Biophysical characterization of anion channels in mitochondrion-endoplasmic-reticulum contact sites

The mitochondrion-endoplasmic reticulum (ER) contact sites (MERCs, also known as mitochondrial-associated membranes [MAMs]) are specialized regions of the ER that are in close proximity to the mitochondrion. These organelle structures play essential roles in a variety of processes, such as calcium signaling, lipid metabolism, renin-angiotensin-aldosterone system control, the unfolded protein response, and autophagy. MERCs are known to actively participate in ion transport between the ER and mitochondria. Although active calcium channels in MERCs have been detected, limited studies have been carried out to identify or characterize functional anion channels. Here, we tested whether functional anion channels are present in MERCs. We isolated MERCs from mouse organs (heart and brain) and reconstituted them in planar bilayers. The single-channel properties were recorded in the presence of various anion channel blockers or antagonists (IAA-94, DIDS, A9C, and NPPB). We corroborated the presence of anion channels targeted by these drugs using immunoblotting and immunocytochemistry. Biochemical analysis and immunocytochemistry corroborate that CLIC4, CLIC3, and VDACs are present in MERCs. Our results indicate that anion channels are active in MERCs, which could play a pertinent role in intracellular organelle communication.