Poster Presentation 2014 International Biophysics Congress

Unsaturated fatty acids drastically accelerate the channel opening of voltage-gated proton channel (#295)

Akira Kawanabe 1 , Yasushi Okamura 1
  1. Graduate School of Medicine, Osaka University, Osaka, Japan

Proton conduction through voltage-gated proton channel (Hv1/VSOP), which is expressed in immunocytes, is controlled by membrane voltage and pH.  Activated phagocytes release arachidonic acid (AA), one species of unsaturated fatty acids, which induces rapid production of reactive oxygen species (ROS) to kill invading pathogens.  Previous studies using whole-cell recording showed that the proton conductance was increased several-fold by AA in neutrophils, macrophages, and eosinophils.  These suggest that the activity of Hv1/VSOP is controlled by AA for efficient ROS generation in phagocytes.  However, the molecular mechanisms of AA modulation for Hv1/VSOP remain unclear.  In order to understand the detailed mechanisms, we quantitatively examined AA effect on Hv1/VSOP heterologously expressed in HEK293T by inside-out patch clamp recording with rapid-perfusion system.

By the application of 50 μM AA, approximately 25-fold increase was observed in the proton current which was evoked by short depolarizing pulse (1 sec) at 100 mV.  The AA-induced increase of proton conductance was significantly larger in this study than in previous studies of native currents.  Analysis with a longer step pulse (60 sec) showed that the increase of the proton current by AA was caused by 2 order shortening of activation time constant and 10 ~ 20 mV negative shift in the voltage-dependence of activation.  Mutation study suggested that the above accelerating kinetics and shift of voltage-dependence were induced by distinct mechanisms.  Experiments with AA derivatives revealed that double bonds and hydrophilicity of head group of AA are essential.  We would discuss the molecular mechanisms of modulation of Hv1/VSOP by unsaturated fatty acids.