Molecular mechanisms of gating in the calcium-activated chloride channel bestrophin

Abstract

Bestrophin (BEST1-4) ligand-gated chloride (Cl-) channels are activated by calcium (Ca2+). Mutation of BEST1 causes retinal disease. Partly because bestrophin channels have no sequence or structural similarity to other ion channels, the molecular mechanisms underlying gating are unknown. Here, we present a series of cryo-electron microscopy structures of chicken BEST1, determined at 3.1 Å resolution or better, that represent the channel’s principal gating states. Unlike other channels, opening of the pore is due to the repositioning of tethered pore-lining helices within a surrounding protein shell that dramatically widens a neck of the pore through a concertina of amino acid rearrangements. The neck serves as both the activation and the inactivation gate. Ca2+ binding instigates opening of the neck through allosteric means whereas inactivation peptide binding induces closing. An aperture within the otherwise wide pore controls anion permeability. The studies define a new molecular paradigm for gating among ligand-gated ion channels.

Mutation-Dependent Pathomechanisms Determine the Phenotype in the Bestrophinopathies

“…Based on criteria as defined in the method section, we selected 56 BD-, 16 ARB-, and the known five ADVIRC-associated mutations (Supplementary Table S1). For the autosomal dominant BD-and, in particular, the ADVIRC-associated mutations there is an apparent trend towards a location of the mutations around the neck and the structurally adjacent flexible helices [20] (Figure 1) implicating an important role of these amino acids in the ion gating processes. In contrast, most of the autosomal recessive mutations are located rather outside of this region (Figure 1).…”

“…Monoallelic carriers of these two classes generally show no ocular symptoms. In contrast, heterozygous carriers of class III, IV, and V mutations exert a dominant-negative effect on the functionality of the channel due to their contribution of defective BEST1 subunits to the final channel complex formation [18][19][20].…”

“…The discovery of the crystal structure of BEST1 orthologues [18,19], and the availability of electron microscopy and electrophysiological recordings revealed that the introduction of various mutations into the chicken BEST1 produces channels with dramatically altered gating properties and diminishes channel inactivation [20]. In cell culture model systems disease-causing BEST1 mutations have been reported to affect protein localization [21][22][23], stability [24][25][26], and ion gating properties of the forming anion channel [27][28][29][30][31].…”

DNA methylation landscape of triple-negative ductal carcinoma in situ (DCIS) progressing to the invasive stage in canine breast cancer

“…BEST1, UCP2 and FXYD1 are genes associated with nutrient and energy transport that have previously been identified as upregulated in cancer cells [60][61][62] . BEST1 and UCP2 encode for transport proteins involved in ligand-gated chloride channels and proton leakage during oxidative phosphorylation, respectively 63,64 , whereas FXYD1 is a subunit of sodium/potassium ATPase, important for active transfer through cell membranes. As for genes involved in signaling pathways whose alterations have the potential to disrupt a wide variety of cell processes, TRAF5 and FUT5 are two candidates that could have a robust impact.…”

“…While BEST1 is hypomethylated in invasive breast cancer compared to healthy breast tissue, it is hypermethylated in ADH and without any change in DCIS stage. BEST1 is involved in the transport of ions through the cell membrane by forming calcium-activated chloride ion channels in epithelial cells 63 . The upregulation of BEST1 in colon cancer cells has been shown to significantly increase cell growth, indicating BEST1 as an essential accelerator of cell proliferation 60 .…”