LELAND STANFORD JUNIOR UNIVERSITY, THE
This project seeks to understand how the energy carried by every touch activates force-sensitive ion channels in sensory neurons. These protein complexes, known as mechano-electrical transduction or MeT channels, are critical not only for touch sensation, but also for joint and position sensing (proprioception), regulation of blood pressure, and bladder control. Though force sensation is fundamental to our health and daily perception of the world, very little is known about the mechanisms of this process in any animal, including humans. The senses of touch and vibration are compromised in normal aging and by chronic diseases such as diabetes. Recent estimates suggest that the health costs due to diabetes- and age-related dysfunction of touch and vibration sensation are more than $28 billion annually. This proposal seeks to improve understanding of touch sensation by studying the roundworm C. elegans, a simple animal that whose sense of touch is better studied than our own. What is learned from this research has the potential to provide new insight into possible diagnostic tools and treatments for the degradation of touch sensation.
A core innovation of the parent proposal was to fuse state-of-the-art techniques in cellular electrophysiology, single-channel recording, electron microscopy, and genetics. During the previous funding period, we used this unique fusion of experimental techniques to show that the MeT channel activated by force in vivo is a multisubunit ion channel complex. The known subunits of this complex comprise two pore-forming proteins, MEC-4 and MEC-10, and two accessory subunits, MEC-2 and MEC-6. MEC-4 and MEC-10 belong to the DEG/ENaC super-family, which is conserved in animals, but absent from plants, fungi, and bacteria. MEC-4 is sufficient to form channels in heterologous cells and in vivo, but assembles with MEC-10 when both subunits are expressed. MEC-2 is a cholesterol-binding, integral membrane protein related to stomatin and podocin in mammals. MEC-6, a transmembrane protein with a large extracellular domain, is related to human paraxonase. Both MEC-2 and MEC-6 are required for channel function in vivo, but dispensible for activity in heterologous cells. Today, this MEC-4-dependent ion channel complex remains the only animal MeT channel whose protein subunits are known. As related channel proteins are expressed by sensory neurons in mammals and appear to play essential roles in regulating vascular tone, using C. elegans as a model system to investigate how such channels work is a unique opportunity.
Two goals of the parent award were 1) to determine the contribution of a conserved motif in MEC-4, the essential pore-forming subunit, to force sensitivity in vivo and 2) to identify genes encoding proteins that connect microtubules to each other and to the plasma membrane. A planned enhancement was to develop (in collaboration with Dr. Beth Pruitt?s group) new devices for controlled force delivery and to integrate this novel force clamp with our in vivo electrophysiological analysis of force sensation. We proposed to use supplemental funding to accelerate the pace of our research, increase the scale of our genetic dissection of force sensation, and resolve to two experimental bottlenecks.