Evolutionary characteristics, expression patterns of wheat receptor-like kinases and functional analysis of TaCrRLK1L16

Abstract

Reverse genetics research in complex hexaploid wheat often encounters challenges in determining the priority of gene functional characterization. This study aims to systematically analyze the wheat (Triticum aestivum) receptor-like kinase (TaRLK) gene family and develop an effective strategy to identify key candidate genes for further investigation. We identified 3,424 TaRLKs using bioinformatics methods and analyzed the diverse and conserved evolutionary relationships of RLKs among Arabidopsis, rice and wheat. Based on publicly available and our laboratory’s transcriptome data, we comprehensively analyzed the transcriptional expression patterns of TaRLKs in response to various stresses, particularly Puccinia striiformis f. sp. tritici (Pst). The TaCrRLK1L16, which is upregulated during Pst infection and triggered cell death in Nicotiana benthamiana, has been identified as a key candidate gene for further functional characterization. Furthermore, our results suggested that the transgenic wheat overexpressing TaCrRLK1L16 significantly enhanced resistance to Pst. This study will provide valuable insights into understanding the evolutionary characteristics and expression patterns of TaRLKs while offering a novel strategy for determining the priority of key candidate TaRLKs.

Keywords

Wheat / Receptor-like kinase / Puccinia striiformis f. sp. tritici / Plant immunity / Biological Sciences / Genetics

References

[1] Almagro ArmenterosJJ, TsirigosKD, SønderbyCK, et al. . SignalP 5.0 improves signal peptide predictions using deep neural networks. Nat Biotechnol, 2019, 37: 420-423.

[2] AusubelFM. Are innate immune signaling pathways in plants and animals conserved?. Nat Immunol, 2005, 6: 973-979.

[3] BaiX, ZhanG, TianS, et al. . Transcription factor BZR2 activates chitinase Cht20.2 transcription to confer resistance to wheat stripe rust. Plant Physiol, 2021, 187: 2749-2762.

[4] BorrillP, Ramirez-GonzalezR, UauyC. expVIP: A customizable RNA-seq data analysis and visualization platform. Plant Physiol, 2016, 170: 2172-2186.

[5] ChenC, WuY, LiJ, et al. . TBtools-II: A “one for all, all for one” bioinformatics platform for biological big-data mining. Mol Plant, 2023, 16: 1733-1742.

[6] CheungAY, WuH-M. THESEUS 1, FERONIA and relatives: a family of cell wall-sensing receptor kinases?. Curr Opin Plant Biol, 2011, 14: 632-641.

[7] ChisholmST, CoakerG, DayB, StaskawiczBJ. Host-microbe interactions: shaping the evolution of the plant immune response. Cell, 2006, 124: 803-814.

[8] DanglJL, JonesJDG. Plant pathogens and integrated defence responses to infection. Nature, 2001, 411: 826-833.

[9] DievartA, GottinC, PérinC, et al. . Origin and diversity of plant receptor-like kinases. Annu Rev Plant Biol, 2020, 71: 131-156.

[10] DouD, ZhouJ-M. Phytopathogen effectors subverting host immunity: different foes, similar battleground. Cell Host Microbe, 2012, 12: 484-495.

[11] EmmsDM, KellyS. OrthoFinder: phylogenetic orthology inference for comparative genomics. Genome Biol, 2019, 20: 238.

[12] FanJ, BaiP, NingY, et al. . The monocot-specific receptor-like kinase SDS2 controls cell death and immunity in rice. Cell Host Microbe, 2018, 23: 498-510.e5.

[13] GanP, TangC, LuY, et al. . Quantitative phosphoproteomics reveals molecular pathway network in wheat resistance to stripe rust. Stress Biol, 2024, 4: 32.

[14] GaoC-H, YuG, CaiP. ggVennDiagram: An intuitive, easy-to-use, and highly customizable r package to generate venn diagram. Front Genet, 2021, 12. 706907

[15] GasteigerE, HooglandC, GattikerAWalkerJM. Protein identification and analysis tools on the ExPASy server. The Proteomics Protocols Handbook, 2005TotowaHumana Press571-607.

[16] Gómez-GómezL, BollerT. FLS2: An LRR receptor–like kinase involved in the perception of the bacterial elicitor flagellin in Arabidopsis. Mol Cell, 2000, 5: 1003-1011.

[17] GuJ, SunJ, LiuN, et al. . A novel cysteine-rich receptor-like kinase gene, TaCRK2 , contributes to leaf rust resistance in wheat. Mol Plant Pathol, 2020, 21: 732-746.

[18] Guo J, Islam MA, Lin H, Ji C, Duan Y, Liu P, Zeng Q, Day B, Kang Z, Guo J (2018) Genome-wide identification of cyclic nucleotide-gated ion channel gene family in wheat and functional analyses of TaCNGC14 and TaCNGC16. Front Plant Sci 9:18. https://doi.org/10.3389/fpls.2018.00018

[19] HaytaS, SmedleyMA, DemirSU, et al. . An efficient and reproducible Agrobacterium-mediated transformation method for hexaploid wheat (Triticum aestivum L.). Plant Methods, 2019, 15: 121.

[20] HooperCM, CastledenIR, AryamaneshN, et al. . Finding the subcellular location of barley, wheat, rice and maize proteins: the compendium of crop proteins with annotated locations (cropPAL). Plant Cell Physiol, 2015, 57: e9-e9.

[21] HuangX, LiuY, JiaY, et al. . FERONIA homologs in stress responses of horticultural plants: current knowledge and missing links. Stress Biol, 2024, 4: 28.

[22] JonesJDG, DanglJL. The plant immune system. Nature, 2006, 444: 323-329.

[23] ImaiR, KoikeM, SutohK, et al. . Molecular characterization of a cold-induced plasma membrane protein gene from wheat. Mol Genet Genomics, 2005, 274: 445-453.

[24] KadotaY, SklenarJ, DerbyshireP, et al. . Direct regulation of the NADPH oxidase RBOHD by the PRR-associated kinase BIK1 during plant immunity. Mol Cell, 2014, 54: 43-55.

[25] KatohK, StandleyDM. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol, 2013, 30: 772-780.

[26] KroghA, LarssonB, von HeijneG, SonnhammerELL. Predicting transmembrane protein topology with a hidden markov model: application to complete genomes. J Mol Biol, 2001, 305: 567-580.

[27] LetunicI, BorkP. Interactive tree of life (iTOL) v5: An online tool for phylogenetic tree display and annotation. Nucleic Acids Res, 2021, 49: W293-W296.

[28] LetunicI, KhedkarS, BorkP. SMART: recent updates, new developments and status in 2020. Nucleic Acids Res, 2020, 49: D458-D460.

[29] LiangX, ZhangJ. Regulation of plant responses to biotic and abiotic stress by receptor-like cytoplasmic kinases. Stress Biol, 2022, 2: 25.

[30] LiangX, ZhouJ-M. Receptor-Like Cytoplasmic Kinases: Central players in plant receptor kinase-mediated signaling. Annu Rev Plant Biol, 2018, 69: 267-299.

[31] LiuM-CJ, YehF-LJ, YvonR, et al. . Extracellular pectin-RALF phase separation mediates FERONIA global signaling function. Cell, 2024, 187: 312-330.e22.

[32] LiuQ, FuQ, YanY, et al. . Curation, nomenclature, and topological classification of receptor-like kinases from 528 plant species for novel domain discovery and functional inference. Mol Plant, 2024, 17: 658-671.

[33] LuS, WangJ, ChitsazF, et al. . CDD/SPARCLE: The conserved domain database in 2020. Nucleic Acids Res, 2019, 48: D265-D268.

[34] MachoA, WangP, ZhuJ-K. Modification of the susceptibility gene TaPsIPK1 - a win-win for wheat disease resistance and yield. Stress Biol, 2022, 2: 40.

[35] MinhBQ, SchmidtHA, ChernomorO, et al. . IQ-TREE 2: new models and efficient methods for phylogenetic inference in the genomic era. Mol Biol Evol, 2020, 37: 1530-1534.

[36] MistryJ, ChuguranskyS, WilliamsL, et al. . Pfam: The protein families database in 2021. Nucleic Acids Res, 2020, 49: D412-D419.

[37] NgouBPM, DingP, JonesJDG. Thirty years of resistance: Zig-zag through the plant immune system. Plant Cell, 2022, 34: 1447-1478.

[38] NguyenQ-N, LeeY-S, ChoL-H, et al. . Genome-wide identification and analysis of Catharanthus roseus RLK1-like kinases in rice. Planta, 2014, 241: 603-613.

[39] PetutschnigEK, JonesAME, SerazetdinovaL, et al. . The lysin motif receptor-like kinase (LysM-RLK) CERK1 is a major chitin-binding protein in Arabidopsis thaliana and subject to Chitin-induced phosphorylation. J Biol Chem, 2010, 285: 28902-28911.

[40] PitsiliE, PhukanUJ, CollNS. Cell death in plant immunity. Cold Spring Harb Perspect Biol, 2020, 12: a036483.

[41] PriceMN, DehalPS, ArkinAP. FastTree 2 – approximately maximum-likelihood trees for large alignments. PLoS ONE, 2010, 5: e9490.

[42] Raivo K (2010) pheatmap: Pretty Heatmaps. 1.0.12. https://doi.org/10.32614/CRAN.package.pheatmap

[43] Ramírez-GonzálezRH, BorrillP, LangD, et al. . The transcriptional landscape of polyploid wheat. Science, 2018, 361: eaar6089.

[44] Rao S, Zhou Z, Miao P, Bi G, Hu M, Wu Y, Feng F, Zhang X, Zhou JM (2018) Roles of receptor-like cytoplasmic kinase VII members in pattern-triggered immune signaling. Plant Physiol pp.00486.2018. https://doi.org/10.1104/pp.18.00486

[45] SaintenacC, CambonF, AouiniL, et al. . A wheat cysteine-rich receptor-like kinase confers broad-spectrum resistance against Septoria tritici blotch. Nat Commun, 2021, 12: 433.

[46] SchlessingerJ. Receptor tyrosine kinases: legacy of the first two decades. Cold Spring Harb Perspect Biol, 2014, 6: a008912-a008912.

[47] ShiY, BaoX, SongX, et al. . The leucine-rich repeat receptor-like kinase protein TaSERK1 positively regulates high-temperature seedling plant resistance to Puccinia striiformis f. sp. tritici by interacting with TaDJA7. Phytopathology, 2023, 113: 1325-1334.

[48] ShiuS-H, BleeckerAB. Plant receptor-like kinase gene family: diversity, function, and signaling. Sci STKE, 2001, 2001: re22.

[49] ShiuS-H, KarlowskiWM, PanR, et al. . Comparative analysis of the receptor-like kinase family in Arabidopsis and Rice. Plant Cell, 2004, 16: 1220-1234.

[50] TeixeiraPJP, ColaianniNR, FitzpatrickCR, DanglJL. Beyond pathogens: microbiota interactions with the plant immune system. Curr Opin Microbiol, 2019, 49: 7-17.

[51] TianW, HouC, RenZ, et al. . A calmodulin-gated calcium channel links pathogen patterns to plant immunity. Nature, 2019, 572: 131-135.

[52] TianH, WuZ, ChenS, et al. . Activation of TIR signalling boosts pattern-triggered immunity. Nature, 2021, 598: 500-503.

[53] WalkerJC, ZhangR. Relationship of a putative receptor protein kinase from maize to the S-locus glycoproteins of Brassica. Nature, 1990, 345: 743-746.

[54] Wang C-F, Huang L-L, Buchenauer H, Han Q-M, Zhang H-C, Kang Z-S (2007) Histochemical studies on the accumulation of reactive oxygen species (O₂⁻ and H₂O₂) in the incompatible and compatible interaction of wheat-Puccinia striiformis f. sp. tritici. Physiol Mol Plant Pathol 71:230–239. https://doi.org/10.1016/j.pmpp.2008.02.006

[55] Wang J, Wang J, Shang H, Chen X, Xu X, Hu X (2019) TaXa21, a leucine-rich repeat receptor-like kinase gene associated with TaWRKY76 and TaWRKY62, plays positive roles in wheat high-temperature seedling plant resistance to Puccinia striiformis f. sp. tritici. Mol Plant-Microbe Interactions 32:1526–1535. https://doi.org/10.1094/MPMI-05-19-0137-R

[56] WangN, TangC, FanX, et al. . Inactivation of a wheat protein kinase gene confers broad-spectrum resistance to rust fungi. Cell, 2022, 185: 2961-2974.e19.

[57] WangY, AbroukM, GourdoupisS, et al. . An unusual tandem kinase fusion protein confers leaf rust resistance in wheat. Nat Genet, 2023, 55: 914-920.

[58] Wang Y, Liu X, Yuan B, Chen X, Zhao H, Ali Q, Zheng M, Tan Z, Yao H, Zheng S, Wu J, Xu J, Shi J, Wu H, Gao X, Gu Q (2024) Fusarium graminearum rapid alkalinization factor peptide negatively regulates plant immunity and cell growth via the FERONIA receptor kinase. Plant Biotechnol J 22:1800–1811. https://doi.org/10.1111/pbi.14303

[59] Wu Z, Zhang G, Zhao R, Gao Q, Zhao J, Zhu X, Wang F, Kang Z, Wang X (2023) Transcriptomic analysis of wheat reveals possible resistance mechanism mediated by Yr10 to stripe rust. Stress Biol 3:44. https://doi.org/10.1007/s44154-023-00115-z

[60] XieJ, ChenY, CaiG, et al. . Tree Visualization By One Table (tvBOT): a web application for visualizing, modifying and annotating phylogenetic trees. Nucleic Acids Res, 2023, 51: W587-W592.

[61] Xu L, Wang J, Xiao Y, Han Z, Chai J (2023) Structural insight into chitin perception by chitin elicitor receptor kinase 1 of Oryza sativa. J Integr Plant Biol 65:235–248. https://doi.org/10.1111/jipb.13279

[62] YadetaKA, ElmoreJM, CreerAY, et al. . A cysteine-rich protein kinase associates with a membrane immune complex and the cysteine residues are required for cell death. Plant Physiol, 2017, 173: 771-787.

[63] YinZ, LiuJ, DouD. RLKdb: A comprehensively curated database of plant receptor-like kinase families. Mol Plant, 2024, 17: 513-515.

[64] ZhangL, WangJ-C, HouL, et al. . Functional role of histidine in the conserved His-x-Asp motif in the catalytic core of protein kinases. Sci Rep, 2015, 5: 10115.

[65] ZhangR, ShiP-T, ZhouM, et al. . Rapid alkalinization factor: function, regulation, and potential applications in agriculture. Stress Biol, 2023, 3: 16.

[66] ZhangL, ZhuQ, TanY, et al. . Mitogen-activated protein kinases MPK3 and MPK6 phosphorylate receptor-like cytoplasmic kinase CDL1 to regulate soybean basal immunity. Plant Cell, 2024, 36: 963-986.

[67] ZhaoJ, KangZ. Fighting wheat rusts in China: a look back and into the future. Phytopathol Res, 2023, 5: 6.

[68] ZhouH, LiS, DengZ, et al. . Molecular analysis of three new receptor-like kinase genes from hexaploid wheat and evidence for their participation in the wheat hypersensitive response to stripe rust fungus infection. Plant J, 2007, 52: 420-434.

[69] ZhuS, FuQ, XuF, et al. . New paradigms in cell adaptation: decades of discoveries on the Cr RLK1L receptor kinase signalling network. New Phytol, 2021, 232: 1168-1183.

[70] ZipfelC. Plant pattern-recognition receptors. Trends Immunol, 2014, 35: 345-351.

[71] ZipfelC, KunzeG, ChinchillaD, et al. . Perception of the Bacterial PAMP EF-Tu by the Receptor EFR Restricts Agrobacterium-Mediated Transformation. Cell, 2006, 125: 749-760.

Funding

National Natural Science Foundation(32172381)

National Natural Science Foundation of China(32102175)

Innovation Capability Support Program of Shaanxi(2023-CX-TD-56)

111 Project from the Ministry of Education of China(BP0719026)

National Key Research and Development Program of China(2021YFD1401003)

RIGHTS & PERMISSIONS

The Author(s)