Dynamin localization in Caenorhabditis Elegans

Dynamin is a 100-KDa GTPase that is essential for endocytosis. The classic model of endocytosis is a sequential mechanism: first, cell membrane is bent and internalized via the formation of a clathrin coat; next, dynamin facilitates GTP hydrolysis, resulting in membrane scission as the newly formed vesicle is pinched off. My project is an examination of the function and localization of the dynamin protein within the context of the organism Caenorhabditis elegans. I have also worked toward the development of cleaner and more definitive ways to investigate dynamin and how neurons are affected by a loss of the protein. The presumed role of dynamin led us to hypothesize that the protein was needed for viability. Using the MosDEL technique (designed by Christian Frøkjær-Jensen), we generated a targeted knockout of the dynamin gene, and observed the predicted lethal phenotype. Animals lacking the dynamin protein fail to hatch or arrest as L1 animals. We were able to rescue the lethal phenotype by supplying dynamin both as a single-copy MosSCI insert and as an extrachromosomal array. To further investigate the action of dynamin, fluorescently-tagged versions of the dynamin protein were generated. We differentially tagged both splice isoforms of the dyn-1 protein and discovered distinct localization patterns of the splice variants: one isoform appears to be localized mainly to the axons of neurons, while the other persists in cell bodies and dendrites. This pattern is consistent regardless of copy number.