Inhibition of Ca ²⁺ channel surface expression by mutant bestrophin‐1 in RPE cells

Abstract

The BEST1 gene product bestrophin‐1, a Ca2+‐dependent anion channel, interacts with CaV1.3 Ca2+ channels in the retinal pigment epithelium (RPE). BEST1 mutations lead to Best vitelliform macular dystrophy. A common functional defect of these mutations is reduced trafficking of bestrophin‐1 into the plasma membrane. We hypothesized that this defect affects the interaction partner CaV1.3 channel affecting Ca2+ signaling and altered RPE function. Thus, we investigated the protein interaction between CaV1.3 channels and bestrophin‐1 by immunoprecipitation, CaV1.3 activity in the presence of mutant bestrophin‐1 and intracellular trafficking of the interaction partners in confluent RPE monolayers. We selected four BEST1 mutations, each representing one mutational hotspot of the disease: T6P, F80L, R218C, and F305S. Heterologously expressed L‐type channels and mutant bestrophin‐1 showed reduced interaction, reduced CaV1.3 channel activity, and changes in surface expression. Transfection of polarized RPE (porcine primary cells, iPSC‐RPE) that endogenously express CaV1.3 and wild‐type bestrophin‐1, with mutant bestrophin‐1 confirmed reduction of CaV1.3 surface expression. For the four selected BEST1 mutations, presence of mutant bestrophin‐1 led to reduced CaV1.3 activity by modulating pore‐function or decreasing surface expression. Reduced CaV1.3 activity might open new ways to understand symptoms of Best vitelliform macular dystrophy such as reduced electro‐oculogram, lipofuscin accumulation, and vision impairment.

Function of the Retinal Pigment Epithelium in Patients With Neurofibromatosis Type 1

“…Recent studies have suggested that the RPE dysfunction in Best disease results from a disruption in Ca2 + -dependent Cl – channels through intracellular Ca2 + signaling. 31 Interestingly, two studies also recorded EOG in patients with ocular albinism (OA) and oculo-cutaneous albinism (OCA) and found a normal or supra-normal LP/DT ratio with significantly low DT amplitudes, 32 , 33 strikingly similar to the pattern found in patients with NF1, whereas albinism could appear as the clinical opposite of NF1. The authors proposed that these results could be explained by the abnormal melanosomal biogenesis and light-induced retinal damage.…”

Novel roles for voltage‐gated T‐type Ca 2+ and ClC‐2 channels in phagocytosis and angiogenic factor balance identified in human iPSC‐derived RPE

“…85,94 The expression of bestrophin-1, CFTR, and ClC-2 were recently confirmed in the hESC-derived RPE. 95 Furthermore, bestrophin-1 has been extensively characterized in hiPSC-derived RPE, 35,96,97 as mutations in the causative gene BEST1 are responsible for several IRDs, referred to as Bestrophinopathies.…”

Knockout of CaV1.3 L-type calcium channels in a mouse model of retinitis pigmentosa

“…The lack of visual improvement despite the delayed degeneration of photoreceptors in rd10/Cav1.3-KO retinas might be due to other factors beside the reduction of Ca 2+ influx in the photoreceptors. We have to consider that Cav1.3 is not the predominant LTCC in the photoreceptors, whereas its expression in the RPE is well documented 18 , 59 – 62 . Therefore, we cannot exclude that an increase in photoreceptors survival was due to a change in the activation or expression of trophic factors from the RPE.…”