identification-genes-involved-stomatal-development-via-epidermal

Identification of the Genes Involved in Stomatal Development <em>via</em> Epidermal Phenotype Scoring

Stomata are small pores on the surface of land plants that are involved in gas exchange and water vapor release, and their function is critical for plant ...

Concordia University

Peptide ligand-receptor kinase interactions have emerged as a key component of plant growth and development. Now, highly related small signaling peptides have been shown to act antagonistically on the same receptor kinase, providing new insights into how plants optimize developmental processes using competitive peptides.

Fine-Tuning Development Through Antagonistic Peptides: An Emerging Theme.

Receptor kinases play a central role in various biological processes, but due to their low abundance and highly hydrophobic and dynamic nature, only a few of them have been functionally characterized, and their partners and ligands remain unidentified. Receptor protein extraction and purification from plant tissues is one of the most challenging steps for the success of various biochemical analyses to characterize their function. Immunoprecipitation is a widely used and selective method for enriching or purifying a specific protein. Here we describe two different optimized protein purification protocols, batch and on-chip immunoprecipitation, which efficiently isolate plant membrane receptor kinases for functional analysis.

Purification of Plant Receptor Kinases from Plant Plasma Membranes.

Mitogen-activated protein kinase (MAPK) signaling plays important roles in diverse biological processes. In , MPK3/MPK6, MKK4/MKK5, and the MAPKKK YODA (YDA) form a MAPK pathway that negatively regulates stomatal development. Brassinosteroid (BR) stimulates this pathway to inhibit stomata production. In addition, MPK3/MPK6 and MKK4/MKK5 also serve as critical signaling components in plant immunity. Here, we report that MAPKKK3/MAPKKK5 form a kinase cascade with MKK4/MKK5 and MPK3/MPK6 to transduce defense signals downstream of multiple plant receptor kinases. Loss of MAPKKK3/MAPKKK5 leads to reduced activation of MPK3/MPK6 in response to different pathogen-associated molecular patterns (PAMPs) and increased susceptibility to pathogens. Surprisingly, developmental defects caused by silencing of  are suppressed in the  double mutant. On the other hand, loss of YDA or blocking BR signaling leads to increased PAMP-induced activation of MPK3/MPK6. These results reveal antagonistic interactions between a developmental MAPK pathway and an immune signaling MAPK pathway.

Antagonistic interactions between two MAP kinase cascades in plant development and immune signaling.

Stomata on the plant epidermis control gas and water exchange and are formed by MAPK-dependent processes. Although the contribution of MAP KINASE3 (MPK3) and MPK6 (MPK3/MPK6) to the control of stomatal patterning and differentiation in Arabidopsis () has been examined extensively, how they are inactivated and regulate distinct stages of stomatal development is unknown. Here, we identify a dual-specificity phosphatase, MAP KINASE PHOSPHATASE1 (MKP1), which promotes stomatal cell fate transition by controlling MAPK activation at the early stage of stomatal development. Loss of function of  creates clusters of small cells that fail to differentiate into stomata, resulting in the formation of patches of pavement cells. We show that MKP1 acts downstream of YODA (a MAPK kinase kinase) but upstream of MPK3/MPK6 in the stomatal signaling pathway and that MKP1 deficiency causes stomatal signal-induced MAPK hyperactivation in vivo. By expressing MKP1 in the three discrete cell types of stomatal lineage, we further identified that MKP1-mediated deactivation of MAPKs in early stomatal precursor cells directs cell fate transition leading to stomatal differentiation. Together, our data reveal the important role of MKP1 in controlling MAPK signaling specificity and cell fate decision during stomatal development.

MAP KINASE PHOSPHATASE1 Controls Cell Fate Transition during Stomatal Development.

The MAPK signaling cascade is universal among eukaryotes and mediates a variety of environmental and developmental responses. Two Arabidopsis MAPKs, MPK3 and MPK6, have been shown to be activated by various stimuli and suggested as a convergence point of different signaling pathways. It is known that these MAPKs, MPK3/MPK6, control the discrete stages of stomatal development in Arabidopsis, but how they are regulated and how the same MAPK components can achieve signaling specificity is largely unknown. We recently demonstrated that MAP Kinase Phosphatase 1 (MKP1) promotes stomatal differentiation by suppressing activation of MPK3/MPK6 in the stomatal lineage. By expressing MKP1 in discrete stomatal precursor cell types, we further identified that MKP1 plays an important role at the early stage of stomatal development for the cell fate transition leading to stomatal differentiation. While MKP1 was previously known as a key regulator of environmental stress responses, our data illustrate a novel role of MKP1 in plant development: it acts as one of the specificity-determining regulators of MAPK signaling to enforce proper stomatal development in Arabidopsis.

MKP1 acts as a key modulator of stomatal development.

Peptide signaling has emerged as a key component of plant growth and development, including stomatal patterning, which is crucial for plant productivity and survival. Although exciting progress has been made in understanding EPIDERMAL PATTERNING FACTOR (EPF) signaling in Arabidopsis, the mechanisms by which EPF peptides control different stomatal patterns and morphologies in grasses are poorly understood. Here, by examining expression patterns, overexpression transgenics and cross-species complementation, the antagonistic stomatal ligands orthologous to Arabidopsis AtEPF2 and AtSTOMAGEN/AtEPFL9 peptides were identified in Triticum aestivum (wheat) and the grass model organism Brachypodium distachyon. Application of bioactive BdEPF2 peptides inhibited stomatal initiation, but not the progression or differentiation of stomatal precursors in Brachypodium. Additionally, the inhibitory roles of these EPF peptides during grass stomatal development were suppressed by the contrasting positive action of the BdSTOMAGEN peptide in a dose-dependent manner. These results not only demonstrate how conserved EPF peptides that control different stomatal patterns exist in nature, but also suggest new strategies to improve crop yield through the use of plant-derived antagonistic peptides that optimize stomatal density on the plant epidermis.

Jin Suk Lee

Department of Biology

Concordia University

Identification of the Genes Involved in Stomatal Development via Epidermal Phenotype Scoring

Jin Suk Lee

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Biology

Concordia University

Identification of the Genes Involved in Stomatal Development via Epidermal Phenotype Scoring

Department of Biology