State-dependent inhibition of TRPM2 channel by acidic pH

Area of Science:

• Ion Channel Physiology
• Cellular Signaling
• Acid-Base Homeostasis

Background:

• Transient Receptor Potential Melastatin 2 (TRPM2) channels are crucial for oxidative stress signaling in immune cells.
• Extracellular acidosis occurs under physiological and pathological conditions, affecting cell function.
• The modulatory role of acidic pH on TRPM2 channel activity remains largely unexplored.

Purpose of the Study:

• To investigate if extracellular acidic pH modulates the activity of the ADP-ribose-activated TRPM2 channel.
• To elucidate the mechanism and characteristics of TRPM2 channel inhibition by acidic pH.
• To identify key residues in the TRPM2 channel pore involved in pH-dependent modulation.

Main Methods:

• Patch clamp electrophysiology was used to record currents in HEK293 cells expressing human TRPM2 (hTRPM2).
• Cells were exposed to varying acidic pH levels to assess inhibition and activation.
• Alanine substitution mutations were introduced in the outer vestibule of the TRPM2 pore to study residue-specific effects.

Main Results:

• Acidic pH rapidly and completely inhibited induced whole-cell and single-channel currents in a voltage-independent manner.
• Inhibition was largely irreversible upon returning to neutral pH, suggesting inactivation.
• Exposure of closed channels to acidic pH reduced subsequent activation in a pH-dependent manner (IC50 for H+ of 20 μM), with largely reversible inhibition.
• Mutations at Lys(952) and Asp(1002) in the outer pore vestibule significantly reduced or abolished pH-induced inhibition and prevented inactivation.

Conclusions:

• Extracellular acidic pH acts as a negative feedback regulator of TRPM2 channel activity.
• Protons bind to the outer vestibule of the TRPM2 channel pore, inhibiting the channel in a state-dependent manner.
• Specific residues, including Lys(952) and Asp(1002), are critical for mediating this pH-dependent inhibition and inactivation.