyclonal human anti-MRPS18-2 antibodies at 1:5000 dilutions (produced in-house) for 16 h at 4°C.

Techniques: Tandem Mass Spectroscopy, Western Blot, Purification, Sedimentation, SDS Page

Journal: Frontiers in Physiology

Article Title: Identification and characterization of CHCHD1, AURKAIP1, and CRIF1 as new members of the mammalian mitochondrial ribosome

doi: 10.3389/fphys.2013.00183

Figure Lengend Snippet: Relative distribution of new mitochondrial ribosomal proteins using emPAI (Exponentially Modified Protein Abundance Index) values calculated from peptides detected by LC-MS/MS analyses of proteins in 28S and 39S subunits and 55S ribosomes .

Article Snippet: Proteins were transferred to PVDF membranes, which were probed with rabbit polyclonal anti-CHCHD1 antibody at a 1:1000 dilution (Abcam), rabbit anti-AURKAIP1 antibody at a 1:1000 dilution (Sigma-Aldrich), goat anti-CRIF1 antibody at a 1:500 dilution (Santa Cruz Biotechnology), mouse monoclonal anti-MRPS29 (DAP3) and anti-HSP60 antibodies at 1:5000 dilutions (BD Transduction Laboratories), mouse anti-OXPHOS (MITOPROFILE®) at a 1:5000 dilution (Abcam Inc.), or rabbit polyclonal human anti-MRPL47 and mouse polyclonal human anti-MRPS18-2 antibodies at 1:5000 dilutions (produced in-house) for 16 h at 4°C.

Techniques: Modification, Quantitative Proteomics

Journal: Frontiers in Physiology

Article Title: Identification and characterization of CHCHD1, AURKAIP1, and CRIF1 as new members of the mammalian mitochondrial ribosome

doi: 10.3389/fphys.2013.00183

Figure Lengend Snippet: GenBank™ and Swiss-Prot accession numbers of new mitochondrial ribosomal proteins found in various species .

Article Snippet: Proteins were transferred to PVDF membranes, which were probed with rabbit polyclonal anti-CHCHD1 antibody at a 1:1000 dilution (Abcam), rabbit anti-AURKAIP1 antibody at a 1:1000 dilution (Sigma-Aldrich), goat anti-CRIF1 antibody at a 1:500 dilution (Santa Cruz Biotechnology), mouse monoclonal anti-MRPS29 (DAP3) and anti-HSP60 antibodies at 1:5000 dilutions (BD Transduction Laboratories), mouse anti-OXPHOS (MITOPROFILE®) at a 1:5000 dilution (Abcam Inc.), or rabbit polyclonal human anti-MRPL47 and mouse polyclonal human anti-MRPS18-2 antibodies at 1:5000 dilutions (produced in-house) for 16 h at 4°C.

Techniques:

Journal: Frontiers in Physiology

Article Title: Identification and characterization of CHCHD1, AURKAIP1, and CRIF1 as new members of the mammalian mitochondrial ribosome

doi: 10.3389/fphys.2013.00183

Figure Lengend Snippet: Sedimentation of new MRPs with large complexes in human cell lines and mitochondria. (A) Whole cell lysate (WCL) and (B) mitochondrial lysate (MITO) obtained from human cell lines were layered on sucrose cushion preparations. After 16 h centrifugation, post-ribosomal supernatant layers (Layers 1 to 6) and the crude ribosome pellet (P) were collected and analyzed by immunoblotting probed with CHCHD1, AURKAIP1, CRIF1, MRPS29, MRPL47, and HSP60 antibodies. (C) To confirm the RNA-dependent association of new MRPs with mitochondrial ribosomes, the same amount of crude ribosomes, both Control and RNase A-treated, were sedimented on 10–30% sucrose gradients. Equal volumes of sucrose gradient fractions were separated on 12% SDS-PAGE. Sedimentation of new MRPs with ribosomal subunits and 55S ribosomes in the absence and presence of RNase A was analyzed by immunoblotting using CHCHD1, AURKAIP1, CRIF1, MRPS29, and MRPL47 antibodies. HSP60 immunoblotting and the Coomassie Blue stained gel are shown as controls for multimeric mitochondrial complexes sedimenting with the 55S ribosome in control and RNase A-treated gradients.

Article Snippet: Proteins were transferred to PVDF membranes, which were probed with rabbit polyclonal anti-CHCHD1 antibody at a 1:1000 dilution (Abcam), rabbit anti-AURKAIP1 antibody at a 1:1000 dilution (Sigma-Aldrich), goat anti-CRIF1 antibody at a 1:500 dilution (Santa Cruz Biotechnology), mouse monoclonal anti-MRPS29 (DAP3) and anti-HSP60 antibodies at 1:5000 dilutions (BD Transduction Laboratories), mouse anti-OXPHOS (MITOPROFILE®) at a 1:5000 dilution (Abcam Inc.), or rabbit polyclonal human anti-MRPL47 and mouse polyclonal human anti-MRPS18-2 antibodies at 1:5000 dilutions (produced in-house) for 16 h at 4°C.

Techniques: Sedimentation, Centrifugation, Western Blot, Control, SDS Page, Staining

Journal: Frontiers in Physiology

Article Title: Identification and characterization of CHCHD1, AURKAIP1, and CRIF1 as new members of the mammalian mitochondrial ribosome

doi: 10.3389/fphys.2013.00183

Figure Lengend Snippet: Role of CHCHD1, AURKAIP1, and CRIF1 knock-down on mitochondrial protein synthesis. (A) Transcript levels of CHCHD1, AURKAIP1, and CRIF1 were shown as an indication of knock-down efficiency in cell lines transfected with control siRNA and siRNAs for corresponding new MRPs. Mitochondrial encoded ND6, COI, COII, ATP6, and 12S rRNA, and nuclear encoded MRPS29 and MRPL47 were examined by RT-PCR reactions in the same samples. RT-PCR reactions with GAPDH were performed as a positive control and relative quantitation of signal intensities was calculated based on GAPDH mRNA levels (or siRNA control cell line) in each knock-down cell line. (B) Expression levels of new MRPs in control siRNA and MRP-specific siRNA transfected HEK293T cell lysates and total crude ribosome fractions ( * ) were analyzed by immunoblotting using antibodies against AURKAIP1, CHCHD1, CRIF1, MRPS29, MRPL47, and HSP60. Immunoblotting analyses with HSP60, MRPS29, and MRPL47 antibodies were used as loading controls for total protein and total ribosome content using whole cell lysates obtained from control and AURKAIP1 siRNA knock-down cells. Similar results were obtained with immunoblotting analyses of total crude ribosome fractions from CHCHD1 and CRIF1 siRNA knock-down cell lines using HSP60, MRPS29, and MRPL47 antibodies (data not shown). (C) De novo synthesis of mitochondrial proteins was evaluated in control siRNA and the siRNA of corresponding MRPs transfected HEK293T cells by pulse labeling of proteins in the presence of [ 35 S]-methionine and a cytosolic translation inhibitor, emetine. A representative electrophoretic pattern of the de novo synthesized translational products is presented. ND1, −2, −3, −4, −4L, −5, and −6 are subunits of Complex I; Cyt b is a subunit of Complex III; COI, −II, and −III are subunits of Complex IV; ATP6 and ATP8 are subunits of Complex V. Coomassie blue staining of the same gel was performed to ensure equal protein loading in the gel. (D) The combined intensities of the 13 mitochondrially-encoded proteins from each lane were used as an overall quantitation of mitochondrial protein synthesis from three separate experiments. (E) Immunoblotting analysis of whole cell lysates prepared from control siRNA and MRP specific siRNA transfected HEK293T cells was performed to examine the steady state levels of the mitochondrial encoded subunit of Complex IV (COII) and nuclear encoded subunits of Complex V (ATP5A), III (UQCRC2), and II (SDHB). The arrow shows the reduced translation of mitochondrial encoded protein COII in cell lines transfected with CHCHD1, AURKAIP1, and CRIF1 specific siRNAs. The same PVDF was reprobed with MRPS29, MRPL47, and HSP60 antibodies to ensure equal loading.

Article Snippet: Proteins were transferred to PVDF membranes, which were probed with rabbit polyclonal anti-CHCHD1 antibody at a 1:1000 dilution (Abcam), rabbit anti-AURKAIP1 antibody at a 1:1000 dilution (Sigma-Aldrich), goat anti-CRIF1 antibody at a 1:500 dilution (Santa Cruz Biotechnology), mouse monoclonal anti-MRPS29 (DAP3) and anti-HSP60 antibodies at 1:5000 dilutions (BD Transduction Laboratories), mouse anti-OXPHOS (MITOPROFILE®) at a 1:5000 dilution (Abcam Inc.), or rabbit polyclonal human anti-MRPL47 and mouse polyclonal human anti-MRPS18-2 antibodies at 1:5000 dilutions (produced in-house) for 16 h at 4°C.

Techniques: Knockdown, Transfection, Control, Reverse Transcription Polymerase Chain Reaction, Positive Control, Quantitation Assay, Expressing, Western Blot, Labeling, Synthesized, Staining

Journal: Frontiers in Physiology

Article Title: Identification and characterization of CHCHD1, AURKAIP1, and CRIF1 as new members of the mammalian mitochondrial ribosome

doi: 10.3389/fphys.2013.00183

Figure Lengend Snippet: List of mammalian mitochondrial ribosomal proteins with their bacterial homologs .

Article Snippet: Proteins were transferred to PVDF membranes, which were probed with rabbit polyclonal anti-CHCHD1 antibody at a 1:1000 dilution (Abcam), rabbit anti-AURKAIP1 antibody at a 1:1000 dilution (Sigma-Aldrich), goat anti-CRIF1 antibody at a 1:500 dilution (Santa Cruz Biotechnology), mouse monoclonal anti-MRPS29 (DAP3) and anti-HSP60 antibodies at 1:5000 dilutions (BD Transduction Laboratories), mouse anti-OXPHOS (MITOPROFILE®) at a 1:5000 dilution (Abcam Inc.), or rabbit polyclonal human anti-MRPL47 and mouse polyclonal human anti-MRPS18-2 antibodies at 1:5000 dilutions (produced in-house) for 16 h at 4°C.

Techniques: