Microtubule Severing Proteins Mediate Minus-End Growth in Drosophila Neurons

Open Access
Author:
Patel, Mit A
Area of Honors:
Biochemistry and Molecular Biology
Degree:
Bachelor of Science
Document Type:
Thesis
Thesis Supervisors:
  • Melissa Rolls, Thesis Supervisor
  • Joseph Reese, Honors Advisor
Keywords:
  • microtubules
  • minus-end
  • neurons
  • severing proteins
  • Drosophila
Abstract:
Microtubules are critical cytoskeletal elements necessary for cell structure and intracellular transport of biomolecules and organelles throughout the cell. Microtubule regulation and polarity is therefore extremely important for maintaining core functions of neurons throughout their lifespan. Microtubule polarity is defined by specific orientation of microtubule ends in the polarized cells; the fast-growing end is the plus-end and the other end is the minus end. In neurons, several microtubule interacting proteins regulate microtubules and their polarity, and defects in these proteins can lead to diseases like hereditary spastic paraplegia, Parkinson’s, Alzheimer’s and other neurodegenerative diseases. While microtubule plus-ends are now quite well understood, relatively little is known about minus-ends. In many cells minus-ends are anchored and static. Severing proteins associate with microtubules and function to cleave microtubules in half. I hypothesized that severing proteins mediate the production of free growing minus-ends and therefore, knockdown of these proteins and mutations in them will reduce the free growing minus-ends in neurons. Genetic tools and live imaging techniques were used to record the microtubule dynamics of sensory neuron dendrites in Drosophila melanogaster. Reduction in severing proteins, spastin, katanin-80, katanin-60L1 and fidgetin resulted in lower ratio of free growing minus-ends to plus-ends in the main dendrites. Additionally, spastin, katanin-60 and katanin-60like mutants also showed reduced levels of free-growing minus-ends in the dendrites. It was proposed that severing proteins cleave the stable γ-TuRC cap from minus-ends such that the newly generated minus-ends can grow by interacting with proteins like patronin and EB, thus contribute in organizing the polarity in neuron dendrites.