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A High-throughput Cre-Lox Activated Viral Membrane Fusion Assay to Identify Inhibitors of HIV-1 Viral Membrane Fusion
Published on: August 14, 2018
Transforming activity of purinergic receptor P2Y, G-protein coupled, 2 revealed by retroviral expression screening
Hisashi Hatanaka1, Shuji Takada, Young Lim Choi
1Division of Functional Genomics, Jichi Medical University, Shimotsukeshi, Tochigi 329-0498, Japan.
Abstract:
Colorectal cancer (CRC) is one of the leading causes of cancer death in humans. In order to identify novel cancer-promoting genes in CRC, we here constructed a retroviral cDNA expression library from a CRC cell line RKO, and used it for a focus formation assay with mouse 3T3 fibroblasts, leading to the identification of 42 independent cDNAs. One of such cDNAs turned out to encode purinergic receptor P2Y, G-protein coupled, 2 (P2RY2). The oncogenic potential of P2RY2 was confirmed in vitro with the focus formation assay as well as soft agar-growth assay, and also in vivo with a tumorigenicity assay in nude mice. While our P2RY2 cDNA encodes a protein with two amino-acid substitutions compared to the reported one, we have confirmed that the wild-type P2RY2 has a strong transforming potential as well. These results indicate an unexpected role of P2RY2 in the carcinogenesis of human cancers.
Insights
Researchers identified purinergic receptor P2Y2 (P2RY2) as a novel gene promoting colorectal cancer (CRC) development. This finding reveals an unexpected role for P2RY2 in human cancer carcinogenesis.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Colorectal cancer (CRC) is a significant cause of cancer mortality worldwide.
- Identifying novel genes driving CRC progression is crucial for developing targeted therapies.
Purpose of the Study:
- To identify novel cancer-promoting genes in colorectal cancer.
- To investigate the oncogenic potential of the identified purinergic receptor P2Y2 (P2RY2).
Main Methods:
- Construction of a retroviral cDNA expression library from a CRC cell line.
- Focus formation assay in mouse 3T3 fibroblasts to screen for oncogenic cDNAs.
- In vitro (focus formation, soft agar) and in vivo (tumorigenicity assay in nude mice) validation of P2RY2's oncogenic potential.
Main Results:
- Identification of 42 independent cDNAs, including one encoding P2RY2.
- Confirmation of P2RY2's oncogenic potential through multiple assays.
- Demonstration that both the identified P2RY2 variant and wild-type P2RY2 exhibit strong transforming activity.
Conclusions:
- Purinergic receptor P2Y2 (P2RY2) plays an unexpected and significant role in colorectal cancer carcinogenesis.
- P2RY2 represents a potential novel therapeutic target for human cancers.
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