Plant Cell Reports最新文献

Key message: Hydrogen peroxide promoted leaf senescence by sulfenylating the magnesium chelating protease I subunit (CHLI1) in the chlorophyll synthesis pathway, and inhibited its activity to reduce chlorophyll synthesis. Leaf senescence is the final and crucial stage of plant growth and development, during which chlorophyll experiences varying degrees of destruction. It is well-known that the higher ROS accumulation is a key factor for leaf senescence, but whether and how ROS regulates chlorophyll synthesis in the process are unknown. Here, we report that H₂O₂ inhibits chlorophyll synthesis during leaf senescence via the I subunit of magnesium-chelatase (CHLI1). During leaf senescence, the decrease of chlorophyll content is accompanied by the increase of H₂O₂ accumulation, as well as the inhibition of catalase (CAT) genes expression. The mutant cat2-1, with increased H₂O₂ shows an accelerated senescence phenotype and decreased CHLI1 activity compared with the wild type. H₂O₂ inhibits CHLI1 activity by sulfenylating CHLI1 during leaf senescence. Consistent with this, the chli1 knockout mutant displays the same premature leaf senescence symptom as cat2-1, while overexpression of CHLI1 in cat2-1 can partially restore its early senescence phenotype. Taken together, these results illustrate that CAT2-mediated H₂O₂ accumulation during leaf senescence represses chlorophyll synthesis through sulfenylating CHLI1, and thus inhibits its activity, providing a new insight into the pivotal role of chlorophyll synthesis as a participant in orchestrating the leaf senescence.

Key message: The N-terminal transmembrane domain of LPAT1 crosses the inner membrane placing the N terminus in the intermembrane space and the C-terminal enzymatic domain in the stroma. Galactolipids mono- and di-galactosyl diacylglycerol are the major and vital lipids of photosynthetic membranes. They are synthesized by five enzymes hosted at different sub-chloroplast locations. However, localization and topology of the second-acting enzyme, lysophosphatidic acid acyltransferase 1 (LPAT1), which acylates the sn-2 position of glycerol-3-phosphate (G3P) to produce phosphatidic acid (PA), remain unclear. It is not known whether LPAT1 is located at the outer or the inner envelope membrane and whether its enzymatic domain faces the cytosol, the intermembrane space, or the stroma. Even the size of mature LPAT1 in chloroplasts is not known. More information is essential for understanding the pathways of metabolite flow and for future engineering endeavors to modify glycerolipid biosynthesis. We used LPAT1 preproteins translated in vitro for import assays to determine the precise size of the mature protein and found that the LPAT1 transit peptide is at least 85 residues in length, substantially longer than previously predicted. A construct comprising LPAT1 fused to the Venus fluorescent protein and driven by the LPAT1 promoter was used to complement an Arabidopsis lpat1 knockout mutant. To determine the sub-chloroplast location and topology of LPAT1, we performed protease treatment and alkaline extraction using chloroplasts containing in vitro-imported LPAT1 and chloroplasts isolated from LPAT1-Venus-complemented transgenic plants. We show that LPAT1 traverses the inner membrane via an N-terminal transmembrane domain, with its N terminus protruding into the intermembrane space and the C-terminal enzymatic domain residing in the stroma, hence displaying a different membrane topology from its bacterial homolog, PlsC.

Key message: Fluorescence in situ hybridization with frozen sections of root tips showed difference of chromosome territories distribution between autosome and sex-chromosome homologous pairs in Populus trichocarpa. The spatial organization of chromatin within the interphase nucleus and the interactions between chromosome territories (CTs) are essential for various biologic processes. Three-dimensional fluorescence in situ hybridization (3D-FISH) is a powerful tool for analyzing CTs, but its application in plants is limited. In this study, we established a 3D-FISH technique using frozen sections of Populus trichocarpa root tips, which was an improvement over the use of paraffin sections and enabled us to acquire good FISH signals. Using chromosome-specific oligo probes, we were able to analyze CTs in interphase nuclei in three dimensions. The distribution of chromosome pairs 17 and 19 in the 3D-preserved nuclei of P. trichocarpa root tip cells were analyzed and showed that the autosome pair 17 associated more often than sex chromosome 19. This research lays a foundation for further study of the spatial position of chromosomes in the nucleus and the relationship between gene expression and spatial localization of chromosomes in poplar.

Key message: The genomic organization, phylogenetic relationship, expression patterns, and genetic variations of m6A-related genes were systematically investigated in wild emmer wheat and the function of TdFIP37 regulating salt tolerance was preliminarily determined. m6A modification is one of the most abundant and crucial RNA modifications in eukaryotics, playing the indispensable role in growth and development as well as stress response in plants. However, its significance in wild emmer wheat remains elusive. Here, a genome-wide search of m6A-related genes was conducted in wild emmer wheat to obtain 64 candidates, including 21 writers, 17 erasers, and 26 readers. Phylogenetic and collinearity analysis demonstrated that segmental duplication and polyploidization contributed mainly to the expansion of m6A-related genes in wild emmer. A number of cis-acting elements involving in stress and hormonal regulation were found in the promoter regions of them, such as MBS, LTR, and ABRE. Genetic variation of them was also investigated using resequencing data and obvious genetic bottleneck was occurred on them during wild emmer wheat domestication process. Furthermore, the salt-responsive candidates were investigated through RNA-seq data and qRT-PCR validation using the salt-tolerant and -sensitive genotypes and the co-expression analysis showed that they played the hub role in regulating salt stress response. Finally, the loss-function mutant of Tdfip37 displayed the significantly higher salt-sensitive compared to WT and then RNA-seq analysis demonstrated that FIP37 mediated the MAPK pathway, hormone signal transduction, as well as transcription factor to regulate salt tolerance. This study provided the potential m6A genes for functional analysis, which will contribute to better understand the regulatory roles of m6A modification and also improve the salt tolerance from the perspective of epigenetic approach in emmer wheat and other crops.

Key message: Wheat TaCDPK1-5A plays critical roles in mediating drought tolerance through regulating osmotic stress-associated physiological processes. Calcium (Ca²⁺) acts as an essential second messenger in plant signaling pathways and impacts plant abiotic stress responses. This study reported the function of TaCDPK1-5A, a calcium-dependent protein kinase (CDPK) gene in T. aestivum, in mediating drought tolerance. TaCDPK1-5A sensitively responded to drought and exogenous abscisic acid (ABA) signaling, displaying induced transcripts in plants under drought and ABA treatments. Yeast two-hybrid and co-immunoprecipitation assays revealed that TaCDPK1-5A interacts with the mitogen-activated protein kinase TaMAPK4-7D whereas the latter with ABF transcription factor TaABF1-3A, suggesting that TaCDPK1-5A constitutes a signaling module with above partners to transduce signals initiated by drought/ABA stressors. Overexpression of TaCDPK1-5A, TaMAPK4-7D and TaABF1-3A enhanced plant drought adaptation by modulating the osmotic stress-related physiological indices, including increased osmolyte contents, enlarged root morphology, and promoted stomata closure. Yeast one-hybrid assays indicated the binding ability of TaABF1-3A with promoters of TaP5CS1-1B, TaPIN3-5A, and TaSLAC1-3-2A, the genes encoding P5CS enzyme, PIN-FORMED protein, and slow anion channel, respectively. ChIP-PCR and transcriptional activation assays confirmed that TaABF1-3A regulates these genes at transcriptional level. Moreover, transgene analysis indicated that these stress-responsive genes positively regulated proline biosynthesis (TaP5CS1-1B), root morphology (TaPIN3-5A), and stomata closing (TaSLAC1-3-2A) upon drought signaling. Positive correlations were observed between yield and the transcripts of TaCDPK1-5A signaling partners in wheat cultivars under drought condition, with haplotype TaCDPK1-5A-Hap1 contributing to improved drought tolerance. Our study concluded that TaCDPK1-5A positively regulates drought adaptation and is a valuable target for molecular breeding the drought-tolerant cultivars in T. aestivum.

Key message: CRISPR/Cas9-mediated knockout of SlWOX4 gene in tomato enhances tolerance to drought stress. Drought stress is one of the major abiotic factors that seriously affects plant growth and crop yield. WUSCHEL-related homeobox (WOX) transcription factors are involved in plant growth, development and stress response. However, little is known about the role of WOX genes in drought tolerance in tomato. Here, SlWOX4, a member of the WOX family in tomato, was functionally characterized in mediating drought tolerance. SlWOX4 was homologous to Nicotiana tabacum NtWOX4 with a conserved HD domain, and was localized in the nucleus. SlWOX4 was significantly down-regulated by drought and abscisic acid (ABA) treatments. The loss-of-function mutations of SlWOX4 produced using the CRISPR-Cas9 system in tomato improved drought tolerance by reducing water loss rate and enhancing stomatal closure. In addition, the wox4 lines exhibited reduced accumulation of reactive oxygen species (ROS) and malondialdehyde (MDA), increased antioxidant enzyme activity, proline contents and ABA contents under drought stress. Moreover, gene editing of SlWOX4 in tomato enhanced drought tolerance by regulating the expression of genes encoding antioxidants and ABA signaling molecules. In summary, SlWOX4 gene might negatively regulate drought stress tolerance in tomato and has great potential as a drought-resistant crop-breeding target genes.

Key message: The Arabidopsis RNA helicase LOS4 plays a key role in regulating pre-mRNA splicing of the genes EIN2, CTR1, and ERS2 in ethylene signaling pathway. The plant hormone ethylene plays diverse roles in plant growth, development, and responses to stress. Ethylene is perceived by the membrane-bound ethylene receptors complex, and then triggers downstream components, such as EIN2, to initiate signal transduction into the nucleus, leading to the activation of ethylene-responsive genes. Over the past decades, substantial information has been accumulated regarding gene cloning, protein-protein interactions, and downstream gene expressions in the ethylene pathway. However, our understanding of mRNA post-transcriptional processing and modification of key genes in the ethylene signaling pathway remains limited. This study aims to provide evidence demonstrating the involvement of the Arabidopsis RNA helicase LOS4 in pre-mRNA splicing of the genes EIN2, CTR1, and ERS2 in ethylene signaling pathway. Various genetic approaches including RNAi gene silencing, CRISPR-Cas9 gene editing, and amino acid mutations were employed in this study. When LOS4 was silenced or knocked down, the ethylene sensitivity of etiolated seedlings was significantly enhanced. Further investigation revealed errors in the EIN2 pre-mRNA splicing when LOS4 was knocked down. In addition, aberrant pre-mRNA splicing was observed in the ERS2 and CTR1 genes in the pathway. Biochemical assays indicated that the los4-2 (E94K) mutant protein exhibited increased ATP binding and enhanced ATP hydrolytic activity. Conversely, the los4-1 (G364R) mutant had reduced substrate RNA binding and lower ATP binding activities. These findings significantly advanced our comprehension of the regulatory functions and molecular mechanisms of RNA helicase in ethylene signaling.

Key message: ARG6 and ARG10 pea accessions exhibited better tolerance to drought by keeping drought-associated attributes stable and higher, that is, stable chlorophyll content, high antioxidant activity, and the presence of polymorphic bands with stress-responsive EST-SSR markers. Each year, a significant portion of crops is lost due to various abiotic stresses, and even pea (Pisum sativum) crop growth and yield are severely affected by the challenges posed by drought stress. Drought is a critical factor that limits crop growth and development, and its impact is exacerbated by changes in the magnitude of climatic conditions. Drought induces oxidative stress in plants, leading to the accumulation of high concentrations of reactive oxygen species that damage cell structures and vital functioning of cells. The primary objective was to identify stress-tolerant plants by evaluating different morphological and biochemical attributes, such as biomass, chlorophyll content, relative water content, ascorbate peroxidase (APX), superoxide dismutase (SOD), and DPPH scavenging activity, as well as protein, proline, and phenolic content. Our study revealed that pea accessions (ARG6 and ARG10) were more resilient to drought stress as their chlorophyll, relative water, protein, and proline contents increased under drought conditions. Antioxidant enzymes, such as SOD, APX, and DPPH activities, also increased under drought stress in ARG10 and ARG6, suggesting that these accessions could bolster the antioxidant defense system in response to drought stress. Based on putative (cellular, biological, and metabolic) functions, ten EST-SSR primers were selected for the amplification study. Three EST-SSR primers, AUMP06_110, AUMP18_300, and AUMP31_250, were used for ARG6 and ARG10. Based on the correlation between the presence or absence of specific EST-SSR alleles, various physiological and morphological traits, and DPPH scavenging activity, both ARG10 and ARG6 demonstrated resistance to drought stress.

Key message: Analysis of the N-terminome of Physcomitrella reveals N-terminal monomethylation of nuclear-encoded, mitochondria-localized proteins. Post- or co-translational N-terminal modifications of proteins influence their half-life as well as mediating protein sorting to organelles via cleavable N-terminal sequences that are recognized by the respective translocation machinery. Here, we provide an overview on the current modification state of the N-termini of over 4500 proteins from the model moss Physcomitrella (Physcomitrium patens) using a compilation of 24 N-terminomics datasets. Our data reveal distinct proteoforms and modification states and confirm predicted targeting peptide cleavage sites of 1,144 proteins localized to plastids and the thylakoid lumen, to mitochondria, and to the secretory pathway. In addition, we uncover extended N-terminal methylation of mitochondrial proteins. Moreover, we identified PpNTM1 (P. patens alpha N-terminal protein methyltransferase 1) as a candidate for protein methylation in plastids, mitochondria, and the cytosol. These data can now be used to optimize computational targeting predictors, for customized protein fusions and their targeted localization in biotechnology, and offer novel insights into potential dual targeting of proteins.

Key message: We report the mitochondrial genome of 39 diploid potatoes and identify a candidate ORF potentially linked to cytoplasmic male sterility in potatoes. Potato (Solanum tuberosum L.) holds a critical position as the foremost non-grain food crop, playing a pivotal role in ensuring global food security. Diploid potatoes constitute a vital genetic resource pool, harboring the potential to revolutionize modern potato breeding. Nevertheless, diploid potatoes are relatively understudied, and mitochondrial DNA can provide valuable insights into key potato breeding traits such as CMS. In this study, we examine and assemble the mitochondrial genome evolution and diversity of 39 accessions of diploid potatoes using high-fidelity (HiFi) sequencing. We annotated 54 genes for all the investigated accessions, comprising 34 protein-coding genes, 3 rRNA genes, and 17 tRNA genes. Our analyses revealed differences in repeats sequences between wild and cultivated landraces. To understand the evolution of diploid maternal lineage inheritance, we conducted phylogenetic analysis, which clearly distinguished mitochondrial from nuclear gene trees, further supporting the evidence-based of clustering between wild and cultivated landraces accessions. Our study discovers new candidate ORFs associated with CMS in potatoes, including ORF137, which is homologous to other CMS in Solanaceae. Ultimately, this work bridges the gap in mitochondrial genome research for diploid potatoes, providing a steppingstone into evolutionary studies and potato breeding.