In this thesis I describe a novel role for the Endoplasmic Reticulum (ER) in spindle positioning and stability in the early <i>C. elegans</i> embryo. Within the embryo the ER is intimately associated with the microtubule cytoskeleton and I show that the disruption of this association leads to instability of nuclear and spindle positioning. I first describe RNA interference screens that I performed to identify new genes involved in the regulation of spindle positioning. This led to the identification of three ER resident proteins that comprise <i>C. elegans</i> homologs of the Oligosaccharyl Transferase (OST) complex. Through characterisation of the spindle phenotypes induced by knockdown of OST components, I show that these proteins are similar to a diverse collection of mutants affecting the ER, such as the trans-membrane ER protein, <i>ooc-3</i> and the <i>npp</i> (nuclear pore protein) genes. I found that identical nuclear and spindle instability phenotypes could be replicated by using the drug Brefeldin A to rapidly disrupt the ER, suggesting the ER might have a structural role in stabilizing nuclear and spindle position. I also provide evidence suggesting that the association between the ER and the microtubule cytoskeleton involves Clathrin heavy-chain (CHC). CHC may form part of the link between the ER and microtubule networks. Overall my results describe a role for the reticular ER network in stabilizing the assembling mitotic spindle and a potential new role for clathrin in linking the ER and microtubules. I addition, I show that Early Endosomes in the <i>C. elegans</i> embryo are polarized and asymmetrically segregated with the actin-myosin cytoskeleton, and present work optimizing the creation of transgenic lines by microparticle bombardment.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:596117 |
| Date | January 2007 |
| Creators | Andrews, R. K. |
| Publisher | University of Cambridge |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
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