The Activation and Biophysical Properties of Anoctamin 8

Abstract

ANO8 is a member of the Anoctamin/TMEM16 family which was previously defined as a superfamily of calcium-activated chloride channels after the identification of ANO1. However, it was found that the Anoctamin family consist of both cation and anion channels due to the identification of ANO9 as a non-selective cation channel. Among them, ANO8 shows low structural homology from the other Anoctamin members and it has not been well studied. Therefore, the aim of the research was to investigate not only the activation of ANO8 but also its biophysical properties.

The plasma membrane localization of ANO8 in HEK293T cells was visualised by immunofluorescent images. This led to examining the electrophysiological characteristics of ANO8 with whole-cell patch clamp recordings. It was unveiled that ANO8 was activated by intracellular cyclic adenosine monophosphate as well as high concentrations of calcium. Moreover, the cAMP-induced currents of ANO8 was enhanced by the addition of intracellular calcium.

The results from RT-PCR suggest that TMEM16H had an unusually high distribution in brain, spinal cord and dorsal root ganglion neurons. Immunostaining of dorsal root ganglion reveals ANO8 is highly expressed in nociceptive neurons.

Next, the function of ANO8 was examined by calcium imaging and it potentiated the responses of TRPV1 and TRPA1 channels that are involved in the transduction of noxious stimuli.

Taken together, this study reveals how ANO8 is activated and its other biophysical characteristics.