Measuring force in the developing zebrafish embryo using an expressible tension sensor
Abstract/Contents
- Abstract
- Early vertebrate development is a mechanically dynamic process. Embryos undergo radical morphologic changes to mold a ball of cells into the recognizable planes of a frog or fish or mouse. In the last 40 years, the zebrafish has emerged as a powerful system for studying these early movements in a vertebrate system. Its large, optically transparent embryos make it a good candidate for advanced microscopy techniques, and in recent years this model organism has been the setting for a host of cutting edge microscopy methodologies. One technique that has been comparatively overlooked in the fish is the use of Fluorescence Lifetime Imaging (FLIM), specifically in conjunction with Forster Resonance Energy Transfer (FRET). FRET is a powerful technique for studying real time, in vivo protein dynamics on the nanometer scale. FLIM allows us to apply this technique in the noisy embryo background without the need for cumbersome spectral bleed-through corrections. In this work, we adapt previously published controls for use in the zebrafish system and then take a systematic approach to developing the best analytical practices for FLIM-FRET in the zebrafish embryo. After developing FLIM-FRET in the early embryo background, we adapt a recently published expressible, FRET-based stress sensor capable of measuring piconewton levels of force in vivo (Grashoff et. al. 2010). We insert this Tension Sensor Module (TSMod) into the zebrafish Epithelial Cell Adhesion Molecule (EpCAM) and use it to make direct measurements of the tensions experienced during early development. In doing so we show that the EpCAM molecule holds between 0.5 and 1.5 piconewtons of force during zebrafish epiboly and that the leading margin is under greater tension than other regions of the developing embryo. This work represents the first use of FLIM-FRET in the zebrafish embryo and the first direct measure of molecular tension in a developing vertebrate.
Description
| Type of resource | text |
|---|---|
| Form | electronic; electronic resource; remote |
| Extent | 1 online resource. |
| Publication date | 2015 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Associated with | Hamilton, Andrea Leigh |
|---|---|
| Associated with | Stanford University, Department of Molecular and Cellular Physiology. |
| Primary advisor | Riedel-Kruse, Hans |
| Thesis advisor | Riedel-Kruse, Hans |
| Thesis advisor | Dunn, Alexander Robert |
| Thesis advisor | Goodman, Miriam Beth |
| Thesis advisor | Talbot, William |
| Advisor | Dunn, Alexander Robert |
| Advisor | Goodman, Miriam Beth |
| Advisor | Talbot, William |
Subjects
| Genre | Theses |
|---|
Bibliographic information
| Statement of responsibility | Andrea Leigh Hamilton. |
|---|---|
| Note | Submitted to the Department of Molecular and Cellular Physiology. |
| Thesis | Thesis (Ph.D.)--Stanford University, 2015. |
| Location | electronic resource |
Access conditions
- Copyright
- © 2015 by Andrea Leigh Hamilton
- License
- This work is licensed under a Creative Commons Attribution Non Commercial Share Alike 3.0 Unported license (CC BY-NC-SA).
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