Dissertation
New Methodologies for Studying Intercellular Connectivity and Signaling in Plant Tissues
Washington State University
Doctor of Philosophy (PhD), Washington State University
2022
DOI:
https://doi.org/10.7273/000005018
Abstract
Cells are at the core of our biosphere. Their organization into tissues, organs, organ systems, and organisms requires frequent exchange of nutrients and information to promote proper developmental patterning and organismal success. Direct physical connections that unite the cytoplasm of adjacent cells provide the most energy-efficient mechanism for moving substances from one cell to the next. Therefore, it is important to understand both the subcellular mechanisms that govern cell-to-cell transport and the supracellular processes that utilize these physical connections. In a plant organism, intercellular communication is dictated by the numerous pores in a plant cell wall that enable the unity of cytoplasm between adjacent cells within and often between tissues. These so-called plasmodesmata are vital in plant physiology at the supracellular level. As plants cannot escape either biotic or abiotic stressors, they have developed sophisticated chemical signaling processes that utilize plasmodesmata to coordinate a response, if necessary.
The microinjection of fluorescent probes into live cells is an essential component in the toolbox of modern plant cell biology because it allows for a direct assessment of plasmodesmal function. However, traditional methods to introduce fluorescent probes into living cells necessarily introduce artifacts that limit our progress. Herein, I describe the diffusive injection micropipette (DIMP) which was developed to avoid several of the artifacts that impact cell function during traditional injections. I used DIMPs to successfully introduce fluorescent probes into plant, fungal, and animal cells, with a more in-depth investigation into the function of plant cell plasmodesmata. As plasmodesmata are nanoscopic channels with complex anatomical features, they are subject to certain biophysical processes that are absent at the macroscopic level. At the nanoscopic scale, surface charges of the plasma membrane generate an electric field that may be large enough to interact with molecules passing through plasmodesmata. Using the DIMP, I found that positively charged fluorescent probes were greatly impaired in their cell-to-cell movement via plasmodesmata when compared to negatively charged probes. This is intriguing because calcium signaling cascades that are vital to plant stress responses previously had been hypothesized to involve the movement of calcium through plasmodesmata. Therefore, I also investigated the propagation of calcium signals in local cell populations responding to mechanical stimuli. In response to mechanical stimuli that increase cell turgor, calcium signals depended on plasmodesmata for their propagation within the tissue. However, the signaling cascade following turgor decreases were independent of plasmodesmata and likely utilize extracellular pathways.
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Details
- Title
- New Methodologies for Studying Intercellular Connectivity and Signaling in Plant Tissues
- Creators
- Alexander H Howell
- Contributors
- Michael Knoblauch (Advisor)Hans Henning Kunz (Committee Member)Asaph B Cousins (Committee Member)Andrew McCubbin (Committee Member)
- Awarding Institution
- Washington State University
- Academic Unit
- School of Biological Sciences
- Theses and Dissertations
- Doctor of Philosophy (PhD), Washington State University
- Publisher
- Washington State University
- Number of pages
- 157
- Identifiers
- 99901019537701842
- Language
- English
- Resource Type
- Dissertation