The plant immune system is a multi-phase complex network that involves the collaboration of multiple subcellular structures. In the past two decades, the core signaling pathways of the immune process, including pattern-triggered immunity (PTI), effector-triggered immunity (ETI), and systemic acquired resistance (SAR), as well as the behavior of organelles, have been revealed to a level of clarity that is able to describe a general and well-covered process of the immune response. However, there are still many events during the immune response that remain mysterious. For instance, while higher plants live a sessile lifestyle, there are countless intracellular motions mediated by the cytoskeleton (including its associated proteins) in response to the external triggers, such as the invasion of pathogens. As our knowledge of plant immunity accumulates, the deficiency in knowledge on how immune signaling regulates the behavior of the cytoskeleton as a critical aspect of defense response, howbeit, becomes more evident. Therefore, this is a field of research that calls for powerful toolboxes to facilitate the analysis of the cytoskeleton in the context of immunity, as well as instructive biological model(s) that guide the direction of the multifarious studies. In this dissertation, I focus on the summary and prospective discussion on the immune function of the actin cytoskeleton and, more importantly, describe my original studies on two major aspects of this topic. First, a prerequisite to functional study of the actin cytoskeleton in the cytoplasm is the ability to accurately describe the status of the cytoskeleton. To achieve this goal, I developed an algorithm, namely implicit Laplacian of enhanced edge (ILEE), to accurately identify and analyze the biological status of the cytoskeleton from confocal image samples. This method significantly improves the accuracy, stability, and robustness of cytoskeleton segmentation, solves other technical hindrances, and enables abundant information to be extracted from images for biological interpretation (see Chapter 2). The ILEE algorithm will further help me to explore the phenotypes of actin architecture in response to immune signaling, which was not previously available due to the lack of the toolbox. Also, the ILEE has been packaged as a library released publicly to benefit the community with a powerful cytoskeleton analysis platform.For the second project of my total research, I focused on the immune function of the actin cytoskeleton in the nucleus. Previously, some Arabidopsis actin depolymerization factors were reported to genetically contribute to plant immunity by unknown mechanism(s), and my story began with a novel activity identified among Arabidopsis actin depolymerization factors – to interact with WRKYs, the stress-responsive transcription factors. During my research, I proved that certain ADFs can form a complex with WRKYs that binds to targeted promoters, hence regulating the activity of WRKYs and playing a positive role in the immune response. The knowledge obtained through this study, in combination with previous research (Lu et al., 2020; Porter et al., 2012a) of my lab, can be summarized into a biological model, in which ADF mediates a nuclear-cytoplasmic immune regulation that systemically facilitates both cytoskeleton dynamics and pro-immune transcriptome reprogramming. In general, this study reveals a novel yet general pattern of cytoskeleton mediated transcriptional regulation, as ADF and perhaps other components of the actin cytoskeleton can shuttle between the cytoplasm and nucleus to form a network with a higher level of complexity. As a potential broader impact, the application range of this model includes but is not necessarily limited to plant immunity.
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