An Investigation Of The Role Of Nuclear Envelope Proteins On Actin Dynamics
Actin is one of the most abundant proteins in eukaryotic cells. Actin structure and actin dynamics play a critical role in a variety of essential biological functions, including cell mobility, intracellular transport and signal transduction. It has been shown that two nuclear envelope proteins - lamin A/C and emerin may regulate actin dynamics. However, the mechanism behind it remains unclear. Here I tried to elucidate the mechanism of how lamin A/C and emerin regulate actin dynamics from two aspects. In the first part of my work, I focused on the nuclear actin dynamics. I used a nuclear F-actin probe (Utr230-GFP-3xNLS) and DNase I to detect nuclear Factin and nuclear G-actin respectively. Lamin A/C deficient cells expressing this probe showed a punctate structure rather than the short filaments in wild-type cells and emerin deficient cells. DNase I staining also showed a more irregular G-actin distribution in lamin A/C deficient cells compared to circular nuclear G-actin patterns in wild-type and emerin deficient cells. These results indicated lamin A/C may play a role in actin dynamics regulation. I proposed that it may regulate actin dynamics through regulating actin binding proteins. In the second part of this work, I used fluorescence recovery after photobleaching (FRAP) analysis, which is the most widely used method to measure the protein dynamics in live cells, to investigate the cytoplasmic actin dynamics. Unlike previous published data, which showed an enhanced cytoplasmic actin mobility in lamin A/C deficient and emerin deficient cells, my result did not show any difference among them. I proposed that FRAP is not good for cytoplasmic actin dynamics due to the high concentration and mobility of actin.
Actin dynamics; Lamin A/C and emerin; Nuclear actin probe
M.S., Applied Physics
Master of Science
dissertation or thesis