A phosphocode-based dichotomy of BAK1 function in plant signalling

Phosphocode-dependent functional dichotomy of a common co-receptor in plant signalling

Perraki A., DeFalco T. A., Derbyshire P. , Avila J., Séré D., Sklenar J., Qi X., Stransfeld L., Schwessingern B., Kadota Y., Macho A. P.,  Jiang S., Couto D., Torii K. U., Menke F. L. H., Cyril Zipfel C. (2018)

Artemis Perraki, Thomas A. DeFalco, Paul Derbyshire, Julian Avila, David Séré, Jan Sklenar, Xingyun Qi, Lena Stransfeld, Benjamin Schwessinger, Yasuhiro Kadota, Alberto P. Macho, Shushu Jiang, Daniel Couto, Keiko U. Torii, Frank L. H. Menke, Cyril Zipfel,

Nature 561: 248–252 – https://doi.org/10.1038/s41586-018-0471-x

Abstract

Multicellular organisms use cell-surface receptor kinases to sense and process extracellular signals. Many plant receptor kinases are activated by the formation of ligand-induced complexes with shape-complementary co-receptors¹. The best-characterized co-receptor is BRASSINOSTEROID INSENSITIVE 1-ASSOCIATED KINASE 1 (BAK1), which associates with numerous leucine-rich repeat receptor kinases (LRR-RKs) to control immunity, growth and development². Here we report key regulatory events that control the function of BAK1 and, more generally, LRR-RKs. Through a combination of phosphoproteomics and targeted mutagenesis, we identified conserved phosphosites that are required for the immune function of BAK1 in Arabidopsis thaliana. Notably, these phosphosites are not required for BAK1-dependent brassinosteroid-regulated growth. In addition to revealing a critical role for the phosphorylation of the BAK1 C-terminal tail, we identified a conserved tyrosine phosphosite that may be required for the function of the majority of Arabidopsis LRR-RKs, and which separates them into two distinct functional classes based on the presence or absence of this tyrosine. Our results suggest a phosphocode-based dichotomy of BAK1 function in plant signalling, and provide insights into receptor kinase activation that have broad implications for our understanding of how plants respond to their changing environment.

Multiple related yet unique peptides specify stomatal cell fate

Effects of BFA on subcellular membrane structures in the meristemoids.
(A) Representative Z-stack confocal images of an ER marker, GFP-HDEL (green) and FM4-64 (magenta) in true leaf abaxial epidermis from the 7-day-old seedlings expressing CaMV35Spro::GFP-HDEL treated with 5 µM FM4-64 alone (mock) or co-treated with 30, 90, 180 µM BFA for 1 hr. Scale bars = 20 µm. BFA treatment at low concentration (30 µM) does not alter the characteristic, mesh-like ER structure inside the meristemoids and at the edge of nuclei. In contrast, high concentration of BFA results in aberrant spherical structures in the ER. (B) Representative confocal images of an ER marker, GFP-HDEL (green) and FM4-64 (magenta) treated with BFA as described in (A). BFA treatment at low concentration (30 µM) causes the formation of BFA bodies (magenta arrows) without impacting the ER structure. In high BFA concentration confers aberrant spherical ER structure (green arrows). In addition, the FM4-64 signals in the BFA bodies disappear. Scale bars = 5 µm. (C) Representative confocal images of a Golgi marker, ST-YFP (green) and FM4-64 (magenta) in true leaf abaxial epidermis from the 7-day-old seedlings expressing CaMV35Spro::N-ST-YFP treated with 5 µM FM4-64 alone (mock) or co-treated with 30, 90, 180 µM BFA for 1 hr. Scale bars = 5 µm. In a lower and medium concentration (30 and 90 µM) of BFA, Golgi (green arrows) are surrounding BFA bodies (magenta arrows). By contrast, in higher concentration (180 µM), the Golgi marker becomes collapsed (green arrow) to ER. (D) Schematic diagrams depicting probable subcellular membrane structures in the meristemoids based on confocal microscopy.

The manifold actions of signaling peptides on subcellular dynamics of a receptor specify stomatal cell fate

Qi X., Yoshinari A., Bai P., Maes M., Zeng S. M., Torii K. U. (2020)

Xingyun Qi, Akira Yoshinari, Pengfei Bai, Michal Maes, Scott M Zeng, Keiko U Torii,

===

Plant Biology – DOI: 10.7554/eLife.58097 –

https://elifesciences.org/articles/58097

Abstract

Receptor endocytosis is important for signal activation, transduction, and deactivation. However, how a receptor interprets conflicting signals to adjust cellular output is not clearly understood. Using genetic, cell biological, and pharmacological approaches, we report here that ERECTA-LIKE1 (ERL1), the major receptor restricting plant stomatal differentiation, undergoes dynamic subcellular behaviors in response to different EPIDERMAL PATTERNING FACTOR (EPF) peptides. Activation of ERL1 by EPF1 induces rapid ERL1 internalization via multivesicular bodies/late endosomes to vacuolar degradation, whereas ERL1 constitutively internalizes in the absence of EPF1. The co-receptor, TOO MANY MOUTHS is essential for ERL1 internalization induced by EPF1 but not by EPFL6. The peptide antagonist, Stomagen, triggers retention of ERL1 in the endoplasmic reticulum, likely coupled with reduced endocytosis. In contrast, the dominant-negative ERL1 remained dysfunctional in ligand-induced subcellular trafficking. Our study elucidates that multiple related yet unique peptides specify cell fate by deploying the differential subcellular dynamics of a single receptor.

Self-inhibition as a mechanism for ensuring proper stomatal development

Expression patterns of ERL1 during stomatal development.
Expression pattern of ERL1pro::ERL1-YFP. (A–J) Time-lapse live imaging of developing abaxial cotyledon epidermis of the 1-day-old T4 seedling of ERL1pro::ERL1-YFP erl1-2. Time points after image collection are indicated in hours:(A) 0.0 hr; (B) 3.5 hr; (C) 13.0 hr; (D) 16.0 hr; (E) 24.5 hr; (F) 26.0 hr; (G) 36.0 hr; (H) 50.5 hr; (I) 68.0 hr; and (J) 71.0 hr. Arrowheads point to two representative cells. Images for (A–J) are taken at the same magnification. Scale bar, 10 µm. (K–L) High resolution live images of stomatal precursors expressing ERL1-YFP. (K, L) meristemoid mother cells (MMC); (M) meristemoid (M); (N) late meristemoid (late M); (O) late meristemoid to guard mother cell transition (late M~GMC); (P) GMC; (Q) immature guard cells (im GC). Images for (K–L) are taken at the same magnification. Scale bar, 7.5 µm. ERL1-YFP signals are detected at the plasma membrane from early meristemoids (A, K, L; arrowheads) and accumulate high at the asymmetric division site (e.g. D, F, M; cyan asterisks). ERL1-YFP signals boost during the late meristemoid-to-GMC transition (G, H, N, O; cyan plus), and diminishes after GMC symmetric division (I, J, P, Q; arrowheads). See accompanying Video 1. Experiments were repeated three times. Total seedlings analyzed; n = 9.

Autocrine regulation of stomatal differentiation potential by EPF1 and ERECTA-LIKE1 ligand-receptor signaling

Qi X., Han S.-K., Dang J. H., Garrick J. M., Ito M., Hofstetter A. K., Torii K. U. (2017)

Xingyun Qi, Soon-Ki Han, Jonathan H. Dang, Jacqueline M. Garrick, Masaki Ito, Alex K. Hofstetter, Keiko U. Torii,

===

In Plant Biology, Developmental Biology – https://doi.org/10.7554/eLife.24102.001 –

https://elifesciences.org/articles/24102

Absolute co-expression of ERL1 and MUTE confers meristemoid arrests.
Shown are confocal microscopy images of cotyledon abaxial epidermis from 7-day-old seedlings of the following genotypes: (A) er erl1 erl2; (B) MUTEpro::ERL1-YFP in er erl1 erl2; (D) er erl1 erl2 epf1 epf2; (E) MUTEpro::ERL1-YFP in er erl1 erl2 epf1 epf2; (F) MUTEpro::ERL1-YFP in er erl1 erl2 mock treated; (G) MUTEpro::ERL1-YFP in er erl1 erl2 treated with 5 µM Stomagen peptide; (H) er erl1 erl2 tmm; (I) MUTEpro::ERL1-YFP in er erl1 erl2 tmm. T1 transgenic seedlings of MUTEpro::ERL1-YFP er erl1 erl2; MUTEpro::ERL1-YFP er erl1 erl2 epf1 epf2; and MUTEpro::ERL1-YFP er erl1 erl2 tmm were used for the analysis. T2 seedlings of MUTEpro::ERL1-YFP er erl1 erl2 were used for the mock or Stomagen treatment. Scale bars, 10 µm (A, B, D, E, H, I), 25 µm (F, G). (C) Quantitative analysis. Stomatal index (SI) of the cotyledon abaxial epidermis from 7-day-old seedlings of respective genotypes. For each genotype, images from six seedlings were analyzed. Welch’s Two Sample T-test was performed for mock vs. Stomagen application (Left). One-way ANOVA followed by Tukey’s HSD test was performed for comparing all other genotypes and classify their phenotypes into three categories (a, b, and c).

Abstract

Development of stomata, valves on the plant epidermis for optimal gas exchange and water control, is fine-tuned by multiple signaling peptides with unique, overlapping, or antagonistic activities. EPIDERMAL PATTERNING FACTOR1 (EPF1) is a founding member of the secreted peptide ligands enforcing stomatal patterning. Yet, its exact role remains unclear. Here, we report that EPF1 and its primary receptor ERECTA-LIKE1 (ERL1) target MUTE, a transcription factor specifying the proliferation-to-differentiation switch within the stomatal cell lineages. In turn, MUTE directly induces ERL1. The absolute co-expression of ERL1 and MUTE, with the co-presence of EPF1, triggers autocrine inhibition of stomatal fate. During normal stomatal development, this autocrine inhibition prevents extra symmetric divisions of stomatal precursors likely owing to excessive MUTE activity. Our study reveals the unexpected role of self-inhibition as a mechanism for ensuring proper stomatal development and suggests an intricate signal buffering mechanism underlying plant tissue patterning.

Signals guiding stomatal development

Fig. 1. Summary of the effects that diverse range of signals have on stomatal development. a A cartoon showing stomatal cell-lineage transitions from a protodermal cell, a meristemoid mother cell (MMC), meristemoids undergoing asymmetric amplifying divisions and producing stomatal-lineage ground cells (SLGCs), and a guard mother cell (GMC) to a stoma with paired guard cells (GCs). A protodermal cell could differentiate into a pavement cell, and SLGCs could become pavement cells. Cartoons are modified from Han and Torii [11]. b An Arabidopsis seedling with stomata highlighted in green is in the center. Signals that negatively regulate stomatal development are shown on the left, indicated with red arrows. Signals that promote stomatal formation are shown on the right, indicated with green arrows. The black and yellow boxes indicate darkness (or signals that inhibit stomatal development) and light (or signals that promote stomatal development), respectively. When a signal is deficient, a minus sign is put in front of it. Top left: cotyledon epidermis with pavement cell only. Middle left: cotyledon epidermis with arrested meristemoids. Bottom left: hypocotyl epidermis with pavement cell only. Top right: cotyledon epidermis with clustered stomata. Middle right: cotyledon epidermis with high stomatal density. Bottom right: hypocotyl epidermis with clustered stomata. Confocal microscopy images of the cotyledon and hypocotyl epidermis of wild-type and various mutant seedlings were taken using a Leica SP5 WLL and false colored using Adobe Photoshop CS6

 

Hormonal and environmental signals guiding stomatal development

by Qi X., Torii K. U. (2018)

 

• Xingyun Qi,
• Keiko U. Torii, Howard Hughes Medical Institute and Department of Biology, University of Washington, Seattle, WA 98195, USA

 

===

in BMC Biology 16: 21 – https://doi.org/10.1186/s12915-018-0488-5 –

https://bmcbiol.biomedcentral.com/articles/10.1186/s12915-018-0488-5

Fig. 2. How hormonal and environmental cues are integrated into the core stomatal developmental pathway. Components involved in the same pathway are grouped with the same color. The experimentally confirmed steps are shown as solid lines, and the steps that are uncertain are shown as broken lines. An arrow indicates a positive regulation, while a ‘T’ indicates negative regulation

Abstract

Stomata are pores on plant epidermis that facilitate gas exchange and water evaporation between plants and the environment.

Given the central role of stomata in photosynthesis and water-use efficiency, two vital events for plant growth, stomatal development is tightly controlled by a diverse range of signals.

A family of peptide hormones regulates stomatal patterning and differentiation. In addition, plant hormones as well as numerous environmental cues influence the decision of whether to make stomata or not in distinct and complex manners.

In this review, we summarize recent findings that reveal the mechanism of these three groups of signals in controlling stomatal formation, and discuss how these signals are integrated into the core stomatal development pathway.

MUTE, Cell-State Switch and the Single Symmetric Division to Create Stomata

Transcriptomic Profiling of MUTE Target Genes Reveals a Framework of Stomatal Cell-State Switch

(A–E) Epidermal phenotypes of 3-day-old seedlings. Mock (A and C), iMUTE (B and D), and iSPCH (E). Mature stomata of mock (C) and iMUTE (D) cotyledon epidermis expressing GC GFP marker E994. Scale bars, 50 μm.

MUTE Directly Orchestrates Cell-State Switch and the Single Symmetric Division to Create Stomata

by Han S. K., Qi X., Sugihara K., Dang J. D., Endo T. A., Miller K. L., Kim E.-D., Miura T., Torii K. U. (2018)

Soon-Ki Han⁶ Xingyun Qi⁶ Kei Sugihara Jonathan H. Dang Takaho A. Endo Kristen L. Miller Eun-Deok Kim Takashi Miura Keiko U. Torii⁷

===

in Dev. Cell 45(3): 303-315 –

https://www.cell.com/developmental-cell/fulltext/S1534-5807(18)30285-5 

Highlights

• •

  Comprehensive inventories of gene expression in stomatal differentiation reported

• •

  MUTE switches stomatal patterning program initiated by its sister bHLH, SPEECHLESS

• •

  MUTE directly induces cell-cycle genes and their direct transcriptional repressors

• •

  Incoherent feed-forward loop by MUTE ensures stomata composed of paired guard cells

Summary

Precise cell division control is critical for developmental patterning. For the differentiation of a functional stoma, a cellular valve for efficient gas exchange, the single symmetric division of an immediate precursor is absolutely essential. Yet, the mechanism governing this event remains unclear. Here we report comprehensive inventories of gene expression by the Arabidopsis bHLH protein MUTE, a potent inducer of stomatal differentiation.

MUTE switches the gene expression program initiated by SPEECHLESS. MUTE directly induces a suite of cell-cycle genes, including CYCD5;1, in which introduced expression triggers the symmetric divisions of arrested precursor cells in mute, and their transcriptional repressors, FAMA and FOUR LIPS.

The regulatory network initiated by MUTE represents an incoherent type 1 feed-forward loop. Our mathematical modeling and experimental perturbations support a notion that MUTE orchestrates a transcriptional cascade leading to a tightly restricted pulse of cell-cycle gene expression, thereby ensuring the single cell division to create functional stomata.

Autocrine regulation of stomatal differentiation

 

 

 

Autocrine regulation of stomatal differentiation potential by EPF1 and ERECTA-LIKE1 ligand-receptor signaling

Qi X., Han S.-K., Dang J. H., Garrick J. M., Ito M., Hofstetter A. K., Torii K. U. (2017)

1. Xingyun Qi,
2. Soon-Ki Han,
3. Jonathan H Dang,
4. Jacqueline M Garrick,
5. Masaki Ito,
6. Alex K Hofstetter,
7. Keiko U Torii, 

 

in  eLife 2017;6:e24102 DOI: 10.7554/eLife.24102

https://elifesciences.org/articles/24102

Abstract

 

Development of stomata, valves on the plant epidermis for optimal gas exchange and water control, is fine-tuned by multiple signaling peptides with unique, overlapping, or antagonistic activities.

EPIDERMAL PATTERNING FACTOR1 (EPF1) is a founding member of the secreted peptide ligands enforcing stomatal patterning. Yet, its exact role remains unclear.

Here, we report that EPF1 and its primary receptor ERECTA-LIKE1 (ERL1) target MUTE, a transcription factor specifying the proliferation-to-differentiation switch within the stomatal cell lineages.

In turn, MUTE directly induces ERL1.

The absolute co-expression of ERL1 and MUTE, with the co-presence of EPF1, triggers autocrine inhibition of stomatal fate.

During normal stomatal development, this autocrine inhibition prevents extra symmetric divisions of stomatal precursors likely owing to excessive MUTE activity.

Our study reveals the unexpected role of self-inhibition as a mechanism for ensuring proper stomatal development and suggests an intricate signal buffering mechanism underlying plant tissue patterning.