Visualizing actin regulatory proteins & the mechanoelectrical Ion channel in inner ear stereocilia

My thesis was a continuation of the “3D Virtual Ear Animation” project for the Imagine RIT Festival held in spring 2010. I worked with Dr. David Corey, a Howard Hughes Medical Institute (HHMI) Investigator, whose research publications I had been reading since the spring of 2010. I learned about information that explains how hearing is transformed into neural signals ultimately perceived by the brain. Motivated by the presence of hearing-impaired students on the campus, I decided to continue this animation project. Another motivation for this topic was the lack of illustrations and animations that captured the latest discoveries, particularly the essential protein molecules that are involved. I had taken advantage of the discoveries made in Dr. Corey’s laboratory. I expect it will be useful for teaching and educating the public on websites devoted to hearing loss. The goal was to illustrate the new molecular understanding of how hearing works. This will help the Deaf become more aware of the causes of hearing loss, and may encourage researchers to invent new solutions to hearing loss that are better than hearing aids. INTRODUCTION The inspiration for my thesis originated with my participation in a “3D Virtual Ear Animation” project for the “Imagine RIT” Festival in the spring of 2010. The goal of the project was to explain how hearing works. It involved a great deal of independent project planning and research, as well as group work, and personal interaction across different disciplines. They included Computer Graphics and Animation, Biomedical Science, Chemistry, and Medical Illustration. The key component of my project was the modeling and simulation of the hair cells transducing their movement into electrical signals that stimulate the auditory nerves. After a careful study of the publications by researchers of the Howard Hughes Medical Institute and the Lawrence Berkeley National Laboratory, I discovered that no one had solved the detailed three-dimensional molecular architecture of the mechano-electrical ion channel in the hair cells of the inner ear. To solve this problem, I adopted the model proposed by Dr. Roderick MacKinnon (Lehninger, 2004) because his model of the potassium ion channel resembles most closely the physiology of this mechano-electrical transduction. When the animation project was completed, the science of the mechano-electrical transduction still intrigued me. During the summer, I was thinking of topics for my Master's thesis. This motivated me to study the movement of hair cells and the various proteins that are involved. Inspired by the presence of hearing-impaired students on the campus, including two of my peers, I decided to continue the animation for my thesis. Motivated by the lack of detailed illustrations and animations that captured the latest discoveries, particularly the essential protein molecule involved, I proposed to create more detailed and accurate models. I expect the animation may help the Deaf become more aware of the causes of hearing loss, and may encourage researchers to invent new solutions to hearing loss. SCIENTIFIC BACKGROUND PART I Unlike the widely studied visual system, a detailed mechanism for hearing has remained elusive to scientists for decades. Hair cells of the inner ear convert the mechanical stimulus of a sound wave into an electrical signal that is sent to the brain. These hair epithelial cells have a bundle of hair-like protrusions emanating from the top surface of the cell, named stereocilia. Stereocilia are connected by fine filaments, called tip links. These tips are stretched every time the hair bundle is deflected by a sound vibration. The tip link is, in turn, connected directly to mechano-electrical ion channel proteins that respond to the stretch by allowing an electrical current into the cell (Schwander et al., 2010). The identity of the mechano-electrical transduction ion channel has been elusive. About seven years ago, a member of the transient receptor potential (TRP) family of ion channels, TRPA1, was proposed as a likely candidate for the transduction channel (Corey et al., 2004). However, a study in 2006 from the same group (Kwan et al., 2006) surprisingly showed that TRPA1 contributes to cold, mechanical, and chemical nociception but is not essential for hair-cell transduction. Other candidates for the transduction channel, transmembrane channel-like 1 and 2 (TMC1 and TMC2), have just been published (Kawashima et al., 2011) but confirmation is still needed. Despite the failure to find the ion channel, some proteins that regulate its transduction function have been identified. Among the many are cadherin 23 (CDH23) and protocadherin 15 (PCDH15). Around the same time, more proteins in hair-cell stereocilia were identified by the study of genes that are linked to deafness, and more were quantified by the technique of mass spectrometry. The progress in understanding hair-cell transduction has been largely driven by the discoveries of genes that are linked to Usher Syndrome (USH). The discoveries have provided insights into the mechanisms that are important for stereocilia development and their functions as mechanosensors (Schwander et al., 2010). Furthermore, scientists now have a better understanding of how the interactions between myosin motor proteins (MYO3A, MYO6A, MYO7A, MYO15A), actin binding proteins (twinfilin, espins), and adaptor proteins (harmonin, whirlin) regulate stereocilia development and function. In summary, our understanding of the cellular and molecular mechanism of hearing has been enhanced both by the study of genes for Usher syndrome deafness, and by the identification with mass spectrometry of about 250 different proteins in stereocilia. Among the actin-binding proteins, twinfilin 2 is the major one in contributing to the staircase architecture of stereocilia by suppressing the length of the stereocilia filamentous actin (F-actin) core of the lower and middle row. TRPA1 was ruled out as the candidate for the mechano-electrical transduction channel; although new candidates have been proposed, the identity of the transduction channel remains elusive (David Corey, personal communication, December 21, 2011).