Calcium coding mechanisms in plant cell growth and immunity

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Calcium (Ca) is a second messenger in all eukaryotes. Defects in Ca signaling cause numerous human diseases including Alzheimer’s disease, heart failure, metabolic diseases, immune disorders, neurodegenerative diseases, and cancer. Despite the importance and broad medical implications, Ca signaling mechanisms remain unclear. The challenging question concerns how Ca encodes specific information coming from different primary signals and translate them into distinct cellular responses. Coding and decoding the specificity of Ca signals remains a long-standing puzzle in the signal transduction field. The PI’s laboratory studies Ca coding and decoding mechanisms using Arabidopsis as a model system and has made breakthroughs in dissecting Ca- coding mechanisms, setting the stage for this application. The proposed studies seek to understand Ca-coding mechanisms in the contexts of pollen tube growth and innate immunity both of which involve cyclic nucleotide- gated channels (CNGCs) in Arabidopsis. The Specific Aim 1 will address the relationship between CNGC-based Ca oscillations and peptide signaling during pollen tube growth. PI’s lab identified two CNGC-type proteins and calmodulin (CaM) forming a Ca “oscillator” in pollen tube growth that also requires autocrine peptide hormones produced by pollen tube. The overarching hypothesis is that peptides bind to their receptors that in turn modulate Ca-oscillator channels. This will be tested through genetic analysis combined with single cell Ca imaging. The Specific Aim 2 will identify Ca transporters that work together with CNGCs in immunity signaling. The importance of Ca signaling has long been recognized in innate immunity for both animal and plant cells. PI’s lab identified a CNGC-type channel that generates cytoplasmic Ca spike in response to bacterial pathogens. Using genetic analysis in Arabidopsis and yeast genetic complementation models, Aim 2 will identify the transporters responsible for removing the Ca signal and study how they coordinate with CNGC-type channels to precisely shape the spatial and temporal dynamics of Ca codes. Specific Aim 3 seeks to understand the mechanisms for activation and inactivation of plant CNGCs. The CNGC-type channels function in both pollen tube and immunity models, but they consist of different subunits and their regulations by CaM are different too. Further, while animal CNGCs are activated by the cyclic nucleotides (cAMP/cGMP), the plant CNGCs in pollen tube and immunity models are insensitive to these nucleotides. The hypothesis is that plant CNGCs are regulated differently from animal counterparts and CaM-based regulation depends on subunit composition of the CNGCs. This hypothesis will be tested in Aim 3 using biochemical and electrophysiological approaches in both pollen tube and immunity model. Arabidopsis is an ideal model to address basic Ca signaling mechanisms, as it provides a plethora of genetic tools and an array of whole-organism and single-cell Ca signaling phenotypes in the genetic mutants. Completion of these aims will reveal new Ca coding mechanisms, contributing to the conceptual framework of Ca signaling highly relevant to human health.