OsKASI-2 is required for the regulation of unsaturation levels of membrane lipids and chilling tolerance in rice.

Chilling stress has seriously limited the global production and geographical distribution of rice. However, the molecular mechanisms associated with plant responses to chilling stress are less known. In this study, we revealed a member of ß-ketoacyl-ACP synthase I family (KASI), OsKASI-2 which confers chilling tolerance in rice. OsKASI-2 encodes a chloroplast-localized KASI enzyme mainly expressed in the leaves and anthers of rice and strongly induced by chilling stress. Disruption of OsKASI-2 led to decreased KAS enzymatic activity and the levels of unsaturated fatty acids, which impairs degree of unsaturation of membrane lipids, thus increased sensitivity to chilling stress in rice. However, the overexpression of OsKASI-2 significantly improved the chilling tolerance ability in rice. In addition, OsKASI-2 may regulate ROS metabolism in response to chilling stress. Natural variation of OsKASI-2 might result in difference in chilling tolerance between indica and japonica accessions, and Hap1 of OsKASI-2 confers chilling tolerance in rice. Taken together, we suggest OsKASI-2 is critical for regulating degree of unsaturation of membrane lipids and ROS accumulation for maintenance of membrane structural homeostasis under chilling stress, and provide a potential target gene for improving chilling tolerance of rice.

bHLH57 confers chilling tolerance and grain yield improvement in rice.

Chilling stress has become a major limiting factor that reduces crop productivity worldwide. In this study, we identified a new gene bHLH57, whose product enhances chilling tolerance in rice at diverse developmental stages. bHLH57 was mainly expressed in leaves and anthers, and its protein was targeted to the nucleus. Overexpression of bHLH57 enhanced chilling tolerance by increasing trehalose synthesis, whereas its mutants by CRISPR/Cas9-mediated mutagenesis were more sensitive to chilling and had reduced trehalose. Meanwhile, bHLH57 may regulate ROS metabolism and CBFs/DREBs- dependent pathways in response to chilling stress. In addition, the overexpression of bHLH57 resulted in increased grain yield under normal and chilling conditions, however, the disruption of bHLH57 displayed decreased grain size and seed setting rate, thus reduced grain yield. Phylogenetic and nucleotide diversity analyses suggested that bHLH57 is relatively conserved in monocotyledons, and may be selected during indica populations adaptation. Taken together, we have identified a new bHLH regulator involved rice chilling tolerance and grain yield, and provide a potential target gene for improving chilling tolerance and grain yield of rice.

ABA biosynthesis gene OsNCED3 contributes to preharvest sprouting resistance and grain development in rice.

Preharvest sprouting (PHS) is an unfavorable trait in cereal crops and causes serious yield loss. However, the molecular mechanism underlying PHS remains largely elusive. Here, we identified a member of 9-cis-epoxycarotenoid dioxygenase family, OsNCED3, which regulates PHS and grain development in rice (Oryza sativa L.). OsNCED3 encodes a chloroplast-localized abscisic acid (ABA) biosynthetic enzyme highly expressed in the embryo of developing seeds. Disruption of OsNCED3 by CRISPR/Cas9-mediated mutagenesis led to a lower ABA and higher gibberellic acid (GA) levels (thus a skewed ABA/GA ratio) in the embryo, promoting embryos growth and breaking seed dormancy before seed maturity and harvest, thus decreased seed dormancy and enhanced PHS in rice. However, the overexpression of OsNCED3 enhanced PHS resistance by regulating proper ABA/GA ratio in the embryo. Intriguingly, the overexpression of OsNCED3 resulted in increased grain size and weight, whereas the disruption of OsNCED3 function decreased grain size and weight. Nucleotide diversity analyses suggested that OsNCED3 may be selected during japonica populations adaptation of seed dormancy and germination. Taken together, we have identified a new OsNCED regulator involved rice PHS and grain development, and provide a potential target gene for improving PHS resistance and grain development in rice.

Characterization and correlation analysis of microbial flora and flavor profile of stinky acid, a Chinese traditional fermented condiment.

To explore the microbial diversity and flavor profiles of stinky acid, we utilized high-throughput sequencing, culture-based techniques, and bionic E-sensory technologies. The results revealed a significant correlation between the acidity levels of stinky acid and the richness of its microbial community. Ten core bacterial genera and three core fungal genera exhibited ubiquity across all stinky acid samples. Through E-nose analysis, it was found that sulfides constituted the principal odor compounds responsible for stinky acid's distinct aroma. Further insights arose from the correlation analysis, indicating the potential contribution of Debaryomyces yeast to the sour taste profile. Meanwhile, three genera-Rhizopus and Thermoascus and Companilactobacillus-were identified as contributors to aromatic constituents. Interestingly, the findings indicated that Rhizopus and Thermoascus could reduce the intensity of the pungent odor of stinky acid. In summary, this investigation's outcomes offer new insights into the complex bacterial diversity of stinky acid.

Agrobacterium-Mediated High-Efficiency Genetic Transformation and Genome Editing of Chaling Common Wild Rice (Oryza rufipogon Griff.) Using Scutellum Tissue of Embryos in Mature Seeds.

Genetic transformation is an important strategy for revealing gene function, and it is used extensively in both functional genomics study and molecular breeding of rice. Demand for its application in wild Oryza species is rising for their extensive genetic diversity. However, genetic transformation of wild Oryza accessions with AA genome using calli induced from scutellum tissue of embryos in mature seeds has not been successfully established. In the present study, we used Chaling common wild rice (CLCWR) (Oryza rufipogon Griff.) with AA genome to successfully establish an Agrobacterium-mediated genetic transformation system based on scutellum tissue of embryos in mature seeds. The calli from embryos in mature seeds of CLCWR were easy to be induced and regenerated. The callus induction rate and texture were optimum under 2.5 mg/L 2,4-D. The optimal hormone combination used for regeneration was 2 mg/L ZT + 0.1 mg/L NAA. Studies on genetic transformation and genome editing showed that the transformation efficiency was 87-94%, the efficiency of single genome editing and multiplex genome editing were about 60-70% and 20-40%, respectively. Compared with Nipponbare (Nip), CLCWR had higher Hygromycin-resistant callus frequency and transformation efficiency. Taken together, our study establishes a highly efficient transformation system for common wild rice with AA genome and provides a good rice material for de novo domestication by genome editing in the future.

OsNCED5, a 9-cis-epoxycarotenoid dioxygenase gene, regulates salt and water stress tolerance and leaf senescence in rice.

9-cis-epoxycarotenoid dioxygenase (NCED) is a rate-limiting enzyme for abscisic acid (ABA) biosynthesis. However, the molecular mechanisms of NCED5 that modulate plant development and abiotic stress tolerance are still unclear, particular in rice. Here, we demonstrate that a rice NCED gene, OsNCED5, was expressed in all tissues we tested, and was induced by exposure to salt stress, water stress, and darkness. Mutational analysis showed that nced5 mutants reduced ABA level and decreased tolerance to salt and water stress and delayed leaf senescence. However, OsNCED5 overexpression increased ABA level, enhanced tolerance to the stresses, and accelerated leaf senescence. Transcript analysis showed that OsNCED5 regulated ABA-dependent abiotic stress and senescence-related gene expression. Additionally, ectopic expression of OsNCED5 tested in Arabidopsis thaliana altered plant size and leaf morphology and delayed seed germination and flowering time. Thus, OsNCED5 may regulate plant development and stress resistance through control of ABA biosynthesis. These findings contribute to our understanding of the molecular mechanisms by which NCED regulates plant development and responses to abiotic stress in different crop species.