Genome-wide identification of the ICS family genes and its role in resistance to Plasmodiophora brassicae in Brassica napus L.

The isochorismate synthase (ICS) proteins are essential regulators of salicylic acid (SA) synthesis, which has been reported to regulate resistance to biotic and abiotic stresses in plants. Clubroot caused by Plasmodiophora brassicae is a common disease that threatens the yield and quality of Oilseed rape (Brassica napus L.). Exogenous application of salicylic acid reduced the incidence of clubroot in oilseed rape. However, the potential importance of the ICS genes family in B. napus and its diploid progenitors has been unclear. Here, we identified 16, 9, and 10 ICS genes in the allotetraploid B. napus, diploid ancestor Brassica rapa and Brassica oleracea, respectively. These ICS genes were classified into three subfamilies (I-III), and member of the same subfamilies showed relatively conserved gene structures, motifs, and protein domains. Furthermore, many hormone-response and stress-related promoter cis-acting elements were observed in the BnaICS genes. Exogenous application of SA delayed the growth of clubroot galls, and the expression of BnaICS genes was significantly different compared to the control groups. Protein-protein interaction analysis identified 58 proteins involved in the regulation of ICS in response to P. brassicae in B. napus. These results provide new clues for understanding the resistance mechanism to P. brassicae.

The chloroplast singlet oxygen-triggered biosynthesis of salicylic acid and jasmonic acid is mediated by EX1 and GUN1 in Arabidopsis.

Reactive oxygen species (ROS) and defence hormones like salicylic acid (SA) and jasmonic acid (JA) play pivotal roles in triggering cell death. However, the precise mechanism governing the interaction between ROS and SA/JA remains elusive. Recently, our research revealed that RNAi mutants with suppressed expression of PROGRAMMED CELL DEATH8 (PCD8) exhibit an overabundance of tetrapyrrole intermediates, particularly uroporphyrinogen III (Uro III), leading to the accumulation of singlet oxygen (1O2) during the transition from darkness to light, thereby instigating leaf necrosis. In this investigation, we uncovered that 1O2 stimulates biosynthesis of SA and JA, activating SA/JA signalling and the expression of responsive genes in PCD8 RNAi (pcd8) mutants. Introducing NahG or knocking out PAD4 or NPR1 significantly alleviates the cell death phenotype of pcd8 mutants, while coi1 partially mitigates the pcd8 phenotype. Further exploration revealed that EX1 and GUN1 can partially rescue the pcd8 phenotype by reducing the levels of Uro III and 1O2. Notably, mutations in EX1 mutations but not GUN1, substantially diminish SA content in pcd8 mutants compared to the wild type, implying that EX1 acts as the primary mediator of 1O2 signalling-mediated SA biosynthesis. Moreover, the triple ex1 gun1 pcd8 displays a phenotype similar to ex1. Overall, our findings underscore that the 1O2-induced cell death phenotype requires EX1/GUN1-mediated retrograde signalling in pcd8 mutants, providing novel insights into the interplay between ROS and SA/JA.

Root system architecture change in response to waterlogging stress in a 448 global collection of rapeseeds (Brassica napus L.).

MAIN CONCLUSIONS: A novel image-based screening method for precisely identifying genotypic variations in rapeseed RSA under waterlogging stress was developed. Five key root traits were confirmed as good indicators of waterlogging and might be employed in breeding, particularly when using the MFVW approach. Waterlogging is a vital environmental factor that has detrimental effects on the growth and development of rapeseed (Brassica napus L.). Plant roots suffer from hypoxia under waterlogging, which ultimately confers yield penalty. Therefore, it is crucially important to understand the genetic variation of root system architecture (RSA) in response to waterlogging stress to guide the selection of new tolerant cultivars with favorable roots. This research was conducted to investigate RSA traits using image-based screening techniques to better understand how RSA changes over time during waterlogging at the seedling stage. First, we performed a t-test by comparing the relative root trait value between four tolerant and four sensitive accessions. The most important root characteristics associated with waterlogging tolerance at 12 h are total root length (TRL), total root surface area (TRSA), total root volume (TRV), total number of tips (TNT), and total number of forks (TNF). The root structures of 448 rapeseed accessions with or without waterlogging showed notable genetic diversity, and all traits were generally restrained under waterlogging conditions, except for the total root average diameter. Additionally, according to the evaluation and integration analysis of 448 accessions, we identified that five traits, TRL, TRSA, TRV, TNT, and TNF, were the most reliable traits for screening waterlogging-tolerant accessions. Using analysis of the membership function value (MFVW) and D-value of the five selected traits, 25 extremely waterlogging-tolerant materials were screened out. Waterlogging significantly reduced RSA, inhibiting root growth compared to the control. Additionally, waterlogging increased lipid peroxidation, accompanied by a decrease in the activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT). This study effectively improves our understanding of the response of RSA to waterlogging. The image-based screening method developed in this study provides a new scientific guidance for quickly examining the basic RSA changes and precisely predicting waterlogging-tolerant rapeseed germplasms, thus expanding the genetic diversity of waterlogging-tolerant rapeseed germplasm available for breeding.

Maize GOLDEN2-LIKE proteins enhance drought tolerance in rice by promoting stomatal closure.

Drought has become one of the most severe abiotic stresses experienced in agricultural production across the world. Plants respond to water deficit via stomatal movements in the leaves, which are mainly regulated by abscisic acid (ABA). A previous study from our lab showed that constitutive expression of maize (Zea mays L.) GOLDEN2-LIKE (GLK) transcription factors in rice (Oryza sativa L.) can improve stomatal conductance and plant photosynthetic capacity under field conditions. In the present study, we uncovered a function of ZmGLK regulation of stomatal movement in rice during drought stress. We found that elevated drought tolerance in rice plants overexpressing ZmGLK1 or GOLDEN2 (ZmG2) was conferred by rapid ABA-mediated stomatal closure. Comparative analysis of RNA-sequencing (RNA-seq) data from the rice leaves and DNA affinity purification sequencing (DAP-seq) results obtained in vitro revealed that ZmGLKs played roles in regulating ABA-related and stress-responsive pathways. Four upregulated genes closely functioning in abiotic stress tolerance with strong binding peaks in the DAP-seq data were identified as putative target genes of ZmGLK1 and ZmG2 in rice. These results demonstrated that maize GLKs play an important role in regulating stomatal movements to coordinate photosynthesis and stress tolerance. This trait is a valuable target for breeding drought-tolerant crop plants without compromising photosynthetic capacity.

PROGRAMMED CELL DEATH8 interacts with tetrapyrrole biosynthesis enzymes and ClpC1 to maintain homeostasis of tetrapyrrole metabolites in Arabidopsis.

Tetrapyrrole biosynthesis (TBS) is a dynamically and strictly regulated process. Disruptions in tetrapyrrole metabolism influence many aspects of plant physiology, including photosynthesis, programmed cell death (PCD), and retrograde signaling, thus affecting plant growth and development at multiple levels. However, the genetic and molecular basis of TBS is not fully understood. We report here PCD8, a newly identified thylakoid-localized protein encoded by an essential gene in Arabidopsis. PCD8 knockdown causes a necrotic phenotype due to excessive chloroplast damage. A burst of singlet oxygen that results from overaccumulated tetrapyrrole intermediates upon illumination is suggested to be responsible for cell death in the knockdown mutants. Genetic and biochemical analyses revealed that PCD8 interacts with ClpC1 and a number of TBS enzymes, such as HEMC, CHLD, and PORC of TBS. Taken together, our findings uncover the function of chloroplast-localized PCD8 and provide a new perspective to elucidate molecular mechanism of how TBS is finely regulated in plants.