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Principal Investigator: Tingting Yang
Organization: COLUMBIA UNIVERSITY HEALTH SCIENCES
Fiscal Year: 2024
Award: $411,250
Funding agency: National Institute of General Medical Sciences
Project Summary/Abstract
The bestrophin proteins are a family of Ca2+-activated anion channels widely distributed from bacteria to
mammals, with representatives in most metazoans including four in humans (Best1-4). They open in response
to increases of intracellular Ca2+ to mediate the passive flow of Cl− and other anions. Best1 and Best2 both have
critical roles in the eye. Best1 is predominantly expressed in retinal pigment epithelium (RPE) of the retina playing
an essential role in generating a vision-related electrical signal named “light peak”, while mutations in the human
BEST1 gene have been genetically linked to at least five retinal degenerative disorders collectively known as
bestrophinopathies. Best2 is highly expressed in non-pigmented epithelium (NPE) of the ciliary body, and is
involved in aqueous humor formation and drainage, which ultimately determine intra-ocular pressure (IOP).
Knockout of Best2 in mice leads to a reduction of IOP, suggesting the pharmaceutical potential of targeting Best2
and its regulators for relieving ocular hypertension, which is a common risk factor for numerous eye diseases
including open-angle glaucoma. However, despite bestrophins’ biological and pharmaceutical significance, little
is known about their cellular regulation in a physiological context. More specifically, in a given cell/tissue, how is
the bestrophin channel modulated to conduct which anion(s) for what downstream purpose(s)? The answer to
this question lies within the protein network that interacts with bestrophins in different tissues/cells. Although
ample evidence suggest the existence of tissue-specific interacting regulators of bestrophins, no protein is known
to interact with Best2, while only a few have been reported to interact with Best1. Our lab studies the biophysics
and regulation of bestrophin channels using a multidisciplinary platform consisting of cryogenic electron
microscopy (cryo-EM), mass spectrometry, electrophysiological recording, CRISPR/Cas9-mediated genome
editing, and stem cell reprogramming/differentiation. The goal of the next five years is to identify Best1 and Best2
interacting protein complexes in RPE and NPE, respectively, and to understand the functional consequences of
these interactions. Overall, the proposed work will make contributions to multiple fields of research including
calcium signaling, ion transport, membrane protein structure and ocular physiology, and the pipelines established
in this work can be generally applied to study different ion channels and other membrane proteins of interest.
Terms: <Anions><Aqueous Humor><Bacteria><Biological><Body Tissues><CRISPR approach><CRISPR based approach><CRISPR method><CRISPR methodology><CRISPR technique><CRISPR technology><CRISPR tools><CRISPR-CAS-9><CRISPR-based method><CRISPR-based technique><CRISPR-based technology><CRISPR-based tool><CRISPR/CAS approach><CRISPR/Cas method><CRISPR/Cas technology><CRISPR/Cas9><CRISPR/Cas9 technology><Calcium Ion Signaling><Calcium Signaling><Cas nuclease technology><Cell Body><Cell Communication and Signaling><Cell Reprogramming><Cell Signaling><Cells><Cellular Regulation><Ciliary Body><Ciliary epithelium><Clustered Regularly Interspaced Short Palindromic Repeats approach><Clustered Regularly Interspaced Short Palindromic Repeats method><Clustered Regularly Interspaced Short Palindromic Repeats methodology><Clustered Regularly Interspaced Short Palindromic Repeats technique><Clustered Regularly Interspaced Short Palindromic Repeats technology><Compensated Glaucoma><Compensative Glaucoma><Cryo-electron Microscopy><Cryoelectron Microscopy><Degenerative Disorder><Drainage><Drainage procedure><Electron Cryomicroscopy><Electrophysiology><Electrophysiology (science)><Epithelium><Eye><Eye diseases><Eyeball><Family><Genes><Genetic Alteration><Genetic Change><Genetic defect><Glaucoma Simplex><Goals><Human><Intracellular Communication and Signaling><Intraocular Fluid><Intraocular Pressure><Ion Channel><Ion Transport><Ionic Channels><Knock-out><Knockout><Knowledge><Light><Link><Mammalia><Mammals><Mass Photometry/Spectrum Analysis><Mass Spectrometry><Mass Spectroscopy><Mass Spectrum><Mass Spectrum Analyses><Mass Spectrum Analysis><Mediating><Membrane Channels><Membrane Protein Gene><Membrane Proteins><Membrane-Associated Proteins><Mice><Mice Mammals><Modern Man><Murine><Mus><Mutation><Names><Neurophysiology / Electrophysiology><Ocular Hypertension><Ocular Physiology><Ocular Tension><Open-Angle Glaucoma><Outer pigmented layer of retina><Pharmaceutical Agent><Pharmaceuticals><Pharmacologic Substance><Pharmacological Substance><Photoradiation><Physiologic><Physiologic Intraocular Pressure><Physiological><Physiology of the Eye><Pigment cell layer of retina><Pigmented layer of retina><Play><Progenitor Cells><Proteins><Regulation><Reporting><Research><Retina><Retinal Degeneration><Retinal Pigment Epithelium><Retinal pigment epithelial cells><Risk Factors><Role><Sight><Signal Transduction><Signal Transduction Systems><Signaling><Simple Glaucoma><Structure of retinal pigment epithelium><Surface Proteins><Tissues><Vision><Visual Physiology><Work><biologic><biological signal transduction><biophysical analysis><biophysical studies><cell growth regulation><cellular reprogramming><cryo-EM><cryoEM><cryogenic electron microscopy><degenerative condition><degenerative disease><degenerative retina diseases><electrophysiological><eye disorder><genome editing><genome mutation><genomic editing><interest><intra-ocular pressure><multidisciplinary><name><named><naming><ocular disease><ocular disorder><ocular hypertensive><ophthalmopathy><pharmaceutical><protein complex><protein structure><protein structures><proteins structure><response><retina degeneration><retinal degenerative><retinal degenerative diseases><social role><stem cells><visual function>