Book Description

Mechanical signaling plays an important role in cell shape and volume regulation, touch sensation, hearing, proprioception, gravitaxis, and turgor regulation. C. elegans provides a powerful model for elaborating mechanisms of eukaryotic mechanotransduction. Genetic screening identified candidate touch-transducing channels (DEG/ENaCs and TRP channels). In C. elegans, six touch neurons (ALML/R, AVM, PLML/R, PVM) are located in specific places in the body, optimized to detect forces delivered to those parts of the body. MEC-4 is expressed in six touch sensory neurons. MEC-10, on the other hand, is expressed in these six neurons, as well as in two extra pairs of neurons, PVDL/R and FLPL/R. Laser ablation studies showed that the six touch neurons respond to gentle and harsh body touch and suggested that FLP and PVD neurons are responsible for the harsh touch response. MEC-10 encodes a component of the core gentle touch sensory channel that is expressed in both gentle touch and harsh touch neurons. I studied the first mec-10 null mutant and showed that MEC-10 is required for both gentle and harsh touch sensation in C. elegans since the mec-10 null mutant is gentle touch insensitive and reduces harsh touch responses. We also used the intracellular calcium reporter cameleon to show that responses of gentle touch neurons and PVD/FLP to touch stimuli decreased in mec-10 null mutant. However, mec-10 null mutation has no significant impact on proprioception and mec-10(d)-induced neurodegeneration. I also made mec-4 and mec-10 hybrid proteins by switching their extracellular and transmembrane domains and checked their function by rescuing assay. Failure to complement the touch sensation function suggested that specific sequences are required for the normal functions of mec-4 and mec-10; smaller perturbation may be needed to recover protein function in chimeras. Based on the solved MEC-4 N-terminal NMR structure prediction, I introduced point mutations into this domain and studied biological consequences in genetic rescue assays and by monitoring dominant negative effects normally seen when the N-terminal is expressed alone. I found that generally, the amino acid substitutions predicted to perturb structure disrupt channel function as predicted. The disrupted mutant strains can also exhibit a significantly decreased density of immuno-stained channel puncta distributed along touch neuron processes. However, the rescue of channel function and the dominant negative effects are not well correlated. Overall, my data advance understanding of MEC-10 and MEC-4 function on mechanosensation.