As core components of ABA signaling pathway, SnRK2s (Sucrose nonfermenting1–Related protein Kinase 2) bind to and phosphorylate AREB/ABF (ABA responsive element binding protein/ABRE-binding factor) transcriptional factors, particularly bZIPs (basic region-leucine zipper), to participate in various biological processes, including flowering. Rice contains 10 SnRK2 members denoted as SAPK1-10 (Stress-Activated Protein Kinase) and dozens of bZIPs. However, which of the SAPKs and bZIPs pair and involve in ABA signaling remains largely unknown. In this study, we carried out a systematical protein-protein interactomic analysis of 10 SAPKs and 9 ABA-inducible bZIPs using yeast-two-hybrid technique, and identified 14 positive interactions. The reliability of Y2H work was verified by in vitro pull-down assay of the key flowering regulator bZIP77 with SAPK9 and SAPK10, respectively. Moreover, SAPK10 could phosphorylate bZIP77 in vitro. Over-expression of SAPK10 resulted in earlier flowering time, at least partially through regulating the FAC-MADS15 pathway. Conclusively, our results provided an overall view of the SAPK-bZIP interactions, and shed novel lights on the mechanisms of ABA-regulated rice flowering.

Cyclin-dependent kinase inhibitors known as KRPs (kip-related proteins) control the progression of plant cell cycles and modulate various plant developmental processes. However, the function of KRPs in rice remains largely unknown. In this study, two rice KRPs members, KRP1 and KRP2, were found to be predominantly expressed in developing seeds and were significantly induced by exogenous abscisic acid (ABA) and Brassinosteroid (BR) applications. Sub-cellular localization experiments showed that KRP1 was mainly localized in the nucleus of rice protoplasts. KRP1 overexpression transgenic lines (OxKRP1), krp2 single mutant (crkrp2), and krp1/krp2 double mutant (crkrp1/krp2) all exhibited significantly smaller seed width, seed length, and reduced grain weight, with impaired seed germination and retarded early seedling growth, suggesting that disturbing the normal steady state of KRP1 or KRP2 blocks seed development partly through inhibiting cell proliferation and enlargement during grain filling and seed germination. Furthermore, two cyclin-dependent protein kinases, CDKC;2 and CDKF;3, could interact with KRP1 in a yeast-two-hybrid system, indicating that KRP1 might regulate the mitosis cell cycle and endoreduplication through the two targets. In a word, this study shed novel insights into the regulatory roles of KRPs in rice seed maturation and germination.

Rice seed is a pivotal reproductive organ that directly determines yield and quality. Long non-coding RNAs (lncRNAs) have been recognized as key regulators in plant development, but the roles of lncRNAs in rice seed development remain unclear. In this study, we performed a paired-end RNA sequencing in samples of rice pistils and seeds at three and seven days after pollination (DAP) respectively. A total of 540 lncRNAs were obtained, among which 482 lncRNAs had significantly different expression patterns during seed development. Results from semi-qPCR conducted on 15 randomly selected differentially expressed lncRNAs suggested high reliability of the transcriptomic data. RNA interference of TCONS_00023703, which is predominantly transcribed in developing seeds, significantly reduced grain length and thousand-grain weight. These results expanded the dataset of lncRNA in rice and enhanced our understanding of the biological functions of lncRNAs in rice seed development

Spotted-leaf mutants are important to reveal programmed cell death and defense-related pathways in rice. We previously characterized the phenotype performance of a rice spotted-leaf mutant spl21 and narrowed down the causal gene locus spl21(t) to an 87-kb region in chromosome 12 by map-based cloning. We showed that a single base substitution from A to G at position 836 in the coding sequence of Oryza sativa beta-1,6-N-acetylglucosaminyl transferase (OsGCNT), effectively mutating Tyr to Cys at position 279 in the translated protein sequence, was responsible for the spotted-leaf phenotype as it could be rescued by functional complementation. Compared to the wild type IR64, the spotted-leaf mutant spl21 exhibited loss of chlorophyll, breakdown of chloroplasts, down-regulation of photosynthesis-related genes, and up-regulation of senescence associated genes, which indicated that OsGCNT regulates premature leaf senescence. Moreover, the enhanced resistance to the bacterial leaf blight pathogen Xanthomonas oryzae pv. oryzae, up-regulation of pathogenesis-related genes and increased level of jasmonate which suggested that OsGCNT is a negative regulator of defense response in rice. OsGCNT was expressed constitutively in the leaves, sheaths, stems, roots, and panicles, and OsGCNT-GFP was localized to the Golgi apparatus. High throughput RNA sequencing analysis provided further evidence for the biological effects of loss of OsGCNT function on cell death, premature leaf senescence and enhanced disease resistance in rice. Thus, we demonstrated that the novel OsGCNT regulated rice innate immunity and immunity-associated leaf senescence probably by changing the jasmonate metabolic pathway. These results reveal that a novel gene Oryza sativa beta-1,6-N-acetylglucosaminyl transferase (OsGCNT) is responsible for the spotted-leaf mutant spl21, and OsGCNT acts as a negative-regulator mediating defense response and immunity-associated premature leaf senescence probably by activating jasmonate signaling pathway.

The leaf blade is the main photosynthetic organ and its morphology is related to light energy capture and conversion efficiency. We isolated a novel rice Dynamic Narrow-Rolled Leaf 1 (dnrl1) mutant showing reduced width of leaf blades, rolled leaves and lower chlorophyll content. The narrow-rolled leaf phenotype resulted from the reduced number of small longitudinal veins per leaf, smaller size and irregular arrangement of bulliform cells compared with the wild-type. DNRL1 was mapped to chromosome 7 and encoded a putative 3-deoxy-7-phosphoheptulonate synthase (DAHPS) which catalyzes the conversion of phosphoenolpyruvate and D-erythrose 4-phosphate to DAHP and phosphate. Sequence analysis revealed that a single base substitution (T–A) was detected in dnrl1, leading to a single amino acid change (L376H) in the coding protein. The mutation led to a lower expression level of DNRL1 as well as the lower activity of DAHPS in the mutant compared with the wild type. Genetic complementation and over-expression of DNRL1 could rescue the narrow-rolled phenotype. DNRL1 was constitutively expressed in all tested organs and exhibited different expression patterns from other narrow-rolled leaf genes. DNRL1-GFP located to chloroplasts. The lower level of chlorophyll in dnrl1 was associated with the downregulation of the genes responsible for chlorophyll biosynthesis and photosynthesis. Furthermore, dnrl1 showed significantly reduced levels of aromatic amino acids including Trp, Phe and Tyr. We conclude that OsDAHPS, encoded by DNRL1, plays a critical role in leaf morphogenesis by mediating the biosynthesis of amino acids in rice.