Effect of the Golgi on Drosophila dendritic microtubule nucleation

Open Access
Author:
McCracken, Christie Joy
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
  • microtubule polarity
  • microtubule nucleation
  • Golgi
  • Drosophila
Abstract:
Microtubule arrays are best studied during mitosis, at which time the centrosome nucleates the minus ends of microtubules while the plus ends grow out in a radial array. Contrary to this model, fully differentiated Drosophila cells, including neurons, do not contain centrosomes (Rogers et al., 2008), so they must use some other means of microtubule nucleation. It has been shown in various cell types that the Golgi apparatus can nucleate microtubules independently of the centrosome (Efimov et al., 2007). Golgi outposts have also been found in neuronal Drosophila dendrites (Ye et al., 2007), a region in which most microtubules are oriented with minus ends directed distally from the cell body. Thus, it has been proposed that Golgi could be nucleating microtubules in Drosophila dendrites. In order to test this hypothesis, I cloned a kinesin/Golgi fusion protein into fly lines with the assumption that the fusion would pull Golgi out of the dendrites (towards microtubule plus ends). Based on antibody staining, several of the fly lines were shown to be expressing the fusion proteins in larvae. These fusions moved about half of the Golgi outposts out of the dendrites and some Golgi into axons, as was seen using fluorescence microscopy to image class IV dendritic arborization (da) neurons. However, imaging Golgi in wild type class I da neurons revealed very few Golgi in the dendrites. This lack of dendritic Golgi in the class I da neurons makes the Golgi an unlikely candidate for nucleating microtubules in dendrites. This inference was supported by analyzing microtubule dynamics using lava lamp (lva) RNAi to knock down Golgi function in the dendrites. In these neurons, microtubule dynamics remained very similar to control neurons. So far, dendritic Golgi outposts show no evidence of being microtubule nucleation sites. However, it would still be interesting to further investigate the kinesin/Golgi fusion lines and their phenotypes in dendrites and axons.