Anion channel SLAH3 is a regulatory target of chitin receptor-associated kinase PBL27 in microbial stomatal closure
- The Sainsbury Laboratory, United Kingdom
- University of Wuerzburg, Germany
- Ludwig-Maximilian-University of Munich, Germany
- John Innes Centre, United Kingdom
- University of Tuebingen, Germany
Figures
Figure 1 with 2 supplements
The LYK5-CERK1-PBL27 receptor complex and SLAH3 are required for chitin-induced stomatal closure.
(A–C) Stomatal aperture measurements in mutants of all members of the LysM-RLK family (A), pbl27 (B), slac1 and slah3 (C). Mature leaf discs were soaked in opening buffer (10 mM MES, 50 mM KCl, pH …
-
Figure 1—source data 1
Source data for stomatal measurements shown in Figure 1 and Figure 1—figure supplements 1 and 2, and for Figure 4.
- https://doi.org/10.7554/eLife.44474.005
Figure 1—figure supplement 1
Functional complementation and guard cell expression of the LYK5-CERK1-PBL27 receptor complex.
Stomatal aperture measurements in transgenic cerk1/pCERK1::CERK1-eGFP, lyk5/pLYK5::LYK5-eGFP and pbl27-1/pPBL27::PBL27-eGFP. Mature leaf discs of six independent transgenic T1 lines were soaked in …
Figure 1—figure supplement 2
Current ejection of chitosan induces stomatal closure in Arabidopsis.
Chitosan was current ejected into the cell wall of guard cells in intact leaves via a microelectrode, by application of 1 nA positive current for 1 min. The tip of electrode was filled with 0.3 …
Figure 2 with 1 supplement
PBL27 interacts with and phosphorylates SLAH3.
(A) Confocal microscopy of N. benthamiana leaves transiently expressing the indicated split-YFP constructs. Representative images are shown. (B) Co-immunoprecipitation of PBL27 and SLAH3 transiently …
-
Figure 2—source data 1
Source data for BiFC images shown in Figure 2.
- https://doi.org/10.7554/eLife.44474.008
-
Figure 2—source data 2
Source data for co-IP blots images shown in Figure 2.
- https://doi.org/10.7554/eLife.44474.009
-
Figure 2—source data 3
Source data for blots on in vitro phosphorylation shown in Figure 2.
- https://doi.org/10.7554/eLife.44474.010
-
Figure 2—source data 4
Source data for blots on in vitro-in vivo phosphorylation shown in Figure 2.
- https://doi.org/10.7554/eLife.44474.011
-
Figure 2—source data 5
Source data for blots on in vivo phosphorylation shown in Figure 2.
- https://doi.org/10.7554/eLife.44474.012
Figure 2—figure supplement 1
Identification of SLAH3 tryptic phospho-peptides.
(A) Summary of tryptic phospho-peptides by LC-MS/MS analysis of incubated recombinant His-MBP-PBL27 with recombinant His-SLAH3-N or GST-SLAH3-C (GST-SLAH3-C-His) in in vitro kinase assays. …
Figure 3 with 3 supplements
S-type anion currents are activated by co-injection of SLAH3 and PBL27 in oocytes.
(A) Macroscopic currents of Xenopus oocytes expressing SLAH3 in the presence or absence of PBL27 in response to the standard voltage protocol. Currents were recorded in 30 mM nitrate-based buffers. …
-
Figure 3—source data 1
Source data for current measurements shown in Figure 3 and Figure 3—figure supplements 1 and 2.
- https://doi.org/10.7554/eLife.44474.017
Figure 3—figure supplement 1
Activation of SLAH3 and its close relative SLAC1 by different kinases.
(A) Instantaneous currents of Xenopus oocytes injected with WT SLAH3 alone, with CIPK23/CBL1 or PBL27, respectively. The effect of ABI1 on the SLAH3-derived was studied by co-expression of the ABI1 …
Figure 3—figure supplement 2
CERK1 enhances PBL27 activation of SLAH3.
(A) Steady-state currents (ISS) of oocytes co-injected with SLAH3 and PBL27 in the presence or absence of CERK1. Currents were recorded in 30 mM nitrate-based buffers (n ≥ 4 experiments, mean ± SD). …
Figure 3—figure supplement 3
Activation of SLAH3 wild type and the SLAH3 mutants S127A, S189A and S601A.
(A), (B) and (C) Instantaneous currents of Xenopus oocytes injected with WT SLAH3 or (A) S127A, (B) S189A or (C) S601A alone or together with PBL27, CIPK23/CBL1, CPK21DEF or SLAH1. (A) As PBL27 …
Figure 4 with 1 supplement
SLAH3 phospho-sites S127 and S189 are necessary for chitin-induced stomatal closure.
(A–D) Stomatal aperture measurements in transgenic slah3-1/35S::SLAH3−3 × FLAG wild type (A), S127A (B), S189A (C) and S601A (D) variants of SLAH3. Mature leaf discs of six independent transgenic T1 …
Figure 4—figure supplement 1
Functional complementation of SLAH3 transgenic lines.
RT-PCR analysis of T1 transgenic slah3-1/p35S::SLAH3-3xFLAG lines and respective T1 lines expressing the S127A, S189A and S601A SLAH3 variants. RNA was extracted from six independent transgenic …
Figure 5
SLAH3 mediates resistance to Botrytis cinerea in leaves.
(A) Lesion diameter measurements in slah3-1 mutants and transgenic slah3-1/35S::SLAH3−3 × FLAG wild type (WT), S127A, S189A and S601A variants of SLAH3. Each four mature leaves of 8 to 10 …
-
Figure 5—source data 1
Source data for Botrytis infection shown in Figure 5.
- https://doi.org/10.7554/eLife.44474.021
-
Figure 5—source data 2
Source data for ethylene measurements shown in Figure 5.
- https://doi.org/10.7554/eLife.44474.022
-
Figure 5—source data 3
Source data for ROS measurements shown in Figure 5.
- https://doi.org/10.7554/eLife.44474.023
Additional files
-
Transparent reporting form
- https://doi.org/10.7554/eLife.44474.024
