P2X7 receptor-mediated calcium dynamics in HEK293 cells: experimental characterization and modelling approach
A Di Garbo1, S Alloisio, M Nobile
1CNR-Institute of Biophysics, via G Moruzzi 1, 56124 Pisa, Italy. digarbo@pi.ibf.cnr.it
Abstract:
The P2X7 receptor (P2X7R) induces ionotropic Ca²⁺ signalling in different cell types. It plays an important role in the immune response and in the nervous system. Here, the mechanisms underlying intracellular Ca²⁺ variations evoked by 3'-O-(4-benzoyl)benzoyl-ATP (BzATP), a potent agonist of the P2X7R, in transfected HEK293 cells, are investigated both experimentally and theoretically. We propose a minimal model of P2X7R that is capable of reproducing, qualitatively and quantitatively, the experimental data. This approach was also adopted for the P2X7R variant, which lacks the entire C-terminus tail (trP2X7R). Then we introduce a biophysical model describing the Ca²⁺ dynamics in HEK293. Our model gives an account of the ionotropic Ca²⁺ influx evoked by BzATP on the basis of the kinetics model of P2X7R. To explain the complex Ca²⁺ responses evoked by BzATP, the model predicted that an impairment in Ca²⁺ extrusion flux through the plasma membrane is a key factor for Ca²⁺ homeostasis in HEK293 cells.
Insights
The P2X7 receptor (P2X7R) mediates calcium signaling. Modeling revealed that impaired calcium extrusion is crucial for complex calcium responses to BzATP in HEK293 cells.
Area of Science:
- Cellular and Molecular Biology
- Biophysics
- Immunology
Background:
- The P2X7 receptor (P2X7R) is a key mediator of ionotropic calcium (Ca²⁺) signaling.
- P2X7R plays significant roles in immune responses and the nervous system.
- Understanding P2X7R-mediated Ca²⁺ dynamics is crucial for cellular function.
Purpose of the Study:
- To investigate the mechanisms of intracellular Ca²⁺ variations induced by the P2X7R agonist BzATP in HEK293 cells.
- To develop and validate computational models for P2X7R kinetics and Ca²⁺ dynamics.
- To elucidate the factors contributing to complex Ca²⁺ responses.
Main Methods:
- Experimental investigation of Ca²⁺ signaling in HEK293 cells transfected with P2X7R.
- Development of a minimal P2X7R kinetics model.
- Construction of a biophysical model for Ca²⁺ dynamics.
- Analysis of a P2X7R variant lacking the C-terminus (trP2X7R).
Main Results:
- A minimal P2X7R model accurately reproduced experimental Ca²⁺ signaling data.
- The biophysical model explained BzATP-evoked Ca²⁺ influx based on P2X7R kinetics.
- Model predictions highlighted the critical role of impaired Ca²⁺ extrusion in cellular Ca²⁺ homeostasis.
Conclusions:
- The developed models provide a quantitative framework for understanding P2X7R-mediated Ca²⁺ signaling.
- Impaired Ca²⁺ extrusion flux is identified as a key determinant of complex Ca²⁺ responses to BzATP.
- This study offers insights into Ca²⁺ homeostasis regulation in HEK293 cells via P2X7R.
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