Microtubule polarity and dynamics in Drosophila neuronal dendrites

Open Access
Author:
Folker, Christin Theresa
Area of Honors:
Biochemistry and Molecular Biology
Degree:
Bachelor of Science
Document Type:
Thesis
Thesis Supervisors:
  • Melissa Rolls, Thesis Supervisor
  • Chen Pei David Tu, Honors Advisor
  • Scott Brian Selleck, Faculty Reader
Keywords:
  • neurons
  • development
  • microtubules
  • polarity
  • dynamics
Abstract:
The neuron is a polar cell, with axons and dendrites sending and receiving signals, respectively. It has been determined that microtubules play a key role sub-compartmental neuron organization by serving as tracks for motor proteins like kinesin and dynein to transport cargo throughout the cell. As such, microtubule orientation, or polarity, is important to the organization of the cell as a whole. In most cells, the microtubule array is nucleated at the centrosome, with the microtubule minus ends anchored to the centrioles (Alberts et al. 2008). However, in neurons the dendrites have microtubules with minus ends out, nucleated at an unknown source. My first objective in this study was to screen various candidate proteins to identify those that may be involved in determining microtubule orientation in the dendrite. RNA interference (RNAi) was used to reduce expression of each of the genes that may affect microtubule polarity. Of the proteins screened, only three: pk, esn, and kat-60L1, showed a statistically significant phenotype (p<0.05). During the screen, it was observed that reducing levels of Dgrip84, a component of the gamma-tubulin small complex (γ-TuSC) and gamma-tubulin ring complex (γ-TuRC) caused slight changes in the number of growing microtubules in the axon. A more aggressive injury assay was developed to determine more conclusively whether Dgrip84 is involved in microtubule nucleation in the dendrites. The typical injury response involves dramatically increased microtubule dynamics, however I hypothesized that cells lacking γ-TuRC and γ-TuSC proteins will not be able to up-regulate dynamics efficiently. Results showed that all Dgrip RNAi followed a trend of reduced ability to up-regulate dendrite microtubule dynamics in response to axotomy, however the phenotype was only statistically significant (p<0.05) for Dgrip91 RNAi and Dgrip128 RNAi. This finding suggests that not only the γ-TuSC, but the γ-TuRC as a whole, is important for non-centrosomal MT nucleation.