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Displaying results 1 to 9 of 9.

Publications

Erickson, J.; Weckwerth, P.; Romeis, T.; Lee, J.; What’s new in protein kinase/phosphatase signalling in the control of plant immunity? Essays in Biochemistry 66, 621-634, (2022) DOI: 10.1042/ebc20210088

Plant immunity is crucial to plant health but comes at an expense. For optimal plant growth, tight immune regulation is required to prevent unnecessary rechannelling of valuable resources. Pattern- and effector-triggered immunity (PTI/ETI) represent the two tiers of immunity initiated after sensing microbial patterns at the cell surface or pathogen effectors secreted into plant cells, respectively. Recent evidence of PTI-ETI cross-potentiation suggests a close interplay of signalling pathways and defense responses downstream of perception that is still poorly understood. This review will focus on controls on plant immunity through phosphorylation, a universal and key cellular regulatory mechanism. Rather than a complete overview, we highlight “what’s new in protein kinase/phosphatase signalling” in the immunity field. In addition to phosphoregulation of components in the pattern recognition receptor (PRR) complex, we will cover the actions of the major immunity-relevant intracellular protein kinases/phosphatases in the ‘signal relay’, namely calcium-regulated kinases (e.g. calcium-dependent protein kinases, CDPKs), mitogen-activated protein kinases (MAPKs), and various protein phosphatases. We discuss how these factors define a phosphocode that generates cellular decision-making ‘logic gates’, which contribute to signalling fidelity, amplitude, and duration. To underscore the importance of phosphorylation, we summarize strategies employed by pathogens to subvert plant immune phosphopathways. In view of recent game-changing discoveries of ETI-derived resistosomes organizing into calcium-permeable pores, we speculate on a possible calcium-regulated phosphocode as the mechanistic control of the PTI-ETI continuum.
Publications

Sopeña-Torres, S.; Jordá, L.; Sánchez-Rodríguez, C.; Miedes, E.; Escudero, V.; Swami, S.; López, G.; Piślewska-Bednarek, M.; Lassowskat, I.; Lee, J.; Gu, Y.; Haigis, S.; Alexander, D.; Pattathil, S.; Muñoz-Barrios, A.; Bednarek, P.; Somerville, S.; Schulze-Lefert, P.; Hahn, M. G.; Scheel, D.; Molina, A.; YODA MAP3K kinase regulates plant immune responses conferring broad-spectrum disease resistance New Phytol. 218, 661-680, (2018) DOI: 10.1111/nph.15007

Mitogen‐activated protein kinases (MAPKs) cascades play essential roles in plants by transducing developmental cues and environmental signals into cellular responses. Among the latter are microbe‐associated molecular patterns perceived by pattern recognition receptors (PRRs), which trigger immunity.We found that YODA (YDA) – a MAPK kinase kinase regulating several Arabidopsis developmental processes, like stomatal patterning – also modulates immune responses. Resistance to pathogens is compromised in yda alleles, whereas plants expressing the constitutively active YDA (CA‐YDA) protein show broad‐spectrum resistance to fungi, bacteria, and oomycetes with different colonization modes. YDA functions in the same pathway as ERECTA (ER) Receptor‐Like Kinase, regulating both immunity and stomatal patterning.ER‐YDA‐mediated immune responses act in parallel to canonical disease resistance pathways regulated by phytohormones and PRRs. CA‐YDA plants exhibit altered cell‐wall integrity and constitutively express defense‐associated genes, including some encoding putative small secreted peptides and PRRs whose impairment resulted in enhanced susceptibility phenotypes. CA‐YDA plants show strong reprogramming of their phosphoproteome, which contains protein targets distinct from described MAPKs substrates.Our results suggest that, in addition to stomata development, the ER‐YDA pathway regulates an immune surveillance system conferring broad‐spectrum disease resistance that is distinct from the canonical pathways mediated by described PRRs and defense Hormones.
Publications

Zembek, P.; Danilecka, A.; Hoser, R.; Eschen-Lippold, L.; Benicka, M.; Grech-Baran, M.; Rymaszewski, W.; Barymow-Filoniuk, I.; Morgiewicz, K.; Kwiatkowski, J.; Piechocki, M.; Poznanski, J.; Lee, J.; Hennig, J.; Krzymowska, M.; Two Strategies of Pseudomonas syringae to Avoid Recognition of the HopQ1 Effector in Nicotiana Species Front. Plant Sci. 9, 978, (2018) DOI: 10.3389/fpls.2018.00978

Pseudomonas syringae employs a battery of type three secretion effectors to subvert plant immune responses. In turn, plants have developed receptors that recognize some of the bacterial effectors. Two strain-specific HopQ1 effector variants (for Hrp outer protein Q) from the pathovars phaseolicola 1448A (Pph) and tomato DC3000 (Pto) showed considerable differences in their ability to evoke disease symptoms in Nicotiana benthamiana. Surprisingly, the variants differ by only six amino acids located mostly in the N-terminal disordered region of HopQ1. We found that the presence of serine 87 and leucine 91 renders PtoHopQ1 susceptible to N-terminal processing by plant proteases. Substitutions at these two positions did not strongly affect PtoHopQ1 virulence properties in a susceptible host but they reduced bacterial growth and accelerated onset of cell death in a resistant host, suggesting that N-terminal mutations rendered PtoHopQ1 susceptible to processing in planta and, thus, represent a mechanism of recognition avoidance. Furthermore, we found that co-expression of HopR1, another effector encoded within the same gene cluster masks HopQ1 recognition in a strain-dependent manner. Together, these data suggest that HopQ1 is under high host-pathogen co-evolutionary selection pressure and P. syringae may have evolved differential effector processing or masking as two independent strategies to evade HopQ1 recognition, thus revealing another level of complexity in plant – microbe interactions.
Publications

Pecher, P.; Eschen-Lippold, L.; Herklotz, S.; Kuhle, K.; Naumann, K.; Bethke, G.; Uhrig, J.; Weyhe, M.; Scheel, D.; Lee, J.; The Arabidopsis thaliana mitogen-activated protein kinases MPK3 and MPK6 target a subclass of ‘VQ-motif’-containing proteins to regulate immune responses New Phytol. 203, 592-606, (2014) DOI: 10.1111/nph.12817

Mitogen‐activated protein kinase (MAPK) cascades play key roles in plant immune signalling, and elucidating their regulatory functions requires the identification of the pathway‐specific substrates.We used yeast two‐hybrid interaction screens, in vitro kinase assays and mass spectrometry‐based phosphosite mapping to study a family of MAPK substrates. Site‐directed mutagenesis and promoter‐reporter fusion studies were performed to evaluate the impact of substrate phosphorylation on downstream signalling.A subset of the Arabidopsis thaliana VQ‐motif‐containing proteins (VQPs) were phosphorylated by the MAPKs MPK3 and MPK6, and renamed MPK3/6‐targeted VQPs (MVQs). When plant protoplasts (expressing these MVQs) were treated with the flagellin‐derived peptide flg22, several MVQs were destabilized in vivo. The MVQs interact with specific WRKY transcription factors. Detailed analysis of a representative member of the MVQ subset, MVQ1, indicated a negative role in WRKY‐mediated defence gene expression – with mutation of the VQ‐motif abrogating WRKY binding and causing mis‐regulation of defence gene expression.We postulate the existence of a variety of WRKY‐VQP‐containing transcriptional regulatory protein complexes that depend on spatio‐temporal VQP and WRKY expression patterns. Defence gene transcription can be modulated by changing the composition of these complexes – in part – through MAPK‐mediated VQP degradation.
Publications

Maldonado-Bonilla, L. D.; Eschen-Lippold, L.; Gago-Zachert, S.; Tabassum, N.; Bauer, N.; Scheel, D.; Lee, J.; The Arabidopsis Tandem Zinc Finger 9 Protein Binds RNA and Mediates Pathogen-Associated Molecular Pattern-Triggered Immune Responses Plant Cell Physiol. 55, 412-425, (2014) DOI: 10.1093/pcp/pct175

Recognition of pathogen-associated molecular patterns (PAMPs) induces multiple defense mechanisms to limit pathogen growth. Here, we show that the Arabidopsis thaliana tandem zinc finger protein 9 (TZF9) is phosphorylated by PAMP-responsive mitogen-activated protein kinases (MAPKs) and is required to trigger a full PAMP-triggered immune response. Analysis of a tzf9 mutant revealed attenuation in specific PAMP-triggered reactions such as reactive oxygen species accumulation, MAPK activation and, partially, the expression of several PAMP-responsive genes. In accordance with these weaker PAMP-triggered responses, tzf9 mutant plants exhibit enhanced susceptibility to virulent Pseudomonas syringae pv. tomato DC3000. Visualization of TZF9 localization by fusion to green fluorescent protein revealed cytoplasmic foci that co-localize with marker proteins of processing bodies (P-bodies). This localization pattern is affected by inhibitor treatments that limit mRNA availability (such as cycloheximide or actinomycin D) or block nuclear export (leptomycin B). Coupled with its ability to bind the ribohomopolymers poly(rU) and poly(rG), these results suggest involvement of TZF9 in post-transcriptional regulation, such as mRNA processing or storage pathways, to regulate plant innate immunity.
Publications

Reddy, P. S.; Kavi Kishor, P. B.; Seiler, C.; Kuhlmann, M.; Eschen-Lippold, L.; Lee, J.; Reddy, M. K.; Sreenivasulu, N.; Unraveling Regulation of the Small Heat Shock Proteins by the Heat Shock Factor HvHsfB2c in Barley: Its Implications in Drought Stress Response and Seed Development PLOS ONE 9, e89125, (2014) DOI: 10.1371/journal.pone.0089125

The rapid increase in heat shock proteins upon exposure to damaging stresses and during plant development related to desiccation events reveal their dual importance in plant development and stress tolerance. Genome-wide sequence survey identified 20 non-redundant small heat shock proteins (sHsp) and 22 heat shock factor (Hsf) genes in barley. While all three major classes (A, B, C) of Hsfs are localized in nucleus, the 20 sHsp gene family members are localized in different cell organelles like cytoplasm, mitochondria, plastid and peroxisomes. Hsf and sHsp members are differentially regulated during drought and at different seed developmental stages suggesting the importance of chaperone role under drought as well as seed development. In silico cis-regulatory motif analysis of Hsf promoters showed an enrichment with abscisic acid responsive cis-elements (ABRE), implying regulatory role of ABA in mediating transcriptional response of HvsHsf genes. Gene regulatory network analysis identified HvHsfB2c as potential central regulator of the seed-specific expression of several HvsHsps including 17.5CI sHsp. These results indicate that HvHsfB2c is co-expressed in the central hub of small Hsps and therefore it may be regulating the expression of several HvsHsp subclasses HvHsp16.88-CI, HvHsp17.5-CI and HvHsp17.7-CI. The in vivo relevance of binding specificity of HvHsfB2C transcription factor to HSE-element present in the promoter of HvSHP17.5-CI under heat stress exposure is confirmed by gel shift and LUC-reporter assays. Further, we isolated 477 bp cDNA from barley encoding a 17.5 sHsp polypeptide, which was predominantly upregulated under drought stress treatments and also preferentially expressed in developing seeds. Recombinant HvsHsp17.5-CI protein was expressed in E. coli and purified to homogeneity, which displayed in vitro chaperone activity. The predicted structural model of HvsHsp-17.5-CI protein suggests that the α-crystallin domain is evolutionarily highly conserved.
Publications

Schenke, D.; Böttcher, C.; Lee, J.; Scheel, D.; Verticillin A is likely not produced by Verticillium sp. J. Antibiot. 64, 523-524, (2011) DOI: 10.1038/ja.2011.36

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Publications

Brock, A. K.; Willmann, R.; Kolb, D.; Grefen, L.; Lajunen, H. M.; Bethke, G.; Lee, J.; Nürnberger, T.; Gust, A. A.; The Arabidopsis Mitogen-Activated Protein Kinase Phosphatase PP2C5 Affects Seed Germination, Stomatal Aperture, and Abscisic Acid-Inducible Gene Expression Plant Physiol. 153, 1098-1111, (2010) DOI: 10.1104/pp.110.156109

Abscisic acid (ABA) is an important phytohormone regulating various cellular processes in plants, including stomatal opening and seed germination. Although protein phosphorylation via mitogen-activated protein kinases (MAPKs) has been suggested to be important in ABA signaling, the corresponding phosphatases are largely unknown. Here, we show that a member of the Protein Phosphatase 2C (PP2C) family in Arabidopsis (Arabidopsis thaliana), PP2C5, is acting as a MAPK phosphatase. The PP2C5 protein colocalizes and directly interacts with stress-induced MPK3, MPK4, and MPK6, predominantly in the nucleus. Importantly, altered PP2C5 levels affect MAPK activation. Whereas Arabidopsis plants depleted of PP2C5 show an enhanced ABA-induced activation of MPK3 and MPK6, ectopic expression of PP2C5 in tobacco (Nicotiana benthamiana) resulted in the opposite effect, with the two MAPKs salicylic acid-induced protein kinase and wound-induced protein kinase not being activated any longer after ABA treatment. Moreover, depletion of PP2C5, whose gene expression itself is affected by ABA treatment, resulted in altered ABA responses. Loss-of-function mutation in PP2C5 or AP2C1, a close PP2C5 homolog, resulted in an increased stomatal aperture under normal growth conditions and a partial ABA-insensitive phenotype in seed germination that was most prominent in the pp2c5 ap2c1 double mutant line. In addition, the response of ABA-inducible genes such as ABI1, ABI2, RD29A, and Erd10 was reduced in the mutant plants. Thus, we suggest that PP2C5 acts as a MAPK phosphatase that positively regulates seed germination, stomatal closure, and ABA-inducible gene expression.
Publications

Varet, A.; Hause, B.; Hause, G.; Scheel, D.; Lee, J.; The Arabidopsis NHL3 Gene Encodes a Plasma Membrane Protein and Its Overexpression Correlates with Increased Resistance to Pseudomonas syringae pv. tomato DC3000 Plant Physiol. 132, 2023-2033, (2003) DOI: 10.1104/pp.103.020438

The Arabidopsis genome contains a family of NDR1/HIN1-like (NHL) genes that show homology to the nonrace-specific disease resistance (NDR1) and the tobacco (Nicotiana tabacum) harpin-induced (HIN1) genes. NHL3 is a pathogen-responsive member of this NHL gene family that is potentially involved in defense. In independent transgenic NHL3-overexpressing plant lines, a clear correlation between increased resistance to virulent Pseudomonas syringae pv. tomato DC3000 and enhanced NHL3 transcript levels was seen. These transgenic plants did not show enhanced pathogenesis-related gene expression or reactive oxygen species accumulation. Biochemical and localization experiments were performed to assist elucidation of how NHL3 may confer enhanced disease resistance. Gene constructs expressing amino-terminal c-myc-tagged or carboxyl-terminal hemagglutinin epitope (HA)-tagged NHL3 demonstrated membrane localization in transiently transformed tobacco leaves. Stable Arabidopsis transformants containing the NHL3-HA construct corroborated the findings observed in tobacco. The detected immunoreactive proteins were 10 kD larger than the calculated size and could be partially accounted for by the glycosylation state. However, the expected size was not attained with deglycosylation, suggesting possibly additional posttranslational modification. Detergent treatment, but not chemicals used to strip membrane-associated proteins, could displace the immunoreactive signal from microsomal fractions, showing that NHL3 is tightly membrane associated. Furthermore, immunofluorescence and immunogold labeling, coupled with two-phase partitioning techniques, revealed plasma membrane localization of NHL3-HA. This subcellular localization of NHL3 positions it at an initial contact site to pathogens and may be important in facilitating interception of pathogen-derived signals.
  • Die Leibniz-Gemeinschaft
  • Digitalization in agriculture
  • Universität Halle
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