Journal: Oncology Reports

Article Title: Increased expression of GIPC2 in colon adenocarcinoma is associated with a favorable prognosis and high levels of immune cell infiltration

doi: 10.3892/or.2023.8503

Figure Lengend Snippet: Expression levels of GIPC2 in COAD. (A) Expression levels of GIPC2 (mean ± SD) in 33 types of cancer based on The Cancer Genome Atlas data. (B) Unpaired analysis of GIPC2 expression (mean ± SD) between tumor and normal tissues in 478 patients with COAD. (C) Paired analysis of GIPC2 expression (scatter points represent expression levels of individual samples) between tumor and normal tissues (n=41). (D) Comparison of GIPC2 protein expression levels between normal and tumor tissues using immunohistochemistry (100× magnification). *P<0.05, **P<0.01, ***P<0.001 vs. normal. COAD, colon adenocarcinoma; GIPC2, Gα-interacting protein C-terminus PDZ-domain-containing family member 2; ns, not significant.

Article Snippet: The concentrated rabbit polyclonal antibody against human GIPC2 protein was purchased from BIOSS (cat no. KT22301; 1:200).

Techniques: Expressing, Immunohistochemistry

Journal: Oncology Reports

Article Title: Increased expression of GIPC2 in colon adenocarcinoma is associated with a favorable prognosis and high levels of immune cell infiltration

doi: 10.3892/or.2023.8503

Figure Lengend Snippet: Association between GIPC2 expression levels and clinicopathological variables. Association of GIPC2 with (A) pathological stage, (B) T stage, (C) N stage, (D) M stage, (E) sex, (F) age, (G) lymphatic invasion and (H) perineural invasion. Data are presented as the mean ± SD. *P<0.05, **P<0.01, ***P<0.001. GIPC2, Gα-interacting protein C-terminus PDZ-domain-containing family member 2; ns, not significant.

Article Snippet: The concentrated rabbit polyclonal antibody against human GIPC2 protein was purchased from BIOSS (cat no. KT22301; 1:200).

Techniques: Expressing

Journal: Oncology Reports

Article Title: Increased expression of GIPC2 in colon adenocarcinoma is associated with a favorable prognosis and high levels of immune cell infiltration

doi: 10.3892/or.2023.8503

Figure Lengend Snippet: Association between GIPC2 expression levels and prognosis. Increased GIPC2 expression in COAD was associated with favorable (A) overall survival, (B) disease-specific survival and (C) progression-free interval. (D) Multivariate Cox regression analysis between GIPC2 expression and clinicopathological factors. Visualization of 1, 3 and 5-year survival probabilities and risk coefficients for each variable of the Cox survival model using forest plots. GIPC2, Gα-interacting protein C-terminus PDZ-domain-containing family member 2.

Article Snippet: The concentrated rabbit polyclonal antibody against human GIPC2 protein was purchased from BIOSS (cat no. KT22301; 1:200).

Techniques: Expressing

Journal: Oncology Reports

Article Title: Increased expression of GIPC2 in colon adenocarcinoma is associated with a favorable prognosis and high levels of immune cell infiltration

doi: 10.3892/or.2023.8503

Figure Lengend Snippet: Cox regression analysis for overall survival.

Article Snippet: The concentrated rabbit polyclonal antibody against human GIPC2 protein was purchased from BIOSS (cat no. KT22301; 1:200).

Techniques:

Journal: Oncology Reports

Article Title: Increased expression of GIPC2 in colon adenocarcinoma is associated with a favorable prognosis and high levels of immune cell infiltration

doi: 10.3892/or.2023.8503

Figure Lengend Snippet: Analysis of genes co-expressed with GIPC2 in COAD. Relationship between GIPC2 and the top five co-expressed genes: (A) EPCAM, (B) LRRC8D, (C) EPB41L4B, (D) ACSL5 and (E) CDS1. (F) Top 100 co-expressed genes of GIPC2 were selected to conduct the enrichment analysis. GIPC2, Gα-interacting protein C-terminus PDZ-domain-containing family member 2.

Article Snippet: The concentrated rabbit polyclonal antibody against human GIPC2 protein was purchased from BIOSS (cat no. KT22301; 1:200).

Techniques:

Journal: Oncology Reports

Article Title: Increased expression of GIPC2 in colon adenocarcinoma is associated with a favorable prognosis and high levels of immune cell infiltration

doi: 10.3892/or.2023.8503

Figure Lengend Snippet: Relationship between GIPC2 expression levels and tumor-infiltrating immune cells. (A) Immune cell score heat map; different colors represent the expression trend in different samples according to the grouping of GIPC2 expression levels (high GIPC2 expression vs. low GIPC2 expression group). *P<0.05, **P<0.01, ***P<0.001. The significance between the two groups was determined using the Mann-Whitney U test. (B) Abundance of tumor-infiltrating immune cells in each sample, with different colors referring to different types of immune cells. The abscissa represents the sample, and the ordinate represents the percentage of immune cells in a single sample. GIPC2, Gα-interacting protein C-terminus PDZ-domain-containing family member 2; NK, natural killer.

Article Snippet: The concentrated rabbit polyclonal antibody against human GIPC2 protein was purchased from BIOSS (cat no. KT22301; 1:200).

Techniques: Expressing, MANN-WHITNEY

Journal: Oncology Reports

Article Title: Increased expression of GIPC2 in colon adenocarcinoma is associated with a favorable prognosis and high levels of immune cell infiltration

doi: 10.3892/or.2023.8503

Figure Lengend Snippet: Relationship between GIPC2 expression and the expression levels of immune checkpoint-related genes. Data are presented as the mean ± SD. *P<0.05, **P<0.01, ***P<0.001 vs. high GIPC2 expression. GIPC2, Gα-interacting protein C-terminus PDZ-domain-containing family member 2.

Article Snippet: The concentrated rabbit polyclonal antibody against human GIPC2 protein was purchased from BIOSS (cat no. KT22301; 1:200).

Techniques: Expressing

Journal: Oncogene

Article Title: GIPC2 interacts with Fzd7 to promote prostate cancer metastasis by activating WNT signaling.

doi: 10.1038/s41388-022-02255-4

Figure Lengend Snippet: Fig. 2 GIPC2 promotor methylation regulated GIPC2 expression in prostate cancer. a Schematic representation of the locations of CpG islands. The red bar shows the CpG sites. The corresponding numbers indicate the genomic locations designated in the UCSC database (hg38). b The promoter-methylation ratios of GIPC2 in prostate cancer samples were detected by EpiTYPER methylation analysis. The methylation ratios were significantly higher in normal adjacent tissues than in prostate cancer tumors. c Correlation between methylation of GIPC2-promoter CpG islands and their expression levels in all samples (n = 89). d The promoter-methylation ratios of GIPC2 were determined by MSP in four cell lines treated with or without DAC. Unmethylated (Unmeth) and methylated (Meth) PCR products were detected. After DAC treatment, GIPC2 mRNA- and protein-expression levels were detected using RT-PCR (e) and western blotting (f, g). The error bars represent the mean ± SD. h Effect of DAC expression on the methylation status of the GIPC2 promoter. DNA from control- or DAC-treated RWPE-1 cells was collected at the indicated time points, cloned, and sequenced to detect CpG-island methylation at the GIPC2 promoter. A summary of bisulfite-treated gDNA-sequencing results from RWPE-1 cells treated with DAC for increasing times is shown, where the amplified region contained 14 CpG sites (represented by circles located along the region) were analyzed by DNA sequencing. The black and white circles represent methylated and unmethylated CpG dinucleotides, respectively. Each line represents the DNA sequence of a random clone, for which the black and white circles represent unmethylated and methylated CpG sites in these regions, respectively. i, j RWPE-1 cells were treated with DAC, and GIPC2 protein expression was detected at 0, 24, 48, or 72 h.

Article Snippet: Paraffin sections (8 μm) were stained with antibodies against GIPC2 (sc515441, Santa Cruz) or BMI-1 (ab126783, Abcam) at a 1:100 dilution.

Techniques: Methylation, Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot, Control, Clone Assay, Sequencing, DNA Sequencing

Journal: Oncogene

Article Title: GIPC2 interacts with Fzd7 to promote prostate cancer metastasis by activating WNT signaling.

doi: 10.1038/s41388-022-02255-4

Figure Lengend Snippet: Fig. 3 GIPC2 did not impact prostate cancer proliferation or apoptosis in vitro. a–d EdU staining and (e, f) CCK8 assays were performed to evaluate the effect of GIPC2 on cell proliferation. RWPE-1 and C4-2 cells were transfected with pcDNA3.1-GIPC2 or siRNA of GIPC2 (siGIPC2-1/ siGIPC2-2), and cell proliferation was detected by EdU staining and a cell-viability kit at different time points. Apoptosis was determined using a Cell Death Detection Kit (g, h) and FCM (i, j). GIPC2 overexpression in RWPE-1 cells and GIPC2 knockdown in C4-2 cells. All experiments were performed at least thrice and the data shown represent the mean ± SD. *p < 0.05, **p < 0.01, two-sided paired t test.

Article Snippet: Paraffin sections (8 μm) were stained with antibodies against GIPC2 (sc515441, Santa Cruz) or BMI-1 (ab126783, Abcam) at a 1:100 dilution.

Techniques: In Vitro, Staining, Transfection, Over Expression, Knockdown

Journal: Oncogene

Article Title: GIPC2 interacts with Fzd7 to promote prostate cancer metastasis by activating WNT signaling.

doi: 10.1038/s41388-022-02255-4

Figure Lengend Snippet: Fig. 5 GIPC2 promoted WNT–β-catenin-pathway activation. a, b RWPE-1 and C4-2 cells were transfected with pcDNA-GIPC2 or siGIPC2-1/ siGIPC2-2 for 48 h. Then, the protein-expression levels of GIPC2, p-GSK-3β, total GSK-3β, activated β-catenin, and total β-catenin were analyzed by western blotting, with ACTB serving as a loading control. c, d Graphical display of target protein-expression levels normalized to ACTB expression. e, f Treating RWPE-1 cells with 0.25 μg DKK1 (an inhibitor of the Wnt–β-catenin-signaling pathway), with or without GIPC2 overexpression. Total proteins were harvested at the indicated time points for western blot analysis. g, h C4-2 cells were treated with 5 mM LiCl (an activator of the Wnt–β-catenin-signaling pathway) for 24 h, following GIPC2 downregulation. Protein-expression levels of GIPC2, p-GSK-3β, total GSK-3β, activated β-catenin, and total β-catenin were analyzed by western blotting, with ACTB serving as a loading control. i, j In tumor tissue homogenates from mouse xenografts, protein expression levels of GIPC2, p-GSK-3β, and activated β-catenin were measured to verify the effects of GIPC2 knockdown. Western blot-band intensities were measured using Image J software. Normalization was done by dividing the target signal by the ACTB signal. P values were determined by Student’s t test. The results are presented as the mean ± SD. *p < 0.05, n = 3.

Article Snippet: Paraffin sections (8 μm) were stained with antibodies against GIPC2 (sc515441, Santa Cruz) or BMI-1 (ab126783, Abcam) at a 1:100 dilution.

Techniques: Activation Assay, Transfection, Expressing, Western Blot, Control, Over Expression, Knockdown, Software

Journal: Oncogene

Article Title: GIPC2 interacts with Fzd7 to promote prostate cancer metastasis by activating WNT signaling.

doi: 10.1038/s41388-022-02255-4

Figure Lengend Snippet: Fig. 6 GIPC2 interacted with the PDZ domain of Fzd7. a, b Exogenous GIPC2 associated with exogenous Fzd7. Total lysates from C4-2 cells expressing GIPC2-HA and/or Fzd7-Myc were immunoprecipitated with antibodies against Myc (a) or the HA tag (b), followed by western blotting using the indicated antibodies. c Endogenous GIPC2 interacted with endogenous Fzd7. C4-2 cell lysates were immunoprecipitated with an anti-Fzd7 antibody, followed by western blotting using an anti-GIPC2 antibody. d C4-2 cells were treated with or without DAC. Endogenous GIPC2 from C4-2 cells was analyzed by western blotting with an anti-Fzd7 antibody or immunoprecipitated with an anti-GIPC2 antibody. e Positive FRET images. C4-2 cells, co-expressing pmCherry-Fzd7 and three different GFP–GIPC2 domains, were analyzed by confocal microscopy. Positive FRET image of C4-2 cells co-transfected with pairs of GFP-C1 and Red-N1 fusion crystallins. f Negative FRET images. Images were acquired before and after photobleaching. A nonbleached region and a corresponding bleached region (shown by an arrow) were used for the data analysis, compared with an unbleached cellular region as a negative control. g C4-2 cells co-transfected with pAcGFP- GIPC2-PDZ (green) and pmCherry-Fzd7 (red). h C4-2 cells co-transfected with pAcGFP-GIPC2-GH1 and pmCherry-Fzd7. i The images show C4-2 cells co-transfected with pAcGFP-GIPC2-GH2 and pmCherry-Fzd7. j FRET efficiencies (percentages) for the interaction between two crystallin partners were calculated, and the FRET intensity was analyzed statistically. The FRET intensity expressed as a percent represents the fraction of interacting donor molecules. The results suggested that a significant interaction occurred between GIPC2-PDZ and Red-Fzd7. k Domain architecture of GIPC2. The GIPC2 protein consists of a GH1 domain (red), a PDZ domain (blue), and a GH2 domain (green). The amino acid position is shown on the domain architecture. The GH1 domain in the N-terminal region, the PDZ domain in the middle region, and the GH2 domain in the C-terminal region are well conserved among GIPC family members.

Article Snippet: Paraffin sections (8 μm) were stained with antibodies against GIPC2 (sc515441, Santa Cruz) or BMI-1 (ab126783, Abcam) at a 1:100 dilution.

Techniques: Expressing, Immunoprecipitation, Western Blot, Confocal Microscopy, Transfection, Negative Control

Journal: Oncogene

Article Title: GIPC2 interacts with Fzd7 to promote prostate cancer metastasis by activating WNT signaling.

doi: 10.1038/s41388-022-02255-4

Figure Lengend Snippet: Fig. 7 The GIPC2–PDZ–Fzd7 axis activated the WNT–β-catenin pathway in prostate cancer metastasis. a Luciferase-expressing C4-2 cells (2 × 105) were suspended in 15 μL of sterile PBS and used for intracardiac injections of male BALB/c nu/nu mice with different GIPC2 isoforms (wild-type GIPC2, GIPC2-δGH1, GIPC2-δGH2, and GIPC2-δPDZ, n = 8 of each group). Bioluminescent images of metastatic tumors were monitored. Each picture includes four different time points: 7, 14, 21, and 28 days. The colored scale bars represent low (purple) to high (red) tumor burdens. b The metastasis rate was determined using the Kaplan–Meier method. *p < 0.05 between GIPC2-δPDZ group and three other groups. c Western blotting showed GIPC2, Fzd7, p-GSK-3β, and activated β-catenin expression in C4-2 cells transfected with OE-GIPC2 (GIPC2 overexpression) or si-Fzd7. d Graphical display of target protein-expression levels normalized to ACTB expression. e Cell-migration abilities were measured by performing wound-healing assays. C4-2 cells were transfected with pcDNA-GIPC2 (with or without si-Fzd7) to evaluate the pro-migration and pro-invasion effects of the GIPC2–Fzd7 axis. After scratching a wound and removing the floating cells, prostate cancer cells (untreated or control-treated) were used in wound-scratch assays for 0, 1, or 3 days. f Prostate cancer cells treated as described above were applied to transwell chambers coated with Matrigel and incubated for 24 h. g The microfluidic model consists of two independent microchannels, where tumor cells and 20% FBS were seeded. Between the two channels, 3D Matrigel was seeded to mimic the ECM. Invading cells was observed under a light microscope (200×). The data shown represent the mean ± SD. *p < 0.05; **p < 0.01.

Article Snippet: Paraffin sections (8 μm) were stained with antibodies against GIPC2 (sc515441, Santa Cruz) or BMI-1 (ab126783, Abcam) at a 1:100 dilution.

Techniques: Luciferase, Expressing, Sterility, Western Blot, Transfection, Over Expression, Migration, Control, Incubation, Light Microscopy

Journal: Translational Cancer Research

Article Title: Bioinformatics analysis reveals the clinical significance of GIPC2/GPD1L for colorectal cancer using TCGA database

doi: 10.21037/tcr-21-1933

Figure Lengend Snippet: Relationship between GIPC2 expression and clinical characteristics

Article Snippet: GIPC2 protein expression images in normal tissues were determined from Human Protein Atlas ( www.proteinatlas.org/ENSG00000137960-GIPC2/tissue/colon#img ).

Techniques: Expressing

Journal: Translational Cancer Research

Article Title: Bioinformatics analysis reveals the clinical significance of GIPC2/GPD1L for colorectal cancer using TCGA database

doi: 10.21037/tcr-21-1933

Figure Lengend Snippet: GIPC2 is downregulated in human CRC tissues and cells. (A) Box-plot shows differences in GIPC2 levels between tumor and normal tissues in TCGA CRC database. (B) GIPC2 expression levels were identified in human normal colon epithelial cells (HcoEpiC) and human colorectal cancer cell lines (Lovo, RKO, DLD-1, and HCT116) using qRT-PCR (***, P<0.001). (C) GIPC2 expression levels in colorectal cancer tissues and surrounding tissues were shown by immunohistochemistry (IHC) from the human protein atlas ( www.proteinatlas.org ). CRC, colorectal cancer; qRT-PCR, quantitative real-time polymerase chain reaction;

Article Snippet: GIPC2 protein expression images in normal tissues were determined from Human Protein Atlas ( www.proteinatlas.org/ENSG00000137960-GIPC2/tissue/colon#img ).

Techniques: Expressing, Quantitative RT-PCR, Immunohistochemistry, Real-time Polymerase Chain Reaction

Journal: Translational Cancer Research

Article Title: Bioinformatics analysis reveals the clinical significance of GIPC2/GPD1L for colorectal cancer using TCGA database

doi: 10.21037/tcr-21-1933

Figure Lengend Snippet: GIPC2 participates in CRC progression. (A) Boxplot depicts the expression levels of GIPC2 among different tumor degrees of invasion (T stage, T1&T2, T3&T4) of CRC patients in TCGA database (**, P<0.01). (B) Box-plot shows the expression patterns of GIPC2 according to tumor lymph node metastasis (N stage, N0, N1&N2) in CRC patients in TCGA database (***, P<0.001). (C) Box-plot shows the expression levels of GIPC2 with different clinicopathologic stages according to CRC AJCC classification (stage I & stage II, stage III & stage IV) in TCGA database (***, P<0.001). (D) Boxplot indicates the expression patterns of GIPC2 according to M stage (M0, M1) in TCGA CRC database (ns). CRC, colorectal cancer.

Article Snippet: GIPC2 protein expression images in normal tissues were determined from Human Protein Atlas ( www.proteinatlas.org/ENSG00000137960-GIPC2/tissue/colon#img ).

Techniques: Expressing

Journal: Translational Cancer Research

Article Title: Bioinformatics analysis reveals the clinical significance of GIPC2/GPD1L for colorectal cancer using TCGA database

doi: 10.21037/tcr-21-1933

Figure Lengend Snippet: GPD1L expression was significantly correlated with GIPC2 expression and multiple clinical characteristics of CRC patients. (A) Correlation between GPD1L and GIPC2 in CRC array from TCGA CRC sequence-based gene expression (P=0). (B) Box-plot shows differences in GPD1L levels between tumor and normal tissues in TCGA CRC database. (C) Box-plot shows distribution patterns of GPD1L according to tumor lymph node metastasis (N stage, N0, N1&N2) in CRC patients in TCGA database (**, P<0.01). (D) Boxplot demonstrates the expression level variation of GIPC2 with different clinical-pathological stages according to CRC AJCC classification (stage I & stage II, stage III & stage IV) in TCGA database (**, P<0.01). (E) Boxplot depicts the expression level variation of GIPC2 among different tumor degrees of invasion (T stage, T1&T2, T3&T4) in CRC patients in TCGA database (ns). (F) Boxplot indicates distribution patterns of GIPC2 according to M stage (M0, M1) in TCGA CRC database (ns). CRC, colorectal cancer; ns, not significant.

Article Snippet: GIPC2 protein expression images in normal tissues were determined from Human Protein Atlas ( www.proteinatlas.org/ENSG00000137960-GIPC2/tissue/colon#img ).

Techniques: Expressing, Sequencing, Gene Expression

Journal: Translational Cancer Research

Article Title: Bioinformatics analysis reveals the clinical significance of GIPC2/GPD1L for colorectal cancer using TCGA database

doi: 10.21037/tcr-21-1933

Figure Lengend Snippet: Functional exploration for GIPC2 and GPD1L in CRC. (A) Heatmap displays expression patterns of 698 GIPC2-related genes. (B) Heatmap demonstrates expression patterns of 869 GIPC2-related genes. (C) Venn diagram intersects GIPC2- and GIPC2-related genes. (D) GO annotations of 426 genes correlated with GIPC2 and GPD1L. (E) KEGG enrichment analysis of 426 genes associated with GIPC2 and GPD1L. CRC, colorectal cancer.

Article Snippet: GIPC2 protein expression images in normal tissues were determined from Human Protein Atlas ( www.proteinatlas.org/ENSG00000137960-GIPC2/tissue/colon#img ).

Techniques: Functional Assay, Expressing

Journal: Translational Cancer Research

Article Title: Bioinformatics analysis reveals the clinical significance of GIPC2/GPD1L for colorectal cancer using TCGA database

doi: 10.21037/tcr-21-1933

Figure Lengend Snippet: Evaluation of the diagnostic and prognosis ability of GIPC2 and GPD1L expression using CRC sample from TCGA. (A) ROC curve analysis of GIPC2 expression for diagnosing CRC patients. (B) ROC curve analysis of GPD1L expression for diagnosing CRC patients. (C,D) Kaplan-Meier curve shows OS of CRC patients between low- and high-expression of GIPC2 and GPD1L. (E,F) Kaplan-Meier curve shows PFI of CRC patients between low- and high-expression of GIPC2 and GPD1L.

Article Snippet: GIPC2 protein expression images in normal tissues were determined from Human Protein Atlas ( www.proteinatlas.org/ENSG00000137960-GIPC2/tissue/colon#img ).

Techniques: Diagnostic Assay, Expressing

Journal: Cell Death & Disease

Article Title: GIPC2 is an endocrine-specific tumor suppressor gene for both sporadic and hereditary tumors of RET- and SDHB-, but not VHL-associated clusters of pheochromocytoma/paraganglioma

doi: 10.1038/s41419-021-03731-7

Figure Lengend Snippet: A The copy number variation of GIPC2 in 55 PPGL tumors was verified by qPCR. The internal control gene was the human C2 gene. Copy number relative ratio <1.4 was determined as copy number deletion, while copy number relative ratio between 1.4 and 2.6 was considered as normal copy number. Tumor with germline or somatic RET mutation was indicated with an asterisk. B The mRNA level of GIPC2 was measured in primary PPGL ( n = 54) and normal medulla tissue ( n = 10) by RT-PCR. C GIPC2 mRNA levels were analyzed in copy number deletion tumors ( n = 39) and normal copy number tumors ( n = 15). D Correlation analysis between GIPC2 copy number and its expression in PPGL samples ( n = 54). E Immunohistochemistry of GIPC2 in the normal medulla and in PPGLs with or without GIPC2 deletion. Scale bars represent 100 µm. Two representative samples were selected for each group. F Western blot of representative tumor samples using antibodies against GIPC2 and ACTB. G Methylation levels of GIPC2 promoter were quantified by Sequenome EpiTYPER analysis in normal ( n = 6) and PPGL samples ( n = 53). H Correlation analysis between GIPC2 methylation and its expression in PPGL samples ( n = 59, including 6 normal samples). I Methylation-specific PCR assay for GIPC2 of DNA isolated from PC12 cells treated or untreated with 10 μM 5-AZA. PCR products labeled with “Meth” or “Unmeth” were generated by primers specific for methylated or unmethylated GIPC2. Relative gene expression of GIPC2 was measured by qPCR, and rat adrenal medulla was examined as well for comparison. J GIPC2 relative expression levels in PPGLs under different genetic subgroups. Data from E-MTAB-733 of the GEO database ( http://www.ebi.ac.uk/arrayexpress/ ) on Affymetrix Human U133 plus 2.0 array, normalized by the geometric mean of ACTB and GAPDH expression levels.

Article Snippet: The primary antibodies against pERK1/2, ERK1/2, pMEK, MEK, p27, ACTB, p-pRB, p53, HA-Tag were purchased from Cell Signaling Technology (USA), antibodies against NONO, HIF1A, HIF2A, VHL were purchased from Abcam (UK), antibodies against p18, pRB were purchased from Santa Cruz (USA), GIPC2, and RET antibodies were purchased from LSBio (USA) and Abbiotec (USA) respectively.

Techniques: Control, Mutagenesis, Reverse Transcription Polymerase Chain Reaction, Expressing, Immunohistochemistry, Western Blot, Methylation, Isolation, Labeling, Generated, Gene Expression, Comparison

Journal: Cell Death & Disease

Article Title: GIPC2 is an endocrine-specific tumor suppressor gene for both sporadic and hereditary tumors of RET- and SDHB-, but not VHL-associated clusters of pheochromocytoma/paraganglioma

doi: 10.1038/s41419-021-03731-7

Figure Lengend Snippet: A Subcellular localization of GIPC2 protein in vivo. The distribution of endogenous GIPC2 was detected by immunofluorescent microscopy with anti-GIPC2 antibody in ACC cells (a) and hPheo1 cells (b). DAPI staining was included to visualize the cell nucleus. B PC12 cells and hPheo1 cells were infected with Tet-on lentiviruses systems carrying GIPC2 or vector, or ACC cells were transfected with the control or si-GIPC2. The growth curves of the cells in 96-well plates were measured with CCK-8 assay at an indicated hour. The efficiency of GIPC2 expression was verified by western blot (insets). C Stable cell lines of PC12-GIPC2 or control PC12 were maintained for 14 days in colony formation assay. D EdU imaging was performed and quantified in ACC cells which were transfected with the control or si-GIPC2. DAPI staining was included to visualize the cell nucleus. E , F PC12 cells infected with Tet-on lentiviruses carrying GIPC2 were inoculated into 4-week-old female BALB/c nude mice (3 × 10 6 cells). When the subcutaneous tumors reached 80–100 mm 3 , mice were randomized into 2 groups and treated with saline as a control group or Doxycycline (20 mg/kg daily) to induce GIPC2 overexpression by intraperitoneal injection. The final tumor sizes were measured ( E ). The final tumor weights were determined and the tumor growth curve was plotted according to the tumor volume ( F ). Each bar represented the mean ± S.D. n = 8. G Protein lysates were prepared from PC12, 293T, and HEK293 cells with GIPC2 overexpression or knockdown and analyzed by western blot using antibodies against the indicated proteins. H Protein lysates were prepared from GIPC2-knockdown HEK293 cells with or without ERK inhibitor PD98059 treatment and analyzed by western blot using antibodies against the indicated proteins.

Article Snippet: The primary antibodies against pERK1/2, ERK1/2, pMEK, MEK, p27, ACTB, p-pRB, p53, HA-Tag were purchased from Cell Signaling Technology (USA), antibodies against NONO, HIF1A, HIF2A, VHL were purchased from Abcam (UK), antibodies against p18, pRB were purchased from Santa Cruz (USA), GIPC2, and RET antibodies were purchased from LSBio (USA) and Abbiotec (USA) respectively.

Techniques: In Vivo, Microscopy, Staining, Infection, Plasmid Preparation, Transfection, Control, CCK-8 Assay, Expressing, Western Blot, Stable Transfection, Colony Assay, Imaging, Saline, Over Expression, Injection, Knockdown

Journal: Cell Death & Disease

Article Title: GIPC2 is an endocrine-specific tumor suppressor gene for both sporadic and hereditary tumors of RET- and SDHB-, but not VHL-associated clusters of pheochromocytoma/paraganglioma

doi: 10.1038/s41419-021-03731-7

Figure Lengend Snippet: A PC12 cells infected with Tet-on lentiviruses systems carrying GIPC2 was treated with Doxycycline for 72 h to induce GIPC2 overexpression prior to cell cycle analysis with ACCURI C6 flow cytometry using CFLOWPLUS and MODFIT software, respectively. B The p27 relative expression signal in PPGL under different genetic subgroups. Data from GEO gene expression database (E-MTAB-733) ( http://www.ebi.ac.uk/arrayexpress/ ) . C Total cellular proteins were prepared from PC12, 293T, HEK293, and hPheo1 cells with GIPC2 overexpression or knockdown, and p27 protein expression was analyzed by western blot. D PC12 cells were treated with cycloheximide (CHX) in the presence or absence of overexpressed GIPC2. p27 protein levels were analyzed at 0, 6, 12, and 24 h time points after the addition of CHX by western blot and quantified. E Kaplan–Meier survival analysis of TCGA-PCPG data set for the disease-free survival time of the high 2 signatures ( GIPC2 and p27 ) group and the low 2 signatures group using the online tool GEPIA2 (gepia2.cancer-pku.cn/). The high 2 signatures group contains the samples with the top 30% of the two-gene ( GIPC2 and p27) signature expression values generated by GEPIA2 ( N = 55), while the low 2 signatures group contains the samples with the bottom 25% of the two-gene signature expression values ( N = 46). Disease-free survival time refers to the time until the occurrence of relapse, distant metastases, or positive regional lymph nodes.

Article Snippet: The primary antibodies against pERK1/2, ERK1/2, pMEK, MEK, p27, ACTB, p-pRB, p53, HA-Tag were purchased from Cell Signaling Technology (USA), antibodies against NONO, HIF1A, HIF2A, VHL were purchased from Abcam (UK), antibodies against p18, pRB were purchased from Santa Cruz (USA), GIPC2, and RET antibodies were purchased from LSBio (USA) and Abbiotec (USA) respectively.

Techniques: Infection, Over Expression, Cell Cycle Assay, Flow Cytometry, Software, Expressing, Gene Expression, Knockdown, Western Blot, Generated

Journal: Cell Death & Disease

Article Title: GIPC2 is an endocrine-specific tumor suppressor gene for both sporadic and hereditary tumors of RET- and SDHB-, but not VHL-associated clusters of pheochromocytoma/paraganglioma

doi: 10.1038/s41419-021-03731-7

Figure Lengend Snippet: A Immunoprecipitation-mass spectrometry analysis of GIPC2-associated proteins. Cellular extracts from 293T cells overexpressing GIPC2-HA, GIPC2-ΔPDZ-HA, or control 293T cells were immunoprecipitated with an anti-HA antibody. The eluates were resolved by SDS-PAGE and stained with coomassie brilliant blue. The four different protein bands (arrows) were retrieved and analyzed by mass spectrometry. B Interaction of GIPC2 with the candidate proteins. Top: Fluorescence resonance energy transfer (FRET) experiments were performed in PC12 cells co-transfected with GIPC2-AcGFP and mcherry-indicated proteins. Percentage E refers to the percentage of energy transfer efficiency. Bottom: Co-localization of GIPC2 and NONO protein in vitro. PC12 cells were co-transfected with GIPC2-AcGFP and mcherry-NONO and observed under a confocal microscope. Green represented GIPC2 and red represented NONO. C Subcellular localization of NONO protein in vivo. The distribution of endogenous NONO was detected by immunofluorescent microscopy with antibodies against NONO in PC12 cells. DAPI staining was included to visualize the cell nucleus. D Interaction of GIPC2 with NONO. Whole-cell lysates from 293T cells co-transfected with GIPC2-HA and His-NONO or control plasmids were prepared and immunoprecipitation was performed with Ni-NTA agarose and anti-GIPC2, respectively, followed by immunoblotting with antibodies against indicated proteins. (The WB diagrams of the “Lysate: anti-His” and “Lysate: anti-GIPC2” are identical in the left and right panel, as they are the same experiment of lysate, without IP, probed with the indicated antibody) E Whole-cell lysates from hPheo1 cells were immunoprecipitated with antibodies against GIPC2 or IgG followed by immunoblotting with the antibodies against the indicated proteins. F GST pull-down experiment. 293T lysates were incubated with bacterially expressed GST fused with either GIPC2 or GIPC2-ΔPDZ. Western blot analysis of the GST-fused proteins and interacting NONO protein were shown. G hPheo1 cells were transfected with the si-Control or si-NONO. The growth curves of the cells in 96-well plates were measured with CCK-8 assay. The efficiency of si-NONO knockdown in hPheo1 cells was verified by western blot. H Total cellular proteins were extracted from hPheo1 cells with NONO overexpression or knockdown and western blot was performed with indicated antibodies. I hPheo1 cells with stably transfected Tet-on lentiviruses systems carrying GIPC2 were transfected with si-NONO or si-Control for 24 h, together with or without tetracycline (3 μg/mL) for another 48 h. Western blot analysis was performed, to measure the relative level of p27 protein normalized by ACTB . J hPheo1 cells with stably transfected Tet-on lentiviruses systems carrying GIPC2 were transfected with si-NONO or si-Control, together with the indicated p27-luciferase reporter, and after 6 h treated with tetracycline (3 μg/mL) to induce GIPC2 overexpression. 48 h later, luciferase activity was measured. Relative luciferase activity was calculated as firefly luciferase activity divided by renilla luciferase activity and shown relative to the control (transfected with pGL-3-Basic vector). K Verification of the ChIP-qPCR results in hPheo1 cells. hPheo1 cells were transfected with GIPC2-HA (a) or NONO-HA (b). ChIP-qPCR analysis of the selected p27 promoters was performed using antibodies against HA-tag or IgG. Results were represented as fold enrichment over control IgG with GAPDH as a negative control. The overexpression of GIPC2 or NONO was verified by western blot. Each point and bars of the pictures above represented the mean ± S.D. for triplicate experiments.

Article Snippet: The primary antibodies against pERK1/2, ERK1/2, pMEK, MEK, p27, ACTB, p-pRB, p53, HA-Tag were purchased from Cell Signaling Technology (USA), antibodies against NONO, HIF1A, HIF2A, VHL were purchased from Abcam (UK), antibodies against p18, pRB were purchased from Santa Cruz (USA), GIPC2, and RET antibodies were purchased from LSBio (USA) and Abbiotec (USA) respectively.

Techniques: Immunoprecipitation, Mass Spectrometry, Control, SDS Page, Staining, Fluorescence, Förster Resonance Energy Transfer, Transfection, In Vitro, Microscopy, In Vivo, Western Blot, Incubation, CCK-8 Assay, Knockdown, Over Expression, Stable Transfection, Luciferase, Activity Assay, Plasmid Preparation, ChIP-qPCR, Negative Control

Journal: Cell Death & Disease

Article Title: GIPC2 is an endocrine-specific tumor suppressor gene for both sporadic and hereditary tumors of RET- and SDHB-, but not VHL-associated clusters of pheochromocytoma/paraganglioma

doi: 10.1038/s41419-021-03731-7

Figure Lengend Snippet: A GIPC2 and p27 expressions were significantly reduced in hereditary NF1-, RET-, SDH -related tumors but not in hereditary VHL -related tumors in the data set of 188 PPGLs from the gene expression database (E-MTAB-733). B ACC cells in the presence of 10 μM Dexamethasone (Dex) in the media were transfected with RET634 (p.C634R) or RET918 (p.M918T) mutant plasmid or plasmid with additional GIPC2 knockdown. Cell proliferation was measured with CCK-8 assay at 48 h. C ACC cells were transfected with RET WT or RET634 mutant plasmid in the presence or absence of Dex. Apoptosis was assayed using Cell Death Detection ELISA Kit 48 h later. D , E ACC was treated with 100 ng/mL NGF or 10 μM Dex for 7 days. D Immunofluorescence of GIPC2 in ACC after 7 days. Green represents the nucleus, red represents GIPC2. E shows the protein changes and the apoptosis of ACC when treated with 100 ng/mL NGF in the absence or presence of 0.1, 1, and 10 μM Dex. F Western blot was performed in ACC cells in the presence of Dex with RET wild type or RET634/918 mutation overexpression, using antibodies against the indicated proteins. G GIPC2 protein levels with an increasing amount of RET634 mutant plasmid used during transfection of ACC cells in the presence of Dex were measured by western blot. H Western blot was performed to measure the protein levels of p27 and pErK1/2 under cotransfection of RET634 mutant and an increasing amount of GIPC2 plasmid in ACC cells in the presence of Dex. Proliferation was measured at 48 h.

Article Snippet: The primary antibodies against pERK1/2, ERK1/2, pMEK, MEK, p27, ACTB, p-pRB, p53, HA-Tag were purchased from Cell Signaling Technology (USA), antibodies against NONO, HIF1A, HIF2A, VHL were purchased from Abcam (UK), antibodies against p18, pRB were purchased from Santa Cruz (USA), GIPC2, and RET antibodies were purchased from LSBio (USA) and Abbiotec (USA) respectively.

Techniques: Gene Expression, Transfection, Mutagenesis, Plasmid Preparation, Knockdown, CCK-8 Assay, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Western Blot, Over Expression, Cotransfection

Journal: Cell Death & Disease

Article Title: GIPC2 is an endocrine-specific tumor suppressor gene for both sporadic and hereditary tumors of RET- and SDHB-, but not VHL-associated clusters of pheochromocytoma/paraganglioma

doi: 10.1038/s41419-021-03731-7

Figure Lengend Snippet: A The proliferation of ACC cells overexpressing SDHB-mutant (p. R46Q) or si-SDHD was measured at 48 h. B Western blot was performed in ACC cells with SDHB-mutant overexpression or si-SDHD using antibodies against the indicated proteins. C ACC proliferation under 10 mM DMS and/or 10uM Dex treatment. D GIPC2 protein level with increasing DMS concentrations in ACC cells was measured by western blot. E Western blot was performed to measure the protein levels of p27 and pErK1/2 under 10 mM DMS stimulation and with an increasing amount of GIPC2 plasmid used during transfection of ACC cells. Proliferation was measured at 48 h. F GIPC2 methylation ratio as measured by EpiTyper assay in ACC transfected with RET and SDHB mutants. G Methylation-specific PCR assay was carried out to detect the methylation of GIPC2 promoter in ACC cells treated with 10 mM DMS and 0.1, 1, and 10 μM Dex.

Article Snippet: The primary antibodies against pERK1/2, ERK1/2, pMEK, MEK, p27, ACTB, p-pRB, p53, HA-Tag were purchased from Cell Signaling Technology (USA), antibodies against NONO, HIF1A, HIF2A, VHL were purchased from Abcam (UK), antibodies against p18, pRB were purchased from Santa Cruz (USA), GIPC2, and RET antibodies were purchased from LSBio (USA) and Abbiotec (USA) respectively.

Techniques: Mutagenesis, Western Blot, Over Expression, Plasmid Preparation, Transfection, Methylation, EpiTYPER Assay

Journal: Cell Death & Disease

Article Title: GIPC2 is an endocrine-specific tumor suppressor gene for both sporadic and hereditary tumors of RET- and SDHB-, but not VHL-associated clusters of pheochromocytoma/paraganglioma

doi: 10.1038/s41419-021-03731-7

Figure Lengend Snippet: A Cell proliferation assay was performed in PC12 (a), HEK293, and HT-29 cells (b) with the overexpression of human VHL-type 2A (p.Y98H), Type 2B (p.W117R), Type 2C (p.L188V), and Type I (p.C162F) mutants and VHL-WT. B Under normal or hypoxia conditions, ACC was first transfected with a siRNA against rat VHL (siVHL) for 48 h, followed by transfection of VHL-WT or VHL-Type 2C mutant and was grown under low serum (1%) condition . After 48 h western blot was performed using antibodies against the indicated proteins. C ACC was first transfected with si-VHL for 48 h, followed by transfection of VHL-WT or VHL mutants as indicated, and was grown under low serum (1%) condition. After 48 h Western blot was performed using antibodies against the indicated proteins (top panel). The cell apoptosis was determined using Cell Death Detection ELISA Kit (bottom panel). D PC12 cells and a PC12 line with stable overexpression of GIPC2 (PC12 + GIPC2) were treated as in C , together with a p53-luciferase reporter transfection. After 48 h, the cells were harvested and the luciferase activities were measured by the Dual-Luciferase Reporter Assay System (Promega, USA).

Article Snippet: The primary antibodies against pERK1/2, ERK1/2, pMEK, MEK, p27, ACTB, p-pRB, p53, HA-Tag were purchased from Cell Signaling Technology (USA), antibodies against NONO, HIF1A, HIF2A, VHL were purchased from Abcam (UK), antibodies against p18, pRB were purchased from Santa Cruz (USA), GIPC2, and RET antibodies were purchased from LSBio (USA) and Abbiotec (USA) respectively.

Techniques: Proliferation Assay, Over Expression, Transfection, Mutagenesis, Western Blot, Enzyme-linked Immunosorbent Assay, Luciferase, Reporter Assay

Journal: eLife

Article Title: GIPC proteins negatively modulate Plexind1 signaling during vascular development

doi: 10.7554/eLife.30454

Figure Lengend Snippet: ( A ) Confocal lateral images of the trunk vasculature (green) of 32 hpf embryos (region dorsal to the yolk extension). Anterior, left; dorsal, up. Scale bars (white horizontal lines), 100 μm. Genotypes indicated on top of each image in yellow font. Angiogenesis deficits are indicated as follows: white asterisks (DLAV gaps), magenta asterisks (truncated Se), white greater-than sign (thin Se). Maternal-zygotic (MZ) removal of gipc activity is denoted by the designation ‘MZ’ in superscript. In the WT image (top left), the vessels are designated with the white font as follows: DLAV (Dorsal Longitudinal Anastomotic Vessel), Se (Segmental Vessel), DA (Dorsal Aorta), and PCV (Posterior Cardinal Vein). ( B ) Bar graph. Percentage of Se in 32 hpf embryos of the indicated genotypes belonging to each of the following four phenotypic classes. Truncated: severe (includes missing Se), medium (yellow), and weak (gray). Non-truncated: complete (black). Significance values were calculated using a two-sided Fisher Exact test and significant differences (p<0.0033) assigned using a Bonferroni-type adjustment for 15 pairwise genotype comparisons (0.05/15 = 0.0033). Brackets and asterisks indicate pairs of genotypes with significantly different distributions of these four phenotypic classes. Quantifications. We scored Se angiogenesis in embryos of the following six genotypes: WT (138 Se, 12 embryos; an average of 11.5 Se/embryo), gipc1 skt1 (130 Se, 11 embryos; an average of 11.8 Se/embryo) , gipc1 skt1(MZ) (380 Se, 33 embryos; an average of 11.5 Se/embryo) , gipc2 skt3/skt4 (130 Se, 11 embryos; an average of 11.8 Se/embryo) , gipc1 skt1 ; gipc2 skt3/skt4 (152 Se, 13 embryos; an average of 11.6 Se/embryo), and gipc1 skt1(MZ) ; gipc2 skt3/skt4 (220 Se, 19 embryos; an average of 11.5 Se/embryo). For additional data, graphs, and statistical comparisons related to this figure, see , and . Please note that given the use of different scales for scoring angiogenesis deficits, it is unfeasible to compare the quantifications in and directly.

Article Snippet: Recombinant DNA reagent , GIPC2 shRNA (h) Lentiviral Particles , Santa Cruz Biotechnology , Cat. # sc-75132-V , shRNA pool (three target-specific constructs against human GIPC2 that encode 19–25 nt (plus hairpin) shRNAs). Target sequences (sense sequences 5’ to 3’): (1) CAGACGAAUUUGUCUUUGA (2) GGACACCUUUACUAACUCU (3) CCAACUUUCUCUCUUUGUA.

Techniques: Activity Assay

Journal: eLife

Article Title: GIPC proteins negatively modulate Plexind1 signaling during vascular development

doi: 10.7554/eLife.30454

Figure Lengend Snippet: ( A–D ) Confocal lateral images of the trunk vasculature (green) of 32 hpf embryos (region dorsal to the yolk extension). Anterior, left; dorsal, up. Scale bars (white horizontal lines), 100 μm. Morpholino injection (un-injected or injected with plxnd1 morpholino) indicated on top, genotypes (WT or gipc1 skt1(MZ) ; gipc2 skt4(MZ) ) indicated on the left. The un-injected WT picture ( A ) shows the names of the major vessels in white font: DLAV (Dorsal Longitudinal Anastomotic Vessel), Se (Segmental Vessel), DA (Dorsal Aorta), and PCV (Posterior Cardinal Vein). Vascular defects highlighted as follows: truncated or missing Se (magenta asterisk), thin Se (white greater/less-than signs), DLAV gaps (white asterisk). Quantifications. The following number of embryos were analyzed: WT (four embryos), WT injected with plxnd1 morpholino (four embryos; 4/4 showed a vascular phenotype similar to that of plxnd1 fov01b nulls), gipc1 skt1(MZ) ; gipc2 skt4(MZ) (12 embryos; 7/12 showed angiogenesis deficits), and gipc1 skt1(MZ) ; gipc2 skt4(MZ) injected with plxnd1 morpholino (11 embryos; 11/11 showed a vascular phenotype similar to that of plxnd1 fov01b nulls).

Article Snippet: Recombinant DNA reagent , GIPC2 shRNA (h) Lentiviral Particles , Santa Cruz Biotechnology , Cat. # sc-75132-V , shRNA pool (three target-specific constructs against human GIPC2 that encode 19–25 nt (plus hairpin) shRNAs). Target sequences (sense sequences 5’ to 3’): (1) CAGACGAAUUUGUCUUUGA (2) GGACACCUUUACUAACUCU (3) CCAACUUUCUCUCUUUGUA.

Techniques: Injection

Journal: eLife

Article Title: GIPC proteins negatively modulate Plexind1 signaling during vascular development

doi: 10.7554/eLife.30454

Figure Lengend Snippet: ( A–F ). Representative fluorescent images of HUVEC morphology in cell collapse experiments under the following conditions. No ligand ( A–C ; top) or 45 min stimulation with 10 nM of SEMA3E ( D–F ; bottom). In each picture, the square area marked by yellow dotted sides is shown at twice the magnification at the bottom left corner and delimited by white sides. shRNA treatments as follows. Non-targeting, control ( A, D ), GIPC ( GIPC1, GIPC2, and GIPC3 ; B, E ), and PLXND1 ( C, F ). Scale bars (white horizontal lines), 100 μm. ( A–C ) Without ligand stimulation cells are uncollapsed regardless of the knockdown condition. ( D, E ) Cell collapse under ligand stimulation. Cells treated with non-targeting, control shRNA collapse ( D ). GIPC knockdown cells hypercollapse ( E ). SEMA3E-induced collapse is PLXND1-dependent. PLXND1 knockdown abrogates the morphological response ( F ). Cell collapse data collected from three independent experiments. This figure is related to .

Article Snippet: Recombinant DNA reagent , GIPC2 shRNA (h) Lentiviral Particles , Santa Cruz Biotechnology , Cat. # sc-75132-V , shRNA pool (three target-specific constructs against human GIPC2 that encode 19–25 nt (plus hairpin) shRNAs). Target sequences (sense sequences 5’ to 3’): (1) CAGACGAAUUUGUCUUUGA (2) GGACACCUUUACUAACUCU (3) CCAACUUUCUCUCUUUGUA.

Techniques: shRNA, Control, Knockdown

Journal: eLife

Article Title: GIPC proteins negatively modulate Plexind1 signaling during vascular development

doi: 10.7554/eLife.30454

Figure Lengend Snippet: Western blots for GIPC1, GIPC2, PLXND1, and GAPDH (loading control) from TCLs of cells infected with the indicated shRNA lentiviral particles. Note the effective decrease of GIPC1-2 and PLXND1 levels. GIPC3 expression was absent under all the experimental conditions assayed. Hence, for brevity, the corresponding Western blots are not shown. This figure is related to .

Article Snippet: Recombinant DNA reagent , GIPC2 shRNA (h) Lentiviral Particles , Santa Cruz Biotechnology , Cat. # sc-75132-V , shRNA pool (three target-specific constructs against human GIPC2 that encode 19–25 nt (plus hairpin) shRNAs). Target sequences (sense sequences 5’ to 3’): (1) CAGACGAAUUUGUCUUUGA (2) GGACACCUUUACUAACUCU (3) CCAACUUUCUCUCUUUGUA.

Techniques: Western Blot, Control, Infection, shRNA, Expressing

Journal: eLife

Article Title: GIPC proteins negatively modulate Plexind1 signaling during vascular development

doi: 10.7554/eLife.30454

Figure Lengend Snippet:

Article Snippet: Recombinant DNA reagent , GIPC2 shRNA (h) Lentiviral Particles , Santa Cruz Biotechnology , Cat. # sc-75132-V , shRNA pool (three target-specific constructs against human GIPC2 that encode 19–25 nt (plus hairpin) shRNAs). Target sequences (sense sequences 5’ to 3’): (1) CAGACGAAUUUGUCUUUGA (2) GGACACCUUUACUAACUCU (3) CCAACUUUCUCUCUUUGUA.

Techniques: Transgenic Assay, Plasmid Preparation, Mutagenesis, Derivative Assay, Selection, Stable Transfection, Knock-Out, Recombinant, Control, shRNA, Sequencing, Construct, Synthesized, Positive Control, Sterility, Concentration Assay

Journal: eLife

Article Title: GIPC proteins negatively modulate Plexind1 signaling during vascular development

doi: 10.7554/eLife.30454

Figure Lengend Snippet:

Article Snippet: Recombinant DNA reagent , GIPC2 shRNA (h) Lentiviral Particles , Santa Cruz Biotechnology , Cat. # sc-75132-V , shRNA pool (three target-specific constructs against human GIPC2 that encode 19–25 nt (plus hairpin) shRNAs). Target sequences (sense sequences 5’ to 3’): (1) CAGACGAAUUUGUCUUUGA (2) GGACACCUUUACUAACUCU (3) CCAACUUUCUCUCUUUGUA.

Techniques: Sequencing