The Role of KLP-4 in Regulating Cholinergic Signaling in vivo

Cellular trafficking is dominated by the use of motor proteins trafficking cargo along the microtubules that make up the cellular cytoskeleton. Polarity within these cells controls the direction of the trafficking, and the cargo consists of organelles, new proteins, neurotransmitters, etc.. Motor proteins differ in the cargo they carry, the direction that they move, as well as the rate and distance that they can travel. The trafficking of neurotransmitters occurs along neurons, highly polarized cells, and this research focuses on neuronal trafficking as a method of controlling the release and degradation of acetylcholine and thus regulating the signals that they propagate. KLP-4 is currently identified as a motor protein that contributes to the regulation of acetylcholine trafficking. KLP-4 as a part of the kinesin-3 super family of motor proteins is especially important due cells’ reliance upon its function as a long distance and highly processive motor protein. This characteristic of the kinesin-3 family is essential, especially when considering the extraordinary lengths that neurons can cover in a given organism. Having identified hypersensitivity to acetylcholine was present in mutant klp-4 Caenorhabditis elegans, rescues of the mutant klp-4 in C. elegans were performed in order to further establish the role that the motor protein has in the regulation of this heavily used signaling pathway. C. elegans were used as the model organism due to their homology to the human nervous system as well as the extent of literature on their success as in vivo models. Rescue crosses as well as transgenic C. elegans were created in order to test their behavior and the activity of their cholinergic signaling in comparison to the aforementioned mutant klp-4 counterparts. The rescued crosses showed recovery of the phenotype, consistent with the known literature that describes the complex and heavily regulated cholinergic signaling pathway. Discovering the an in-frame 200 amino acid deletion and one amino acid insertion within the cargo binding domain of klp-4, leads to the hypothesis that the KLP-4 motor is constitutively active and thus contributing to the hypersensitivity phenotype. The participation of CHA1 and Unc17 as co regulators in cholinergic signaling will be explored as possible opposite mutants (hyposensitive mutants to the effects of acetylcholine) when crossed with the original klp-4 mutants will show a true rescue of the cholinergic signaling mutant phenotype.