Localization of Myosin II and identification of its potential upstream regulators in Dictyostelium discoideum
- UNCG Author/Contributor (non-UNCG co-authors, if there are any, appear on document)
- Ashley Herring (Creator)
- Institution
- The University of North Carolina at Greensboro (UNCG )
- Web Site: http://library.uncg.edu/
- Advisor
- Paul Steimle
Abstract: The thesis work presented here aimed to understand how myosin II is regulated in Dictyostelium discoideum within the context of chemotactic signaling and cell migration. Myosin II filaments are regulated via phosphorylation of the heavy chain tails, mediated by myosin heavy chain kinases (MHCKs). The MHCKs are a key point of regulation of myosin II filament disassembly. However, little is known about upstream regulators of myosin II and therefore, is a major component of this project. The GFP-tagged version of myosin II was electroporated into cells lacking putative upstream regulators in order to study cAMP-mediated myosin II translocation. To determine if the lack of one of these regulators indeed affected myosin II translocation, the phenotype of the mutants were compared to wildtype Ax2 cells. Cells lacking GbpC and GskA demonstrated loss of the wildtype phenotype, while PkbA null, RegA null, and VwkA null cells showed defects, but not the loss of the wildtype phenotype, indicating that GbpC and GskA are necessary for proper myosin II translocation. Another aim focused on studying the effect of the loss of all MHCKs on myosin II translocation. The MHCK D knockout plasmid (MHCKD-KO) was electroporated into MHCK A/B/C null cells, however, none of the isolates contained the disrupted version of the mhkD gene as confirmed by diagnostic PCR. The results indicate that the lack of all MHCKs is lethal to the cells and require a knockdown approach instead. In conclusion, myosin II filament regulation is highly coordinated and relies on a variety of upstream regulators, the loss of which inhibits myosin II function. In addition, the loss of all MHCKs seem to be either be lethal to Dictyostelium cells, indicating another method is needed to study this effect on myosin II filament disassembly and regulation. These studies offer a basis of future research directed at understanding and pinpointing upstream regulators of myosin II filament regulation within the context of chemotaxis and cell migration.
Localization of Myosin II and identification of its potential upstream regulators in Dictyostelium discoideum
PDF (Portable Document Format)
785 KB
Created on 8/1/2015
Views: 737
Additional Information
- Publication
- Thesis
- Language: English
- Date: 2015
- Keywords
- Cell Biology, Dictyostelium, MHCK, Myosin
- Subjects
- Myosin
- Dictyostelium discoideum
- Chemotaxis
- Cell migration