Functional Characterization of the Wheat Macrophage Migration Inhibitory Factor TaMIF1 in Wheat-Stripe Rust (Puccinia striiformis) Interaction

Macrophage migration inhibitory factor (MIF), named for its role in inhibiting macrophage/monocyte migration, has multiple functions in modulation of inflammation, cell proliferation, angiogenesis, and tumorigenesis in vertebrates. Although homologs of this gene can be found in plants, the function of MIF in plants remains obscure. Here, we characterized TaMIF1 in Triticum aestivum resembling the MIF secreted from Homo sapiens. Transcript analysis revealed that TaMIF1 responded to stripe rust infection of wheat and was upregulated during the infection stage. TaMIF1 was localized to both the cytosol and nuclei in wheat mesophyll protoplast. Additionally, TaMIF1 possessed significant tautomerase activity, indicating conservation of MIFs across kingdoms. Agrobacterium tumefaciens infiltration assay demonstrated that TaMIF1 was capable of suppressing programmed cell death hinting its role in plant immunity. Heterologous expression of TaMIF1 increased fission yeast sensitivity to oxidative stress. Silencing TaMIF1 decreased the susceptibility of wheat to Pst seemingly through increasing reactive oxygen species accumulation. In conclusion, functions of the TaMIF1 were investigated in this study, which provides significant insight into understanding the role of MIFs across kingdoms.

Stochastic gene expression drives mesophyll protoplast regeneration

Cell pluripotency is fundamental to biology. It has long been known that differentiated somatic plant cells may reacquire pluripotency, but the underlying mechanism remains elusive. In many plant species, a single isolated mesophyll protoplast may regenerate into an entire plant, which is widely used in gene transformation. Here, we identified two transcription factors whose ectopic activation promotes protoplast regeneration. Furthermore, we found that their expression was induced by protoplast isolation but at a very low frequency. Using live imaging and single-cell transcriptomics, we show that isolating protoplasts induces enhanced expression variation at the genome level. Isolating protoplasts also leads to genome-wide increases in chromatin accessibility, which promotes stochastic activation of gene expression and enhances protoplast regeneration. We propose that transcriptome chaos with increased expression variability among cells creates a cellular-level evolutionary driver selecting for regenerating cells.

A cell death assay in barley and wheat protoplasts for identification and validation of matching pathogen AVR effector and plant NLR immune receptors

Background Plant disease resistance to host-adapted pathogens is often mediated by host nucleotide-binding and leucine-rich repeat (NLR) receptors that detect matching pathogen avirulence effectors (AVR) inside plant cells. AVR-triggered NLR activation is typically associated with a rapid host cell death at sites of attempted infection and this response constitutes a widely used surrogate for NLR activation. However, it is challenging to assess this cell death in cereal hosts. Results Here we quantify cell death upon NLR-mediated recognition of fungal pathogen AVRs in mesophyll leaf protoplasts of barley and wheat. We provide measurements for the recognition of the fungal AVRs AvrSr50 and AVRa1 by their respective cereal NLRs Sr50 and Mla1 upon overexpression of the AVR and NLR pairs in mesophyll protoplast of both, wheat and barley. Conclusions Our data demonstrate that the here described approach can be effectively used to detect and quantify death of wheat and barley cells induced by overexpression of NLR and AVR effectors or AVR effector candidate genes from diverse fungal pathogens within 24 h.

Phosphate Status Affects Phosphate Transporter Expression and Glyphosate Uptake and Transport in Grand Eucalyptus (Eucalyptus grandis)

Soluble phosphate availability is a major limiting factor for plant growth, development, and yield. To assure a constant phosphorous supply, plants employ both high- and low-affinity phosphate acquisition mechanisms. Glyphosate is an herbicide widely used throughout the world, and previous studies have suggested that it can be transported across the plasma membrane via phosphate transporters in herbaceous species. The effects of phosphate status on glyphosate uptake were investigated in the tree grand eucalyptus (Eucalyptus grandisW. Hill ex. Maid.). Eucalyptus grandis’s putative phosphate transporters showed differential gene expression in leaves and roots in response to phosphate limitation. Overall, the expression of high-affinity phosphate transporters increased in phosphate-limiting treatments, particularly in roots. More [14C]glyphosate was absorbed and translocated in phosphate-limiting plants compared with control plants grown in phosphate-replete treatments. In leaf mesophyll protoplast assays, rapid uptake of [14C]glyphosate into protoplasts was observed, and addition of phosphate to the assays competed with [14C]glyphosate uptake. These data indicate that phosphate transporters represent one mechanism of glyphosate uptake inE. grandis. These results have implications for best management practices for weed control and glyphosate application under phosphate application regimes.