Protein homeostasis (proteostasis) is crucial for proper cellular function, including the production of peptides with biological functions through controlled proteolysis. Proteostasis has roles in maintenance of cellular functions and plant interactions with the environment under physiological conditions. Plant stress continues to reduce agricultural yields causing substantial economic losses; thus, it is critical to understand how plants perceive stress signals to elicit responses for survival. As previously shown in
Proteome-wide analysis of cysteine reactivity during effector-triggered immunity
A surge in the accumulation of oxidants generates shifts in the cellular redox potential during early stages of plant infection with pathogens and activation of effector-triggered immunity (ETI). The redoxome, defined as the proteome-wide oxidative modifications of proteins caused by oxidants, has a well-known impact on stress responses in metazoans. However, the identity of proteins and the residues sensitive to oxidation during the plant immune response remain largely unknown. Previous studies of the thimet oligopeptidases TOP1 and TOP2 placed them in the salicylic acid dependent branch of ETI, with a current model wherein TOPs sustain interconnected organellar and cytosolic pathways that modulate the oxidative burst and development of cell death. Herein, we characterized the ETI redoxomes in Arabidopsis (Arabidopsis thaliana) wild-type Col-0 and top1top2 mutant plants using a differential alkylation-based enrichment technique coupled with label-free mass spectrometry-based quantification. We identified cysteines sensitive to oxidation in a wide range of protein families at multiple time points after pathogen infection. Differences were detected between Col-0 and top1top2 redoxomes regarding the identity and number of oxidized cysteines, and the amplitude of time-dependent fluctuations in protein oxidation. Our results support a determining role for TOPs in maintaining the proper level and dynamics of proteome oxidation during ETI. This study significantly expands the repertoire of oxidation-sensitive plant proteins and can guide future mechanistic studies.
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- Award ID(s):
- 1714157
- NSF-PAR ID:
- 10093518
- Date Published:
- Journal Name:
- Plant physiology
- Volume:
- 179
- Issue:
- 4
- ISSN:
- 1532-2548
- Page Range / eLocation ID:
- 1248-1264
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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SUMMARY Arabidopsis thaliana , thimet oligopeptidases (TOPs) TOP1 (also referred to as organellar oligopeptidase) and TOP2 (also referred to as cytosolic oligopeptidase) are essential components in plant response to pathogens, but further characterization of TOPs and their peptide substrates is required to understand their contributions to stress perception and defense signaling. Herein, label‐free peptidomics via liquid chromatography‐tandem mass spectrometry was used to differentially quantify 1111 peptides, originating from 369 proteins, between the Arabidopsis Col‐0 wild type andtop1top2 knock‐out mutant. This revealed 350 peptides as significantly more abundant in the mutant, representing accumulation of these potential TOP substrates. Ten direct substrates were validated usingin vitro enzyme assays with recombinant TOPs and synthetic candidate peptides. These TOP substrates are derived from proteins involved in photosynthesis, glycolysis, protein folding, biogenesis, and antioxidant defense, implicating TOP involvement in processes aside from defense signaling. Sequence motif analysis revealed TOP cleavage preference for non‐polar residues in the positions surrounding the cleavage site. Identification of these substrates provides a framework for TOP signaling networks, through which the interplay between proteolytic pathways and defense signaling can be further characterized. -
Plant intracellular immune receptor NLR (nucleotide-binding leucine-rich repeat) proteins sense the presence of pathogens and trigger strong and robust immune responses. NLR genes are known to be tightly controlled at the protein level, but little is known about their dynamics at the transcript level. In this study, we presented a meta-analysis of transcript dynamics of all 207 NLR genes in the Col-0 accession of Arabidopsis thaliana under various biotic and abiotic stresses based on 88 publicly available RNA sequencing datasets from 27 independent studies. We find that about two thirds of the NLR genes are generally induced by pathogens, immune elicitors, or salicylic acid (SA), suggesting that transcriptional induction of NLR genes might be an important mechanism in plant immunity regulation. By contrast, NLR genes induced by biotic stresses are often repressed by abscisic acid, high temperature and drought, suggesting that transcriptional regulation of NLR genes might be important for interaction between abiotic and biotic stress responses. In addition, pathogen-induced expression of some NLR genes are dependent on SA induction. Interestingly, a small group of NLR genes are repressed under certain biotic stress treatments, suggesting an unconventional function of this group of NLRs. This meta-analysis thus reveals the transcript dynamics of NLR genes under biotic and abiotic stress conditions and suggests a contribution of NLR transcript regulation to plant immunity as well as interactions between abiotic and biotic stress responses.more » « less
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Abstract Sigma factor (
SIG ) proteins contribute to promoter specificity of the plastid‐encodedRNA polymerase during chloroplast genome transcription. All six members of theSIG family, that is,SIG 1–SIG 6, are nuclear‐encoded proteins targeted to chloroplasts. Sigma factor 2 (SIG 2) is a phytochrome‐regulated protein important for stoichiometric control of the expression of plastid‐ and nuclear‐encoded genes that impact plastid development and plant growth and development. AmongSIG factors,SIG 2 is required not only for transcription of chloroplast genes (i.e., anterograde signaling), but also impacts nuclear‐encoded, photosynthesis‐related, and light signaling‐related genes (i.e., retrograde signaling) in response to plastid functional status. AlthoughSIG 2 is involved in photomorphogenesis in Arabidopsis, the molecular bases for its role in light signaling that impacts photomorphogenesis and aspects of photosynthesis have only recently begun to be investigated. Previously, we reported thatSIG 2 is necessary for phytochrome‐mediated photomorphogenesis specifically under red (R) and far‐red light, thereby suggesting a link between phytochromes and nuclear‐encodedSIG 2 in light signaling. To explore transcriptional roles ofSIG 2 in R‐dependent growth and development, we performedRNA sequencing analysis to compare gene expression insig2‐2 mutant and Col‐0 wild‐type seedlings at two developmental stages (1‐ and 7‐day). We identified a subset of misregulated genes involved in growth, hormonal cross talk, stress responses, and photosynthesis. To investigate the functional relevance of these gene expression analyses, we performed several comparative phenotyping tests. In these analyses, strongsig2 mutants showed insensitivity to bioactiveGA 3, high intracellular levels of hydrogen peroxide (H2O2) indicative of a stress response, and specific defects in photosynthesis, including elevated levels of cyclic electron flow (CEF ) and nonphotochemical quenching (NPQ ). We demonstrated thatSIG 2 regulates a broader range of physiological responses at the molecular level than previously reported, with specific roles in red‐light‐mediated photomorphogenesis. -
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