scholarly journals Allele-specific endogenous tagging and quantitative analysis of beta-catenin in colorectal cancer cells

2020 ◽  
Author(s):  
Giulia Ambrosi ◽  
Oksana Voloshanenko ◽  
Antonia F. Eckert ◽  
Dominique Kranz ◽  
G. Ulrich Nienhaus ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Wnt Signaling ◽  
Genome Engineering ◽  
Fluorescent Proteins ◽  
Cancer Cell Line ◽  
Correlation Spectroscopy ◽  
Colorectal Cancer Cell Line ◽  
Beta Catenin ◽  
Wild Type ◽  
The One

ABSTRACTWnt signaling plays important roles in development, homeostasis, and tumorigenesis. Mutations in β-catenin that activate Wnt signaling have been found in colorectal and hepatocellular carcinomas. However, the dynamics of wild-type and mutant forms of β-catenin are not fully understood. Here, we genome-engineered fluorescently tagged alleles of the endogenous β-catenin in a colorectal cancer cell line. Wild-type and oncogenic mutant alleles were tagged with different fluorescent proteins, enabling the analysis of both variants in the same cell. We analyzed the properties of both β-catenin alleles using immunoprecipitation, immunofluorescence and fluorescence correlation spectroscopy approaches, revealing distinctly different biophysical properties. In addition, activation of Wnt signaling by treatment with a GSK3β inhibitor or a truncating APC mutation modulated the wild-type allele to mimic the properties of the mutant β-catenin allele. The one-step tagging strategy demonstrates how genome engineering can be employed for the parallel functional analysis of different genetic variants.

scholarly journals Allele-specific endogenous tagging and quantitative analysis of β-catenin in colorectal cancer cells

eLife ◽  
2022 ◽  
Vol 11 ◽  
Author(s):  
Giulia Ambrosi ◽  
Oksana Voloshanenko ◽  
Antonia F Eckert ◽  
Dominique Kranz ◽  
G Ulrich Nienhaus ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Wnt Signaling ◽  
Genome Engineering ◽  
Fluorescent Proteins ◽  
Cancer Cell Line ◽  
Correlation Spectroscopy ◽  
Colorectal Cancer Cell Line ◽  
Wild Type ◽  
The One ◽  
Mutant Forms

Wnt signaling plays important roles in development, homeostasis, and tumorigenesis. Mutations in β-catenin that activate Wnt signaling have been found in colorectal and hepatocellular carcinomas. However, the dynamics of wild-type and mutant forms of β-catenin are not fully understood. Here, we genome-engineered fluorescently tagged alleles of endogenous β-catenin in a colorectal cancer cell line. Wild-type and oncogenic mutant alleles were tagged with different fluorescent proteins, enabling the analysis of both variants in the same cell. We analyzed the properties of both β-catenin alleles using immunoprecipitation, immunofluorescence, and fluorescence correlation spectroscopy approaches, revealing distinctly different biophysical properties. In addition, activation of Wnt signaling by treatment with a GSK3β inhibitor or a truncating APC mutation modulated the wild-type allele to mimic the properties of the mutant β-catenin allele. The one-step tagging strategy demonstrates how genome engineering can be employed for the parallel functional analysis of different genetic variants.

Silencing the wild-type and mutant K-ras increases the resistance to 5-flurouracil in HCT-116 as a colorectal cancer cell line

2015 ◽  
Vol 26 (2) ◽  
pp. 187-196 ◽  
Author(s):  
Ladan Teimoori-Toolabi ◽  
Saba Hashemi ◽  
Kayhan Azadmanesh ◽  
Farnaz Eghbalpour ◽  
Farnaz Safavifar ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Cell Line ◽  
Cancer Cell ◽  
Cancer Cell Line ◽  
Colorectal Cancer Cell ◽  
Colorectal Cancer Cell Line ◽  
Wild Type ◽  
Hct 116

Cytotoxic Evaluation and Molecular Docking studies of Aminopyridine derivatives as Potential Anticancer Agents

2021 ◽  
Vol 22 ◽  
Author(s):  
Umair Ilyas ◽  
Shagufta Naaz ◽  
Syed Aun Muhammad ◽  
Humaira Nadeem ◽  
Reem Altaf ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Cancer Cell ◽  
In Silico ◽  
Cancer Cell Line ◽  
Docking Studies ◽  
Colorectal Cancer Cell ◽  
Colorectal Cancer Cell Line ◽  
Beta Catenin ◽  
Hct 116

Background: The development of resistance to available anticancer drugs is increasingly becoming a major challenge and new chemical entities could be unveiled to compensate for this therapeutic failure. Objectives: The current study demonstrated whether N-protected and deprotected amino acid derivatives of 2-aminopyridine could attenuate tumor development using colorectal cancer cell lines. Methods: Biological assays were performed to investigate the anticancer potential of synthesized compounds. The in silico ADME profiling and docking studies were also performed by docking the designed compounds against the active binding site of beta-catenin (CTNNB1) to analyze the binding mode of these compounds. Four derivatives 4a, 4b, 4c, and 4d were selected for investigation of in vitro anticancer potential using colorectal cancer cell line HCT 116. The anti-tumor activities of synthesized compounds were further validated by evaluating the inhibitory effects of these compounds on the target protein beta-catenin through in vitro enzyme inhibitory assay. Results: The docking analysis revealed favorable binding energies and interactions with the target proteins. The in vitro MTT assay on colorectal cancer cell line HCT 116 and HT29 revealed potential anti-tumor activities with an IC50 range of 3.7-8.1µM and 3.27-7.7 µM, respectively. The inhibitory properties of these compounds on the concentration of beta-catenin by ELISA revealed significant percent inhibition of target protein at 100 µg/ml. Conclusion: In conclusion, the synthesized compounds showed significant anti-tumor activities both in silico and in vitro, having potential for further investigating its role in colorectal cancer.

Cytochalasan Alkaloids as TRAIL Sensitizers from an Endophytic Fungus Chaetomium sp.

Planta Medica ◽  
2021 ◽  
Author(s):  
Hanli Ruan ◽  
Ying Gao ◽  
Ruihua Mao ◽  
Ye Liu ◽  
Ming Zhou
Keyword(s):  
Colorectal Cancer ◽  
Absolute Configuration ◽  
Endophytic Fungus ◽  
Cancer Cell Line ◽  
Ring System ◽  
Ht29 Cell ◽  
Colorectal Cancer Cell Line ◽  
X Ray ◽  
Fibrous Roots ◽  
Trail Sensitivity

Two new cytochalasans with a rare 6/6/5/5/7 pentacyclic ring system, named chaetoconvosins C−D (1−2), together with two known congeners (3−4), were isolated from the fermentation of an endophytic fungus, Chaetomium sp. SG-01, harbored in the fibrous roots of Schisandra glaucescens Diels. Their structures including the absolute configuration were elucidated by extensive spectroscopic (HRESIMS, NMR, and ECD) and X-ray crystallographic analyses. The TRAIL sensitivity of 1–4 in a TRAIL-resistant HT29 colorectal cancer cell line was evaluated, which revealed that co-treatment of 1–4 at 50 µM with TRAIL (150 ng/mL) reduced the HT29 cell viability by 19.0%, 24.1%, 17.9%, and 15.5%, respectively, compared to treatment with 1–4 alone.

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