Journal: bioRxiv

Article Title: Signaling by Ras G12V depends on EGFR activity in vivo

doi: 10.1101/2025.11.12.687982

Figure Lengend Snippet: (A) Schematic representation of the Ras1 genomic region showing the extent of the Ras1 KO deletion (“replacer region”; up) and the molecular structure of Ras1 KO-Kin constructs. From top to bottom: Ras1 WT , Flag-Ras1 WT , Flag-Ras1 G12V , Flag-KRASB WT , Flag-KRASB G12V , FRT Ras1 G12V (referred as Ras1 G12V ) and FRT Ras1 wt FRT Flag-Ras1 G12V (referred as FRT Ras1 ). (B) Representative western blot from third instar larval extracts of the following genotypes hemizygous for Ras1 : Df(3R)by10/+ (control), Flag-Ras1 wt /Df(3R)by10 , Flag-Ras1 G12V /Df(3R)by10 , Ras1 wt /Df(3R)by10 , Ras1 G12V /Df(3R)by10 and FRT Ras1/Df(3R)by10 . Note the differential migration for tagged (Flag-Ras; up) versus untagged (Ras; down) proteins. βTub was used as an internal loading control. (C) Quantification of Ras protein levels relative to βTub corresponding to three biological replicates of the western blots shown in B. Horizontal black lines indicate the mean of biological replicates. Each vertical blue line indicates the standard deviation (SD) of technical replicates, and the circles represents the mean of each biological replicate. (D) Percentage of pupal lethality in control and Ras1 KO-Kin heterozygous backgrounds. The bar graphs display the mean ± SD. (E) Survival curves of heterozygous females for control, Ras1 WT /+ (blue line) , Ras1 G12V /+ (dotted blue line) , Flag-Ras1 WT (pink line) , Flag-Ras1 G12V /+ (dotted pink line) , Flag-KRASB WT /+ (purple line) , and Flag-KRASB G12V /+ (dotted purple line) heterozygous females. Survival statistic were calculated using Gehan-Breslow-Wilcoxon test. (F) Adult female wing phenotypes of heterozygous Ras1 WT and KRASB WT (left), heterozygous Ras1 G12V and KRASB G12V (“Endogenous Ras G12V expression”), sal EPv -Gal4 UAS-GFP/UAS-Ras1 G12V (sal>UAS-Ras1 G12V ) and sal EPv -Gal4 UAS-GFP/UAS-KRASB G12V (sal>UAS-KRASB G12V ; “Ectopic Ras G12V expression”), and functional rescue of Ras1 knockdown ( sal EPv -Gal4 UAS-GFP/UAS-Ras1-RNAi ; sal>UAS-Ras1-i) by endogenous KRASB WT expression in sal EPv -Gal4 UAS-GFP/UAS-Ras1-RNAi; KRASB WT /+ flies (sal>UAS-Ras1-i; KRASB WT /+; “KRASB functional equivalence”).

Article Snippet: Briefly, Drosophila Ras1 genomic DNA from vasa-Cas9 flies, Ras1 cDNA (RE53955, DGCR) and human KRASB cDNA (SC109374, OriGene Technologies) were cloned into reintegration vectors ( RIV white and RIV FRTMCSFRTMCS3 ) ( Baena-Lopez et al , 2013 ) in either wild-type or G12V mutant forms and tagged or not with the Flag or HA epitopes at their 5’ end.

Techniques: Construct, Western Blot, Control, Migration, Standard Deviation, Expressing, Functional Assay, Knockdown

Journal: bioRxiv

Article Title: Signaling by Ras G12V depends on EGFR activity in vivo

doi: 10.1101/2025.11.12.687982

Figure Lengend Snippet: (A) Representative western blots of Raf-RAS binding domain (RBD) pulldowns detecting Ras-GTP from third instar larval extracts of the following genotypes: controls, Ras1 WT /+, Ras1 G12V /+, Flag-Ras1 WT /+, Flag-Ras1 G12V /+, Flag-KRASB WT /+, Flag-KRASB G12V /+ and FRT Ras1/+ (left) and control, Flag-Ras1 WT /+, Flag-Ras1 G12V /+ and Flag-Ras1 G12V-CAAX /+ (right). Each line shows the presence of Flag-Ras and Ras in the pulldown (above) and input (below). (B) Wing of Flag-Ras1 G12V-CAAX /+ genotype showing normal wing size and pattern of veins. (C) Quantification of Ras-GTP (% Ras-GTP) corresponding to the western blots shown in A. The graph displays the mean ± SD of different biological replicates (dots). (D) Representative western blot from third instar larval extracts from hemizygous control ( Df(3R)by10/+ ), Ras1 WT /Df(3R)by10, Ras1 G12V /Df(3R)by10, Flag-Ras1 WT /Df(3R)by10, Flag-Ras1 G12V /Df(3R)by10, Flag-KRASB WT /Df(3R)by10 and Flag-KRASB G12V /Df(3R)by10 . Blots were probed for dpERK, total ERK and βTub (loading control). (E) Quantification of dpERK levels relative to βTub corresponding to the western blots shown in D. No significant differences were observed between genotypes. Horizontal black lines indicate the mean of biological replicates. Each vertical blue line indicates the standard deviation (SD) of technical replicates, and the circles represents the mean of each biological replicate. (F) Schematic representation of a mature third instar larval wing disc indicating the pattern of dpERK accumulation by shades of red. In the wing pouch region, maximal accumulation of dpERK is detected in the developing wing veins (L2-L5), along two stripes abutting the dorso-ventral boundary (D/V) and in the precursor cells of the sensory organs (SOPs) in the dorsal notum region. (G) Immunostaining for dpERK in third instar larval wing imaginal discs of the following genotypes: Ras1 WT /+ , Ras1 G12V /+ , KRASB WT /+ and KRASB G12V /+ (top row from left to right) and Ras1 WT /Df(3R)by10 , Ras1 G12V /Df(3R)by10 , KRASB WT /Df(3R)by10 and KRASB G12V /Df(3R)by10 (bottom row, left to right). Scale bar: 100 μm. (H) Quantification of the ratio of dpERK levels between the vein L4 and the L4-L5 intervein territories in Ras1 WT /+ and Ras1 G12V /+ dorsal and ventral wing disc compartments. Violin plots display the median and the Q1 and Q3 interquartile ranges.

Article Snippet: Briefly, Drosophila Ras1 genomic DNA from vasa-Cas9 flies, Ras1 cDNA (RE53955, DGCR) and human KRASB cDNA (SC109374, OriGene Technologies) were cloned into reintegration vectors ( RIV white and RIV FRTMCSFRTMCS3 ) ( Baena-Lopez et al , 2013 ) in either wild-type or G12V mutant forms and tagged or not with the Flag or HA epitopes at their 5’ end.

Techniques: Western Blot, Binding Assay, Control, Standard Deviation, Immunostaining

Journal: bioRxiv

Article Title: Signaling by Ras G12V depends on EGFR activity in vivo

doi: 10.1101/2025.11.12.687982

Figure Lengend Snippet: (A) Accumulation of dpERK in third instar wing imaginal discs, from younger (A 1 ) to older (A 4 ). (B) Schematic representation of a mature wing disc of ap-Gal4 UAS-GFP/UAS-FLP; FRT Ras1 WT FRT Ras1 G12V /+ genotype showing the generation of dorsal Ras1 G12V /+ (GFP positive) and ventral Ras1 WT /+ (GFP negative) territories. (C) Expression of GFP (green in C 1 -C 3 ) and dpERK (red in C 1 -C 3 and white in C 1 ’-C 3 ’) in progressively older third instar wing discs of ap-Gal4 UAS-GFP/UAS-FLP; FRT Ras1 WT FRT Ras1 G12V /+ genotype. (D) Quantification of dpERK levels in dorsal (Ras1 G12V /+) relative to ventral (Ras1 WT /+) compartments in the L4-L5 intervein region of early and late third instar wing disc. Violin plots display the median and the Q1 and Q3 interquartile ranges. (E) Schematic representation of a mature wing disc of ap-Gal4 UAS-GFP/UAS-FLP; FRT HA-Ras1 WT FRT Flag-KRASB G12V /+ genotype showing dorsal KRASB G12V /+ (GFP positive) and ventral Ras1 WT /+ (GFP negative) territories. (F) Expression of GFP (green in F 1 -F 3 ) and dpERK (red in F 1 -F 3 and white in F 1 ’-F 3 ’) in progressively older third instar wing discs of ap-Gal4 UAS-GFP/UAS-FLP; FRT HA-Ras1 WT FRT Flag-KRASB G12V /+ genotype. (G) Quantification of dpERK levels in dorsal (KRASB G12V /+) relative to ventral (Ras1 WT /+) compartments in the L4-L5 intervein region of early and late third instar wing disc. Violin plots display the median and the Q1 and Q3 interquartile ranges. Scale bar: 100 μm.

Article Snippet: Briefly, Drosophila Ras1 genomic DNA from vasa-Cas9 flies, Ras1 cDNA (RE53955, DGCR) and human KRASB cDNA (SC109374, OriGene Technologies) were cloned into reintegration vectors ( RIV white and RIV FRTMCSFRTMCS3 ) ( Baena-Lopez et al , 2013 ) in either wild-type or G12V mutant forms and tagged or not with the Flag or HA epitopes at their 5’ end.

Techniques: Expressing

Journal: Cancer Cell International

Article Title: Novel function of GTPBP2 in promoting hepatocellular carcinoma progression through inhibition of BTRC-mediated KRAS degradation

doi: 10.1186/s12935-025-04029-w

Figure Lengend Snippet: GTPBP2 promotes KRAS and its associated signaling cascade in HCC. (A) Gene Set Enrichment Analysis (GSEA) was performed to investigate the co-expression network and calculate Pearson’s correlation coefficient between GTPBP2 and various hallmarks of cancer in the Roessler HCC cohort ( GSE14520 ). The lower panel presents an enrichment plot depicting genes that are correlated with GTPBP2 within the KRAS signaling pathway. (B) Knockdown of GTPBP2 in HA22T, Mahlavu, and J7 cells was performed to investigate KRAS expression and KRAS-mediated signaling, including phosphorylation of AKT (p-AKT) and MEK (p-MEK), using western blot analysis. Quantitative results are presented in bar plots on the right panel of the immunoblots. (C) The impact of GTPBP2 overexpression on KRAS and its associated signaling cascade was assessed by western blotting and quantitatively analyzed in HA22T, Mahlavu, and J7 cells. β-Actin served as the loading control. Quantitative results are displayed below each signal. Data are presented as means ± SD from three independent experiments. (D) Migration assays were performed to evaluate the effect of KRAS on GTPBP2-overexpressing Mahlavu and J7 cells, using the pcDNA3 empty plasmid as the transfection control. The quantitative results are presented in the right panel (**p < 0.01)

Article Snippet: Equal amounts of protein (50–150 μg based on different targets) were separated via 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), transferred to PVDF membrane (pH 7.9, Amersham Biosciences Inc., Piscataway, NJ, USA) and incubated with the relevant primary antibodies including anti-GTPBP2 (GTX122509, GeneTex, Irvine, CA, USA), ABclonal KRAS Rabbit pAb ABclonal Inc., USA (A1190), anti-p-AKT (sc-7985, Santa Cruz, Santa Cruz Biotechnology, Inc., Europe), anti-AKT (ab179463, Abcam, CA, USA), anti-p-MEK (#9154, Cell Signaling Technology, Boston, MA, USA), anti-MEK (#4694, Cell Signaling Technology), anti-β-Actin (MAB1501R, Millipore, Billerica, Boston, MA, USA), and anti-BTRC (sc-390629, Santa Cruz).

Techniques: Expressing, Knockdown, Phospho-proteomics, Western Blot, Over Expression, Control, Migration, Plasmid Preparation, Transfection

Journal: Cancer Cell International

Article Title: Novel function of GTPBP2 in promoting hepatocellular carcinoma progression through inhibition of BTRC-mediated KRAS degradation

doi: 10.1186/s12935-025-04029-w

Figure Lengend Snippet: Knockdown of GTPBP2 decreases KRAS protein stability in HCC cells. (A ) J7 cells, both with GTPBP2 knockdown and control, were treated with 20 μM cycloheximide (CHX) for time points ranging from 0 to 12 hours. The collected cell lysates were subsequently analyzed by western blot to evaluate KRAS protein stability, with β-actin serving as the loading control. (B) Control or GTPBP2-knockdown J7 cells were treated with or without MG-132 (10 μM) for 4 hours to evaluate the impact of GTPBP2 expression on KRAS protein stability mediated by proteasomal degradation. (C) GTPBP2-knockdown J7 cells transfected with the HA-Ub plasmid were treated with MG-132 for 4 hours prior to KRAS immunoprecipitation and subsequent immunoblotting against Ubiquitin. (D) The RBD pulldown assay was utilized to determine the effect of GTPBP2 overexpression on KRAS activation and to investigate the protein-protein interaction between GTPBP2 and active KRAS in J7 cells. (E) Protein-protein interactions between KRAS and GTPBP2 were detected using the Duolink PLA technique. The close proximity of the two proteins (less than 40 nm apart) is visualized by red fluorescent dots. Nuclei are counterstained with DAPI. Scale bars: 20 μm. (F) Immunohistochemistry (IHC) was performed to detect the expressions of GTPBP2 and KRAS in human HCC tissues. (G) Immunofluorescence (IF) was used to examine cellular location of GTPBP2 and BTRC in J7 cells using antibodies specifically against GTPBP2 and BTRC. Scale bars: 20 μm. (H) Analysis on the amino acid sequence of GTPBP2 for the presence of a BTRC recognition site, DSGKS motif. Two key serine (Ser) residues to binding capacity were mutated to alanine (Ala) to create GTPBP2 mutant (Mt). (I) Co-immunoprecipitation (Co-IP) experiments were conducted using J7 cells transfected with wild-type (Wt) or mutant GTPBP2 containing mutated BTRC to assess potential protein-protein interaction between GTPBP2 and BTRC

Article Snippet: Equal amounts of protein (50–150 μg based on different targets) were separated via 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), transferred to PVDF membrane (pH 7.9, Amersham Biosciences Inc., Piscataway, NJ, USA) and incubated with the relevant primary antibodies including anti-GTPBP2 (GTX122509, GeneTex, Irvine, CA, USA), ABclonal KRAS Rabbit pAb ABclonal Inc., USA (A1190), anti-p-AKT (sc-7985, Santa Cruz, Santa Cruz Biotechnology, Inc., Europe), anti-AKT (ab179463, Abcam, CA, USA), anti-p-MEK (#9154, Cell Signaling Technology, Boston, MA, USA), anti-MEK (#4694, Cell Signaling Technology), anti-β-Actin (MAB1501R, Millipore, Billerica, Boston, MA, USA), and anti-BTRC (sc-390629, Santa Cruz).

Techniques: Knockdown, Control, Western Blot, Expressing, Transfection, Plasmid Preparation, Immunoprecipitation, Ubiquitin Proteomics, Over Expression, Activation Assay, Protein-Protein interactions, Immunohistochemistry, Immunofluorescence, Sequencing, Binding Assay, Mutagenesis, Co-Immunoprecipitation Assay

Journal: Cancer Cell International

Article Title: Novel function of GTPBP2 in promoting hepatocellular carcinoma progression through inhibition of BTRC-mediated KRAS degradation

doi: 10.1186/s12935-025-04029-w

Figure Lengend Snippet: GTPBP2 inhibits KRAS ubiquitination by attenuating the interaction between BTRC and KRAS. (A) BTRC immunoprecipitation analysis was performed to evaluate the effect of GTPBP2 overexpression on the interaction between BTRC and KRAS in J7 and HA22T cells. (B) The interaction between KRAS and BTRC was assessed using Duolink PLA in control, GTPBP2-Wt, and GTPBP2-Mt overexpressing J7 cells. KRAS antibody alone was used as a negative control. The interaction between KRAS and BTRC was detected using both KRAS and BTRC antibodies. Close proximity (<40 nm) between the two proteins is represented by small, distinct red fluorescent puncta observed by fluorescence microscopy. Nuclei were counterstained with DAPI. Scale bars: 20 μm. (C) Quantification of PLA signals was determined by counting the number of puncta per cell. Histograms show the mean± SEM from 30 cells across three independent experiments (*p < 0.05; **p< 0.01). ( D ) Ubiquitination of KRAS was analyzed in HA-Ub plasmid transfected control or BTRC-knockdown J7 cells by immunoprecipitation with the KRAS-specific antibody, followed by immunoblotting with an HA-Ub antibody. ( E ) GTPBP2 protein levels were analyzed by western blotting in control and BTRC-knockdown HA22T and J7 cells treated with or without MG-132 for 4 hours. β-actin served as the loading control. (F) Ubiquitination of GTPBP2 was assessed using the same procedure as in (D), with immunoprecipitation performed using a GTPBP2-specific antibody. The asterisk (*) indicates the antibody heavy chain

Article Snippet: Equal amounts of protein (50–150 μg based on different targets) were separated via 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), transferred to PVDF membrane (pH 7.9, Amersham Biosciences Inc., Piscataway, NJ, USA) and incubated with the relevant primary antibodies including anti-GTPBP2 (GTX122509, GeneTex, Irvine, CA, USA), ABclonal KRAS Rabbit pAb ABclonal Inc., USA (A1190), anti-p-AKT (sc-7985, Santa Cruz, Santa Cruz Biotechnology, Inc., Europe), anti-AKT (ab179463, Abcam, CA, USA), anti-p-MEK (#9154, Cell Signaling Technology, Boston, MA, USA), anti-MEK (#4694, Cell Signaling Technology), anti-β-Actin (MAB1501R, Millipore, Billerica, Boston, MA, USA), and anti-BTRC (sc-390629, Santa Cruz).

Techniques: Ubiquitin Proteomics, Immunoprecipitation, Over Expression, Control, Negative Control, Fluorescence, Microscopy, Plasmid Preparation, Transfection, Knockdown, Western Blot

Journal: Cancer Cell International

Article Title: Novel function of GTPBP2 in promoting hepatocellular carcinoma progression through inhibition of BTRC-mediated KRAS degradation

doi: 10.1186/s12935-025-04029-w

Figure Lengend Snippet: GTPBP2 expression is correlated to AFP content and is required for sorafenib resistance. (A, C) Representative images from the colony formation assay in control and GTPBP2-knocked down Mahlavu and J7 cells, with or without sorafenib (5 μM) treatment, stained with crystal violet. The right panels display the quantified results. (B, D) MTS assays were performed to assess the impact of GTPBP2 on cellular sensitivity to sorafenib in control and GTPBP2-knockdown Mahlavu and J7 cells over a 3-day period. ( E, F ) Cell viability assays in Mahlavu (E) and J7 (F) cells showing that overexpression of GTPBP2 increased resistance to sorafenib, and this effect was overcome by KRAS knockdown. The corresponding IC50 values are indicated. Data are presented as mean ± SEM from three independent experiments (**p<0.05, ** p < 0.01). Experiments were performed in triplicates. GTPBP2 expression was analyzed using the Roessier Liver array ( G) and Cohort 2 (H) in relation to AFP contents. Serum concentration higher than 300 ng/ml is presented as high, less than or equal to 300 ng/ml is presented as low. (* p< 0.05; ** p < 0.01; *** p < 0.001). (I) A schematic illustration of the proposed model where GTPBP2 attenuates BTRC-mediated KRAS degradation in liver cancer. Abnormal expression of GTPBP2 in liver cancer tissue competes with KRAS for BTRC binding, leading to enhanced KRAS protein stability and activation of its downstream signaling pathways. This, in turn, promotes increased metastasis, exacerbates malignancy, and contributes to sorafenib resistance

Article Snippet: Equal amounts of protein (50–150 μg based on different targets) were separated via 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), transferred to PVDF membrane (pH 7.9, Amersham Biosciences Inc., Piscataway, NJ, USA) and incubated with the relevant primary antibodies including anti-GTPBP2 (GTX122509, GeneTex, Irvine, CA, USA), ABclonal KRAS Rabbit pAb ABclonal Inc., USA (A1190), anti-p-AKT (sc-7985, Santa Cruz, Santa Cruz Biotechnology, Inc., Europe), anti-AKT (ab179463, Abcam, CA, USA), anti-p-MEK (#9154, Cell Signaling Technology, Boston, MA, USA), anti-MEK (#4694, Cell Signaling Technology), anti-β-Actin (MAB1501R, Millipore, Billerica, Boston, MA, USA), and anti-BTRC (sc-390629, Santa Cruz).

Techniques: Expressing, Colony Assay, Control, Staining, Knockdown, Over Expression, Concentration Assay, Binding Assay, Activation Assay, Protein-Protein interactions

Journal: Experimental & molecular medicine

Article Title: Oncogenic KRAS mutation confers chemoresistance by upregulating SIRT1 in non-small cell lung cancer.

doi: 10.1038/s12276-023-01091-0

Figure Lengend Snippet: Fig. 2 SIRT1 upregulation is mediated by c-Myc downstream of KRAS. A HEK293T cells were transfected with pcDNA and KRASG12C plasmids (2 μg). H460 cells were transfected with siCon and siKRAS (80 nM). The cells were harvested with lysis buffer and subjected to western blotting. B, C KRASMut cells (H358, NCIH23, SKLU-1, SW900, A427, H727), KRASWT cells, and KRASG12C cells (H1299G12C) were transfected with siCon, c-Myc specific siRNA (80 nM), pcDNA, or c-Myc plasmid (2 μg) for 48 h, and the levels of the KRAS downstream effectors c-Myc and SIRT1 were measured. D H358 cells were transfected with KRASG12C and siCon or sic-Myc, and cell extracts were immunoprecipitated with an anti-KRAS antibody and immunoblotted with anti-SIRT1, anti-c-Myc, and anti-KRAS antibodies. E Chromatin immunoprecipitation-qPCR analysis of KRAS, SIRT1, and SIRT2 was performed in H358 cells transfected with siCon or siKRAS (80 nM) for 48 h and then immunoprecipitated using an anti-c- Myc antibody or mouse IgG as a negative control. The relative enrichment was calculated by normalizing the qPCR signals. The data are plotted as the mean values determined from at least two independent chromatin immunoprecipitation assays and three independent amplification reactions. Student’s t test, mean ± SD; n = 6; *p < 0.05. F H358 cells were transfected with siCon and siKRAS (80 nM) for 48 h and then fixed after 4 h. c-Myc expression was detected with an RFP emission filter, and SIRT1 expression was detected with a GFP emission filter.

Article Snippet: Small interfering RNAs (siRNAs), including siKRAS (sc35731 and sc-43874), siSIRT1 (sc-40986), siSIRT2 (sc-40988), siHDAC6 (sc35544), sip300 (sc-29431), sic-Myc (sc-29226), siADAM17 (sc-36604), and shSIRT1 (sc-40986), and antibodies, including antibodies specific for SIRT1 (sc-15454), SIRT2 (sc-20966), KRAS (F234, sc-30), Tubulin (sc-32293), PARP (sc-8007), cleaved PARP (sc-56196), and p300 (sc-32244), were purchased from Santa Cruz Biotechnology Inc. (Dallas, TX, USA).

Techniques: Transfection, Lysis, Western Blot, Plasmid Preparation, Immunoprecipitation, Chromatin Immunoprecipitation, Negative Control, Expressing

Journal: Experimental & molecular medicine

Article Title: Oncogenic KRAS mutation confers chemoresistance by upregulating SIRT1 in non-small cell lung cancer.

doi: 10.1038/s12276-023-01091-0

Figure Lengend Snippet: Fig. 3 KRASMut-induced SIRT1 rebinds to KRASMut and increases KRAS activity via deacetylation. A HEK293T cells were transfected with KRASG12C and SIRT1 plasmids (4 μg), and cell extracts were immunoprecipitated with anti-KRAS and anti-SIRT1 antibodies and immunoblotted with the reciprocal antibody. B, C Plasmids (pcDNA, KRASG12C, and SIRT1 each 4 μg) and siRNAs (siCon and siSIRT1, each 80 nM) were transfected into HEK293T cells. Cell extracts were immunoprecipitated with an anti-KRAS antibody and Raf-1 agarose beads and analyzed using anti- acetylated lysine, anti-SIRT1, anti-KRAS, and anti-KRAS-GTP antibodies. D Normal lung epithelial cell, fibroblast, and KRAS Mut cell lysates were immunoprecipitated with an anti-KRAS antibody and immunoblotted with anti-acetyl-lysine and anti-KRAS antibodies.

Article Snippet: Small interfering RNAs (siRNAs), including siKRAS (sc35731 and sc-43874), siSIRT1 (sc-40986), siSIRT2 (sc-40988), siHDAC6 (sc35544), sip300 (sc-29431), sic-Myc (sc-29226), siADAM17 (sc-36604), and shSIRT1 (sc-40986), and antibodies, including antibodies specific for SIRT1 (sc-15454), SIRT2 (sc-20966), KRAS (F234, sc-30), Tubulin (sc-32293), PARP (sc-8007), cleaved PARP (sc-56196), and p300 (sc-32244), were purchased from Santa Cruz Biotechnology Inc. (Dallas, TX, USA).

Techniques: Activity Assay, Transfection, Immunoprecipitation

Journal: bioRxiv

Article Title: Transposon mutagenesis identifies cooperating genetic drivers during keratinocyte transformation and cutaneous squamous cell carcinoma progression

doi: 10.1101/2019.12.24.887968

Figure Lengend Snippet: ( a - b ) Assessment of keratinocyte transformation in response to expression of oncogenic KRAS G12D by anchorage-independent soft agar assay (n=3). ( a ) Images of colonies stained with crystal violet solution. ( b ) Statistical significance was tested by unpaired t-test. * P = 0.015.

Article Snippet: KRAS G12D expression vector (Origene cat# RC400104) was a kind gift from Dr Karen Mann.

Techniques: Transformation Assay, Expressing, Soft Agar Assay, Staining