Zhichao Zhao

Programmed cell death and immunity in plants are finely orchestrated to promote antimicrobial defense while preventing autoimmunity. However, the molecular mechanisms involved are not fully understood. Here, we isolated a rice mutant ecdr1 (enhanced cell death and resistance 1) that displayed an autoimmunity phenotype and enhanced resistance to rice blast and bacterial blight, and we identified ECDR1 as a new shared component in phosphoinositide 3-kinase (PI3K) and PI4K complexes, linking the hyccin-containing protein with plant defense responses. ECDR1 was expressed at all developmental stages and in all tissues examined. The ECDR1 protein was highly conserved in function across monocots and dicots. The 113 bp deletion in the ECDR1 promoter reduced its expression, leading to cell death and enhanced disease resistance. The ECDR1 protein was localized in plasma membrane and interacted with tetratricopeptide repeat protein 1 (TPR1) and TPR2, which in turn interacted with both PI4K1 and PI3K1, suggesting that ECDR1 could function as a component associated with not only PI4K complexes but also PI3K complexes to help catalyze PI into phosphatidylinositol 3-phosphate (PI3P) and PI4P. The lethality of all homozygous ecdr1, pi3k1, tpr1 and tpr2 mutants indicated the crucial roles of these genes in plant normal growth, which restricted our understanding of PI3K and PI4K functions. Alternatively, exogenous application of PI3K and PI4K inhibitors could substantially exacerbate the cell death and enhance disease resistance, implying their roles in plant immunity. Our findings provide novel insights into the regulatory mechanisms of ECDR1 in cell death and defense pathways, will aid in understanding the functions of PI3K and PI4K in plant immunity.

Appropriate heading date is an important safeguard for stabilizing rice yield and quality. Heading integrator Ehd1 and florigen Hd3a are crucial for rice heading. However, the mechanism by which Ehd1 transmits heading signals to Hd3a and the accurate regulation of Hd3a expression remains unclear. Here, we found that OsWOX13 functions as a signal transducer from Ehd1 to Hd3a for the accurate regulation of rice heading date. Ehd1 can directly bind to the OsWOX13 promoter, OsWOX13, in turn, binds to the Hd3a promoter to activate its expression. OsWOX13 interacts with Hd3a to inhibit OsWOX13-mediated activation of Hd3a expression, which precisely controls Hd3a levels. Meanwhile, late-heading phenotypes were observed in the ehd1, Oswox13, Oswox13 ehd1, hd3a, and Oswox13 hd3a, demonstrating that Ehd1, OsWOX13, and Hd3a coordinately regulate rice heading date. Furthermore, OsWOX13 can be divided into 2 haplotypes, Hap-T and Hap-C. Ehd1 elevated OsWOX13 expression in Hap-T compared with Hap-C. Longjing 31T-C (carrying Hap-C) exhibited later heading dates than Longjing 31 (carrying Hap-T). Rice varieties carrying Hap-T are predominantly cultivated in high-latitude regions. This study provides genetic resources and a theoretical foundation for improving the regional and climatic adaptability of rice.

Heading date is a key agronomic trait that affects crop yield and regional adaptability. In this study, we identified a rice (Oryza sativa) early heading mutant and cloned the causal heading inhibitor gene EARLY HEADING AT LONG DAY 4 (ELD4) using the MutMap method. The eld4 CRISPR mutants and ELD4 RNAi plants flowered earlier than the wild type under natural long-day (NLD) conditions. ELD4 is a zinc finger transcription factor that localizes in the nucleus. Biochemical and genetic evidence suggests that ELD4 physically interacts with the pseudo-response regulator (PRR) protein OsPRR95. The Osprr95 mutant also exhibits an earlier heading phenotype under NLD. ELD4 and OsPRR95 co-regulate heading date by directly binding to the promoter and the first intron of OsMADS51, which promotes heading date. Moreover, electrophoretic mobility shift and luciferase complementation assays showed that ELD4 enhances OsPRR95 binding to the first intron of OsMADS51. These results suggest that the ELD4-OsPRR95 module directly represses OsMADS51 in regulating flowering time. Haplotype analysis reveals that OsPRR95 haplotype 3, which is geographically distributed in the north, exhibits a shorter heading date than haplotype 1 found in the south. This indicates that selection at the OsPRR95 locus has enhanced the regional adaptability of rice.

G-protein signaling and brassinosteroid (BR) phytohormones play important roles in regulating rice (Oryza sativa) yield-related plant architecture, such as leaf inclination and grain size. However, the relationship between G-proteins and BR signaling has not been fully elucidated in rice. The present study indicates that the G-protein Gγ subunit DENSE AND ERECT PANICLE 1 (DEP1) positively regulates BR signaling in rice and that BRs promote DEP1 nuclear entry through GRAIN NUMBER ASSOCIATED (GNA). Additionally, DEP1 interacts with and acts upstream of OsMYB86, an R2R3-MYB family transcription factor that positively regulates BR signaling by directly binding to the promoter of its downstream gene BRASSINOSTEROID UPREGULATED 1 (BU1), activating its expression in rice. In the nucleus, DEP1 interacts with OsMYB86 and GNA, significantly enhancing OsMYB86-mediated activation of BU1 expression. Furthermore, BU1 interacts with another HLH protein, INCREASED LEAF INCLINATION1 (ILI1), and a bHLH protein, ILI1 BINDING bHLH (IBH1). Interaction between ILI1 and BU1 facilitates translocation of BU1 from the cytoplasm to the nucleus, where they impede IBH1 binding to the promoter of the AUXIN RESPONSE FACTOR 11 (OsARF11) gene, which is involved in crosstalk between BR and auxin, thus effectively relieving the IBH1-repressed transcription of OsARF11. These findings reveal a DEP1-mediated signaling pathway that links G-proteins to the traditional BR signaling pathway, ensuring the efficient activation of BR responses in rice.

Autophagy is a highly conserved cellular program in eukaryotic cells which mediates the degradation of cytoplasmic components through the lysosome, also named the vacuole in plants. However, the molecular mechanisms underlying the fusion of autophagosomes with the vacuole remain unclear. Here, we report the functional characterization of a rice (Oryza sativa) mutant with defects in storage protein transport in endosperm cells and accumulation of numerous autophagosomes in root cells. Cytological and immunocytochemical experiments showed that this mutant exhibits a defect in the fusion between autophagosomes and vacuoles. The mutant harbors a loss‐of‐function mutation in the rice homolog of Arabidopsis thaliana MONENSIN SENSITIVITY1 (MON1). Biochemical and genetic evidence revealed a synergistic interaction between rice MON1 and AUTOPHAGY‐RELATED 8a in maintaining normal growth and development. In addition, the rice mon1 mutant disrupted storage protein sorting to protein storage vacuoles. Furthermore, quantitative proteomics verified that the loss of MON1 function influenced diverse biological pathways including autophagy and vacuolar transport, thus decreasing the transport of autophagic and vacuolar cargoes to vacuoles. Together, our findings establish a molecular link between autophagy and vacuolar protein transport, and offer insights into the dual functions of the MON1–CCZ1 (CAFFEINE ZINC SENSITIVITY1) complex in plants.

CYP78A, a cytochrome P450 subfamily that includes rice (Oryza sativa L.) BIG GRAIN2 (BG2, CYP78A13) and Arabidopsis thaliana KLUH (KLU, CYP78A5), generate an unknown mobile growth signal (referred to as a CYP78A-derived signal) that increases grain (seed) size. However, the mechanism by which the CYP78A pathway increases grain size remains elusive. Here, we characterized a rice small grain mutant, small grain4 (smg4), with smaller grains than its wild type due to restricted cell expansion and cell proliferation in spikelet hulls. SMG4 encodes a multidrug and toxic compound extrusion (MATE) transporter. Loss of function of SMG4 causes smaller grains while overexpressing SMG4 results in larger grains. SMG4 is mainly localized to endoplasmic reticulum (ER) exit sites (ERESs) and partially localized to the ER and Golgi. Biochemically, SMG4 interacts with coat protein complex Ⅱ (COPⅡ) components (Sar1, Sec23, and Sec24) and CYP78As (BG2, GRAIN LENGTH 3.2 [GL3.2], and BG2-LIKE 1 [BG2L1]). Genetically, SMG4 acts, at least in part, in a common pathway with Sar1 and CYP78As to regulate grain size. In summary, our findings reveal a CYP78As–SMG4–COPⅡ regulatory pathway for grain size in rice, thus providing new insights into the molecular and genetic regulatory mechanism of grain size.

Strigolactones (SLs) constitute a class of plant hormones that regulate many aspects of plant development, including repressing tillering in rice (Oryza sativa). However, how SL pathways are regulated is still poorly understood. Here, we describe a rice mutant dwarf and high tillering1 (dht1), which exhibits pleiotropic phenotypes (such as dwarfism and increased tiller numbers) similar to those of mutants defective in SL signaling. We show that DHT1 encodes a monocotyledon-specific hnRNP-like protein that acts as a previously unrecognized intron splicing factor for many precursor mRNAs (pre-mRNAs), including for the SL receptor gene D14. We find that the dht1 (DHT1^I232F) mutant protein is impaired in its stability and RNA binding activity, causing defective splicing of D14 pre-mRNA and reduced D14 expression, and consequently leading to the SL signaling-defective phenotypes. Overall, our findings deepen our understanding of the functional diversification of hnRNP-like proteins and establish a connection between posttranscriptional splicing and SL signaling in the regulation of plant development.

Abscisic acid and gibberellic acid exert opposite effects on root growth and tillering by antagonistically regulating the APC/CTE-mediated degradation of rice S

GPA5 functions as a plant-specific effector of Rab5a required for mediating tethering and membrane fusion of dense vesicles with protein storage vacuoles in ric