Skip to main content
eScholarship
Open Access Publications from the University of California

Scholarly Works (6 results)

  • Thesis
  • Peer Reviewed
UC Davis Electronic Theses and Dissertations (2024)

The cell wall is an integral and dynamic structure surrounding plant cells, involved in plant growth and development. During cytokinesis, cell plate formation takes place, via fusion of cytokinetic vesicles, that after membrane transformation and deposition of polysaccharides transitions to a new cross wall. Callose, a β(1,3) glucan polysaccharide, is transiently deposited at the cell plate during this fundamental process. Callose deposition is thought to play a vital role in stabilizing the cell plate and contributing to a spreading force during cell plate expansion.

Cytokinesis-specific callose synthases are recalcitrant to study by genetic means, because null mutations in the corresponding genes cause lethality. Additionally, limitations in live cell imaging of polysaccharides add to the difficulties in elucidating the biological role and regulation of callose biosynthesis at the cell plate.

To overcome these challenges, I have taken a multidisciplinary approach. To better understand the transition from a vesicular network to a fenestrated sheet and finally a mature cell plate, we implemented a modeling approach, adopting the Helfrich energy that examines the elastic properties of lipid bilayers. Our model predicts the requirement of a spreading/stabilizing force for cell plate expansion and maturation. The transient accumulation of callose, coinciding with the predicted cell plate stages requiring this spreading force, are consistent with the proposed model. Furthermore, we used a specific cytokinetic callose inhibitor, Endosidin7 (ES7), to dissect the role of callose. ES7 does not inhibit wound or stress-induced callose deposition; however, it specifically causes failure in cell plate expansion and maturation, indicating that callose is a contributor to the stabilizing or spreading force as predicted by our model.

We further used advanced light microscopic techniques to explore cell plate dynamics. I studied cell plate development across four dimensions using a cytokinesis-specific GTPase YFP-RABA2a as a vesicle marker. With the aid of a robust cell plate volume analysis pipeline, I identified three easily trackable cell plate developmental phases that can be interrogated to study cell plate development. Inhibition of callose through ES7 suppressed phase transitions, establishing a critical role and timing of polysaccharide deposition in cell plate expansion and maturation.

Using this suite of interdisciplinary techniques, we contributed to breaking apart the molecular black box surrounding callose deposition during cytokinesis.

    Cover page: Dissection of cell plate dynamics during cytokinesis and the role of callose in cell plate maturation
    • Article
    • Peer Reviewed
    UC Davis Previously Published Works (2018)
    The cell wall, a complex macromolecular composite structure surrounding and protecting plant cells, is essential for development, signal transduction, and disease resistance. This structure is also integral to cell expansion, as its tensile resistance is the primary balancing mechanism against internal turgor pressure. Throughout these processes, the biosynthesis, transport, deposition, and assembly of cell wall polymers are tightly regulated. The plant endomembrane system facilitates transport of polysaccharides, polysaccharide biosynthetic and modifying enzymes and glycoproteins through vesicle trafficking pathways. Although a number of enzymes involved in cell wall biosynthesis have been identified, comparatively little is known about the transport of cell wall polysaccharides and glycoproteins by the endomembrane system. This review summarizes our current understanding of trafficking of cell wall components during cell growth and cell division. Emerging technologies, such as vesicle glycomics, are also discussed as promising avenues to gain insights into the trafficking of structural polysaccharides to the apoplast.
      Cover page: Post-Golgi Trafficking and Transport of Cell Wall Components
      • Article
      • Peer Reviewed
      UC Davis Previously Published Works (2022)
      Plant cytokinesis, a fundamental process of plant life, involves de novo formation of a "cell plate" partitioning the cytoplasm of dividing cells. Cell plate formation is directed by orchestrated delivery, fusion of cytokinetic vesicles, and membrane maturation to form a nascent cell wall by timely deposition of polysaccharides. During cell plate maturation, the fragile membrane network transitions to a fenestrated sheet and finally a young cell wall. Here, we approximated cell plate sub-structures with testable shapes and adopted the Helfrich-free energy model for membranes, including a stabilizing and spreading force, to understand the transition from a vesicular network to a fenestrated sheet and mature cell plate. Regular cell plate development in the model was possible, with suitable bending modulus, for a two-dimensional late stage spreading force of 2-6 pN/nm, an osmotic pressure difference of 2-10 kPa, and spontaneous curvature between 0 and 0.04 nm-1. With these conditions, stable membrane conformation sizes and morphologies emerged in concordance with stages of cell plate development. To reach a mature cell plate, our model required the late-stage onset of a spreading/stabilizing force coupled with a concurrent loss of spontaneous curvature. Absence of a spreading/stabilizing force predicts failure of maturation. The proposed model provides a framework to interrogate different players in late cytokinesis and potentially other membrane networks that undergo such transitions. Callose, is a polysaccharide that accumulates transiently during cell plate maturation. Callose-related observations were consistent with the proposed model's concept, suggesting that it is one of the factors involved in establishing the spreading force.
        Cover page: A biophysical model for plant cell plate maturation based on the contribution of a spreading force.
        • Article
        • Peer Reviewed
        UC Davis Previously Published Works (2024)
        Cell plate formation during cytokinesis entails multiple stages occurring concurrently and requiring orchestrated vesicle delivery, membrane remodelling, and timely deposition of polysaccharides, such as callose. Understanding such a dynamic process requires dissection in time and space; this has been a major hurdle in studying cytokinesis. Using lattice light sheet microscopy (LLSM), we studied cell plate development in four dimensions, through the behavior of yellow fluorescent protein (YFP)-tagged cytokinesis-specific GTPase RABA2a vesicles. We monitored the entire duration of cell plate development, from its first emergence, with the aid of YFP-RABA2a, in both the presence and absence of cytokinetic callose. By developing a robust cytokinetic vesicle volume analysis pipeline, we identified distinct behavioral patterns, allowing the identification of three easily trackable cell plate developmental phases. Notably, the phase transition between phase I and phase II is striking, indicating a switch from membrane accumulation to the recycling of excess membrane material. We interrogated the role of callose using pharmacological inhibition with LLSM and electron microscopy. Loss of callose inhibited the phase transitions, establishing the critical role and timing of the polysaccharide deposition in cell plate expansion and maturation. This study exemplifies the power of combining LLSM with quantitative analysis to decode and untangle such a complex process.
          Cover page: Four-dimensional quantitative analysis of cell plate development in Arabidopsis using lattice light sheet microscopy identifies robust transition points between growth phases.
          • Article
          • Peer Reviewed
          UC Berkeley Previously Published Works (2017)
          The engineering of living plants for visible light emission and sustainable illumination is compelling because plants possess independent energy generation and storage mechanisms and autonomous self-repair. Herein, we demonstrate a plant nanobionic approach that enables exceptional luminosity and lifetime utilizing four chemically interacting nanoparticles, including firefly luciferase conjugated silica (SNP-Luc), d-luciferin releasing poly(lactic-co-glycolic acid) (PLGA-LH2), coenzyme A functionalized chitosan (CS-CoA) and semiconductor nanocrystal phosphors for longer wavelength modulation. An in vitro kinetic model incorporating the release rates of the nanoparticles is developed to maximize the chemiluminescent lifetimes to exceed 21.5 h. In watercress (Nasturtium officinale) and other species, the nanoparticles circumvent limitations such as luciferin toxicity above 400 μM and colocalization of enzymatic reactions near high adenosine triphosphate (ATP) production. Pressurized bath infusion of nanoparticles (PBIN) is introduced to deliver a mixture of nanoparticles to the entire living plant, well described using a nanofluidic mathematical model. We rationally design nanoparticle size and charge to control localization within distinct tissues compartments with 10 nm nanoparticles localizing within the leaf mesophyll and stomata guard cells, and those larger than 100 nm segregated in the leaf mesophyll. The results are mature watercress plants that emit greater than 1.44 × 1012 photons/sec or 50% of 1 μW commercial luminescent diodes and modulate "off" and "on" states by chemical addition of dehydroluciferin and coenzyme A, respectively. We show that CdSe nanocrystals can shift the chemiluminescent emission to 760 nm enabling near-infrared (nIR) signaling. These results advance the viability of nanobionic plants as self-powered photonics, direct and indirect light sources.
            Cover page: A Nanobionic Light-Emitting Plant
            • Article
            • Peer Reviewed
            UC Davis Previously Published Works (2023)
            INTRODUCTION: During proliferative plant cell division, the new cell wall, called the cell plate, is first built in the middle of the cell and then expands outward to complete cytokinesis. This dynamic process requires coordinated movement and arrangement of the cytoskeleton and organelles. METHODS: Here we use live-cell markers to track the dynamic reorganization of microtubules, nuclei, endoplasmic reticulum, and endomembrane compartments during division and the formation of the cell plate in maize leaf epidermal cells. RESULTS: The microtubule plus-end localized protein END BINDING1 (EB1) highlighted increasing microtubule dynamicity during mitosis to support rapid changes in microtubule structures. The localization of the cell-plate specific syntaxin KNOLLE, several RAB-GTPases, as well as two plasma membrane localized proteins was assessed after treatment with the cytokinesis-specific callose-deposition inhibitor Endosidin7 (ES7) and the microtubule-disrupting herbicide chlorpropham (CIPC). While ES7 caused cell plate defects in Arabidopsis thaliana, it did not alter callose accumulation, or disrupt cell plate formation in maize. In contrast, CIPC treatment of maize epidermal cells occasionally produced irregular cell plates that split or fragmented, but did not otherwise disrupt the accumulation of cell-plate localized proteins. DISCUSSION: Together, these markers provide a robust suite of tools to examine subcellular trafficking and organellar organization during mitosis and cell plate formation in maize.
              Cover page: Subcellular positioning during cell division and cell plate formation in maize.
              Top