Disruption of EARLY LESION LEAF 1, encoding a cytochrome P450 monooxygenase, induces ROS accumulation and cell death in rice.

Lesion-mimic mutants (LMMs) provide a valuable tool to reveal the molecular mechanisms determining programmed cell death (PCD) in plants. Despite intensive research, the mechanisms behind PCD and the formation of lesions in various LMMs still remain to be elucidated. Here, we identified a rice (Oryza sativa) LMM, early lesion leaf 1 (ell1), cloned the causal gene by map-based cloning, and verified this by complementation. ELL1 encodes a cytochrome P450 monooxygenase, and the ELL1 protein was located in the endoplasmic reticulum. The ell1 mutant exhibited decreased chlorophyll contents, serious chloroplast degradation, upregulated expression of chloroplast degradation-related genes, and attenuated photosynthetic protein activity, indicating that ELL1 is involved in chloroplast development. RNA sequencing analysis showed that genes related to oxygen binding were differentially expressed in ell1 and wild-type plants; histochemistry and paraffin sectioning results indicated that hydrogen peroxide (H2 O2 ) and callose accumulated in the ell1 leaves, and the cell structure around the lesions was severely damaged, which indicated that reactive oxygen species (ROS) accumulated and cell death occurred in the mutant. TUNEL staining and comet experiments revealed that severe DNA degradation and abnormal PCD occurred in the ell1 mutants, which implied that excessive ROS accumulation may induce DNA damage and ROS-mediated cell death in the mutant. Additionally, lesion initiation in the ell1 mutant was light dependent and temperature sensitive. Our findings revealed that ELL1 affects chloroplast development or function, and that loss of ELL1 function induces ROS accumulation and lesion formation in rice.

UDP-N-acetylglucosamine pyrophosphorylase enhances rice survival at high temperature.

High-temperature stress inhibits normal cellular processes and results in abnormal growth and development in plants. However, the mechanisms by which rice (Oryza sativa) copes with high temperature are not yet fully understood. In this study, we identified a rice high temperature enhanced lesion spots 1 (hes1) mutant, which displayed larger and more dense necrotic spots under high temperature conditions. HES1 encoded a UDP-N-acetylglucosamine pyrophosphorylase, which had UGPase enzymatic activity. RNA sequencing analysis showed that photosystem-related genes were differentially expressed in the hes1 mutant at different temperatures, indicating that HES1 plays essential roles in maintaining chloroplast function. HES1 expression was induced under high temperature conditions. Furthermore, loss-of-function of HES1 affected heat shock factor expression and its mutation exhibited greater vulnerability to high temperature. Several experiments revealed that higher accumulation of reactive oxygen species occurred in the hes1 mutant at high temperature. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and comet experiments indicated that the hes1 underwent more severe DNA damage at high temperature. The determination of chlorophyll content and chloroplast ultrastructure showed that more severe photosystem defects occurred in the hes1 mutant under high temperature conditions. This study reveals that HES1 plays a key role in adaptation to high-temperature stress in rice.

MORE FLORET1 Encodes a MYB Transcription Factor That Regulates Spikelet Development in Rice.

Rice (Oryza sativa) spikelets have a unique inflorescence structure, and the mechanisms regulating their development are not yet fully understood. Moreover, approaches to manipulate spikelet development have the potential to increase grain yield. In this study, we identified and characterized a recessive spikelet mutant, namely more floret1 (mof1). The mof1 mutant has a delayed transition from the spikelet to the floral meristem, inducing the formation of extra lemma-like and palea-like organs. In addition, the main body of the palea was reduced, and the sterile lemma was enlarged and partially acquired hull (lemma and/or palea) identity. We used map-based cloning to identify the MOF1 locus and confirmed our identification by complementation and by generating new mof1 alleles using CRISPR-Cas9 gene editing. MOF1 encodes a MYB domain protein with the typical ethylene response factor-associated amphiphilic repression motifs, is expressed in all organs and tissues, and has a strong repression effect. MOF1 localizes to the nucleus and interacts with TOPLESS-RELATED PROTEINs to possibly repress the expression of downstream target genes. Taken together, our results reveal that MOF1 plays an important role in the regulation of organ identity and spikelet determinacy in rice.

Detecting and pyramiding target QTL for plant- and grain-related traits via chromosomal segment substitution line of rice.

Introduction: Plant height and grain length are important agronomic traits in rice, exhibiting a strong effect on plant architecture and grain quality of rice varieties. Methods: Methods: A novel rice chromosomal segment substitution line (CSSL), i.e., CSSL-Z1357, with significantly increased plant height (PH) and grain length (GL) was identified from CSSLs constructed by using Nipponbare as a receptor and a restorer line Xihui 18 as a donor. Seven agronomic traits of PH, PL, GL, GW, GPP, SPP, and TGW were phenotyped, and REML implemented in HPMIXED of SAS were used to detect the QTL for these traits. Secondary CSSLs were screened out via marker-assisted selection (MAS) to estimate the additive and epistatic effects of detected QTLs, evaluating the potential utilization of pyramiding the target QTLs for yield and quality improvement of rice varieties. Results and Discussion: Results and Discussion: CSSL-Z1357 carried nine segments from Xihui 18 with an average segment length of 4.13 Mb. The results show that the long grain of CSSL-Z1357 was caused by the increased number of surface cells and the length of the inner glume. Thirteen quantitative trait loci were identified via the F2 population of Nipponbare/CSSL-Z1357, including three each for GL (qGL-3, qGL-6, and qGL-7) and PH (qPH-1, qPH-7, and qPH-12I), among which qGL-3 increased GL by 0.23 mm with synergistic allele from CSSL-Z1357. Additionally, three single (S1 to S3), two double (D1, D2), and one triple segment (T1) substitution lines were developed in F3 via MAS. Results show that pyramiding the segments from Chr.3 (qGL-3 and qPH-3), Chr.6 (qGL-6 and qPH-6), and Chr.7 (Null and qPH-7) tended to result in better phenotype of increased GL and PH and decreased grain width, providing a potential basis for enhancing grain yield and quality in rice breeding.

The ell1 mutation disrupts tryptophan metabolism and induces cell death.

Tryptophan metabolism pathways are important components of the plant immune system; for example, serotonin is derived from tryptophan, and plays a vital role in rice (Oryza sativa) innate immunity. Recently, we isolated a rice mutant, early lesion leaf 1 (ell1), which exhibits lesions. RNA-seq analysis revealed that KEGG pathways related to amino acid metabolism were significantly enriched in the transcripts differentially expressed in this mutant. Furthermore, measurements of free amino acid contents revealed the accumulated tryptophan of ell1 mutant. In addition, the transcript levels of genes related to tryptophan biosynthesis were significantly enhanced in the ell1 mutant. These results revealed that ELL1 plays a critical role in tryptophan metabolism. Based on these findings, it is revealed that loss of ELL1 function may disrupt tryptophan metabolism, thereby inducing cell death and forming lesions in rice.

ABNORMAL FLOWER AND GRAIN 1 encodes OsMADS6 and determines palea identity and affects rice grain yield and quality.

The palea and lemma are floral organ structures unique to grasses; these structures form the hull and directly affect grain size and quality. However, the molecular mechanisms controlling the development of the hull are not well understood. In this study, we characterized the rice (Oryza sativa) abnormal flower and grain1 (afg1) mutant, a new allele of OsMADS6. Similar to previously characterized osmads6 alleles, in the afg1 floret, the palea lost its marginal region and acquired the lemma identity. However, in contrast to other osmads6 alleles, the afg1 mutant showed altered grain size and grain quality, with decreased total starch and amylose contents, and increased protein and soluble sugar contents. The analysis of transcriptional activity suggested that AFG1 is a transcriptional activator and may affect grain size by regulating the expression levels of several genes related to cell expansion and proliferation in the afg1 mutant. These results revealed that AFG1 plays an important role in determining palea identity and affecting grain yield and quality in rice.

The C2H2 zinc-finger protein LACKING RUDIMENTARY GLUME 1 regulates spikelet development in rice.

Rice (Oryza sativa) spikelets are a unique inflorescence structure and their development directly determines grain size and yield. Although many genes related to spikelet development have been reported, the molecular mechanisms underlying this process have not been fully elucidated. In this study, we identified a new recessive rice mutant, lacking rudimentary glume 1 (lrg1). The lrg1 spikelets only formed one rudimentary glume, which, along with the sterile lemmas, was homeotically transformed into lemma-like organs and acquired lemma identity. The transition from the spikelet to the floral meristem was delayed in the lrg1 mutant, resulting in the formation of an ectopic lemma-like organ between the sterile lemma and the terminal floret. In addition, we found that the abnormal lrg1 grain phenotype resulted from the alteration of cell numbers and the hull size. LRG1 encodes a ZOS4-06-C2H2 zinc-finger protein with the typical EAR motifs, and is expressed in all organs and tissues. LRG1 localizes to the nucleus and can interact with the TOPLESS-RELATED PROTEINs (TPRs) to repress the expressions of their downstream target genes. Taken together, our results reveal that LRG1 plays an important role in the regulation of spikelet organ identity and grain size.