The hyposensitive N187D P2X7 mutant promotes malignant progression in nude mice
Jing-Hui Chong1, Guo-Guang Zheng, Yuan-Yuan Ma
1State Key Laboratory of Experimental Hematology, Institute of Hematology and Blood Diseases Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, 288 Nanjing Road, Tianjin 300020, China.
Abstract:
Nucleotides are new players in the intercellular communication network. P2X7 is a member of the P2X family of receptors, which are ATP-gated plasma membrane ion channels with diverse biological functions. Abnormal expression and dysfunction of P2X7 have been reported in leukemias. Here, we report a new P2X7 mutant (an A(559)-to-G substitution causing N187D P2X7) cloned from J6-1 leukemia cells. The characteristics of N187D P2X7 were studied by establishing stably transfected K562 cell lines. Our results show that N187D P2X7 required a higher concentration of agonist for its activation, leading to Ca(2+) influx (EC(50) = 293.3 ± 6.6 μm for the mutant and 93.6 ± 2.2 μm for wild-type P2X7) and ERK phosphorylation, which were not caused by differential cell-surface expression or related to high ATPase activity on the cell surface and in the extracellular space. K562 cells expressing this N187D mutant showed a proliferative advantage and reduced pro-apoptosis effects in vitro and in vivo. Furthermore, elevated angiogenesis and CD206-positive macrophage infiltration were found in tumor tissues formed by K562-M cells. In addition, higher expression of VEGF and MCP1 could be detected in tumor tissues formed by K562-M cells. Our results suggest that N187D P2X7, representing mutants hyposensitive to agonist, might be a positive regulator in the progression of hematopoietic malignancies.
Insights
A novel P2X7 receptor mutant (N187D P2X7) found in leukemia cells exhibits reduced sensitivity to ATP. This P2X7 variant promotes cancer cell proliferation and tumor growth, suggesting a role in hematopoietic malignancies.
Area of Science:
- Cellular Biology
- Molecular Biology
- Oncology
Background:
- Nucleotides act as signaling molecules in intercellular communication.
- P2X7 receptors are ATP-gated ion channels involved in various biological processes.
- P2X7 receptor dysfunction is implicated in leukemias.
Purpose of the Study:
- To characterize a newly identified P2X7 receptor mutant (N187D P2X7) from leukemia cells.
- To investigate the functional consequences of N187D P2X7 in leukemia cell lines.
- To assess the role of N187D P2X7 in the progression of hematopoietic malignancies.
Main Methods:
- Cloning of the N187D P2X7 mutant from J6-1 leukemia cells.
- Establishment of stably transfected K562 cell lines expressing wild-type and mutant P2X7.
- Measurement of agonist-induced calcium influx and ERK phosphorylation.
- In vitro and in vivo assays for cell proliferation, apoptosis, angiogenesis, and macrophage infiltration.
- Analysis of VEGF and MCP1 expression in tumor tissues.
Main Results:
- N187D P2X7 exhibits reduced sensitivity to ATP (higher EC50 for Ca(2+) influx).
- Mutant P2X7 expression confers a proliferative advantage and reduces apoptosis in K562 cells.
- Tumors formed by N187D P2X7 expressing cells show increased angiogenesis, CD206+ macrophage infiltration, VEGF, and MCP1 levels.
- Differential cell-surface expression or ATPase activity does not explain the observed effects.
Conclusions:
- The N187D P2X7 mutant, characterized by hyposensitivity to agonists, may promote cell proliferation and survival in hematopoietic malignancies.
- This P2X7 variant could be a positive regulator in the progression of leukemia.
- Targeting P2X7 receptor mutations may offer therapeutic strategies for hematologic cancers.
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