Date of Award
Doctor of Philosophy (PhD)
The advent of single-molecule fluorescence imaging and super-resolution microscopy has produced a range of powerful tools that are able to interrogate biological systems at spatial and temporal resolutions that were previously thought to be well beyond the reach of the visible spectrum of light. While these new imaging techniques have already yielded impressive results across a broad range of studies, they still face a number of limitations. The work described herein aims to address some of these limitations by furthering the use of these techniques for the investigation of in vivo protein dynamics. Specifically, total internal reflection fluorescence (TIRF) microscopy is used on fluorescently tagged proteins in the Chlamydomonas reinhardtii intraflagellar transport (IFT) model system to interrogate both the dynamics involved in IFT, which has been linked to a number of diseases, and to shed light onto some of the dynamics involved with intracellular active transport. The first two studies described here constitute investigations into the dynamics of IFT cargo and IFT motor proteins, respectively, once they have arrived at the plus ends of microtubules at the flagellar tip. The third section consists of the observation and analysis of speed variations along IFT tracks and the potential causes thereof. The final segment touches on a number of nascent explorations into studying protein flagellar entry mechanisms and, separately, the use of intensity profile standard deviations to extract IFT protein complex size and localization information.
Chair and Committee
Yan Mei Wang
Robert Blankenship, Anders Carlsson, Susan Dutcher, Ralf Wessel
Kovacs, Anthony Paul, "Motor and Cargo Dynamics During Intraflagellar Transport: A Single Molecule Investigation" (2015). Arts & Sciences Electronic Theses and Dissertations. 658.