Endogenous pannexin1 channels form functional intercellular cell-cell channels with characteristic voltage-dependent...

Area of Science:

• Cell Biology
• Ion Channel Physiology
• Molecular Biophysics

Background:

• The existence and function of intercellular channels formed by pannexin1 (Panx1) have been debated for over a decade.
• Connexins (Cxs) are well-established gap junction proteins, but Panx1's role in direct cell-cell communication remains less understood.

Purpose of the Study:

• To electrophysiologically characterize exogenous human pannexin1 (hPanx1) cell-cell channels.
• To investigate the properties and functional relevance of hPanx1 channels in endogenous settings.

Main Methods:

• Electrophysiological recordings (single-channel and dual-cell recordings) in HeLa cells lacking connexin45 (KO Cx45) expressing exogenous hPanx1.
• Characterization of hPanx1 channels in TC620 cells (endogenously expressing hPanx1) under varying conditions (temperature, knockdown, overexpression).
• In silico modeling to explore pore properties and potential mechanisms for dye passage.

Main Results:

• Exogenous hPanx1 channels exhibited two distinct states (O-state and S-state) with unique voltage sensitivity and conductance properties.
• S-state channels showed dynamic asymmetry and conductance matching the O-state substate.
• Endogenous hPanx1 channels in TC620 cells displayed temperature-dependent coupling, reduced by Panx1 knockdown/inhibition, and increased by overexpression.
• Octanol, a connexin inhibitor, did not block hPanx1-mediated coupling.
• In silico analysis suggested arginine residues in the pore may facilitate DAPI passage.

Conclusions:

• This study provides robust electrophysiological evidence that hPanx1 forms functional intercellular channels with distinct properties.
• Endogenously expressed hPanx1 contributes to cell-cell coupling, independent of connexins.
• Understanding Panx1 channel characteristics is vital for recognizing its role in various cell types and physiological processes.