# Molecular Physiology of TMEM16/Anoctamin Proteins

> **NIH NIH R01** · EMORY UNIVERSITY · 2021 · $343,200

## Abstract

Understanding the mechanisms by which small molecules are transported across cell membranes is a
fundamental challenge in cell physiology. This application focuses on one family of transport proteins, the
Anoctamins / TMEM16s, because they play diverse and indispensable roles in cellular physiology. The founding
members of the Anoctamin (ANO) family are Ca2+-activated Cl- channels (ANO1 and ANO2). These channels
are ubiquitously expressed and are intimately engaged in keeping our epithelia moist by driving the secretion of
bodily fluids, controlling gut motility, facilitating the secretion of hormones, and regulating neuronal excitability
and smooth muscle contractility, among other functions. Dysfunction of ANO1 has been implicated in a variety
of human disease states including hypertension, colitis, asthma, and lung disease. Genetic disruption of the
ANO1 gene in mice causes major developmental abnormalities, behavioral disorders, altered gastrointestinal
motility, and ability to sense pain. Because ANO1 and ANO2 play such varied but essential roles in cell
physiology, they represent novel targets for therapeutic drug development, but as yet ANOs as drug targets have
received relatively little attention. Recently, 3-D structures of various ANOs including ANO1 have provided
valuable insights into how these proteins work, but major questions remain. The long-range goal of our research
is to understand the structure and function of ANO1 (TMEM16A) and ANO2 (TMEM16B). Specifically in this
application, we focus on the regulation of ANO1 by the phospholipid phosphatidylinositol-(4,5)bisphosphate
(PI(4,5)P2). While PI(4,5)P2 is a minor lipid in the cell membrane, it is clear that it plays a critical, but scantily
understood, role in ANO1 and ANO2 function. We will use a combination of single-cell electrophysiology, directed
mutagenesis, and computational molecular dynamics modeling to elucidate how the opening and closing of
ANO1 is controlled by PI(4,5)P2 and calcium ions. There are 3 aims: (1) We will characterize the biophysical
mechanisms of ANO1 and ANO2 regulation by PI(4,5)P2, the functional interactions between PI(4,5)P2 and
calcium, and the structural requirements of phosphoinositides and inositol phosphates for channel regulation.
(2) We will identify the amino acids involved in PI(4,5)P2 regulation and locate the PI(4,5)P2 binding sites in ANO1
and ANO2. Preliminary data provides strong support for the existence of 3 different PI(4,5)P2 binding sites in
ANO1. (3) We will determine the functional roles for each of the 3 different PI(4,5)P2 binding sites in regulating
ANO1 Ca2+ sensitivity, gating, and inactivation. A compelling reason for comparing ANO1 and ANO2 is that
although these 2 proteins are 70% similar (57% identical) in sequence, ANO1 is stimulated by PI(4,5)P2 while
ANO2 is inhibited. This difference provides a rich opportunity to understand how PI(4,5)P2 binding is coupled to
channel function. These studies will answer pressing ou...

## Key facts

- **NIH application ID:** 10245101
- **Project number:** 5R01GM132598-03
- **Recipient organization:** EMORY UNIVERSITY
- **Principal Investigator:** H. CRISS HARTZELL
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $343,200
- **Award type:** 5
- **Project period:** 2019-09-16 → 2023-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10245101

## Citation

> US National Institutes of Health, RePORTER application 10245101, Molecular Physiology of TMEM16/Anoctamin Proteins (5R01GM132598-03). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10245101. Licensed CC0.

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