NAC (NAM, ATAF1/2, and CUC2) transcription factors are ubiquitously distributed in eukaryotes and play significant roles in stress response. However, the functional verifications of NACs in Picea (P.) wilsonii remain largely uncharacterized. Here, we identified the NAC transcription factor PwNAC11 as a mediator of drought stress, which was significantly upregulated in P. wilsonii under drought and abscisic acid (ABA) treatments. Yeast two-hybrid assays showed that both the full length and C-terminal of PwNAC11 had transcriptional activation activity and PwNAC11 protein cannot form a homodimer by itself. Subcellular observation demonstrated that PwNAC11 protein was located in nucleus. The overexpression of PwNAC11 in Arabidopsis obviously improved the tolerance to drought stress but delayed flowering time under nonstress conditions. The steady-state level of antioxidant enzymes’ activities and light energy conversion efficiency were significantly increased in PwNAC11 transgenic lines under dehydration compared to wild plants. PwNAC11 transgenic lines showed hypersensitivity to ABA and PwNAC11 activated the expression of the downstream gene ERD1 by binding to ABA-responsive elements (ABREs) instead of drought-responsive elements (DREs). Genetic evidence demonstrated that PwNAC11 physically interacted with an ABA-induced protein—ABRE Binding Factor3 (ABF3)—and promoted the activation of ERD1 promoter, which implied an ABA-dependent signaling cascade controlled by PwNAC11. In addition, qRT-PCR and yeast assays showed that an ABA-independent gene—DREB2A—was also probably involved in PwNAC11-mediated drought stress response. Taken together, our results provide the evidence that PwNAC11 plays a dominant role in plants positively responding to early drought stress and ABF3 and DREB2A synergistically regulate the expression of ERD1.

Sugar transport from the source leaf to the sink organ is critical for seed development and crop yield, as well as for responding to abiotic stress. SWEETs (sugar will eventually be exported transporters) mediate sugar efflux into the reproductive sink and are therefore considered key candidate proteins for sugar unloading during seed development. However, the specific mechanism underlying the sugar unloading to seeds in Camellia oleifera remains elusive. Here, we identified a SWEET gene named CoSWEET10, which belongs to Clade III and has high expression levels in the seeds of C. oleifera. CoSWEET10 is a plasma membrane-localized protein. The complementation assay of CoSWEET10 in SUSY7/ura3 and EBY.VW4000 yeast strains showed that CoSWEET10 has the ability to transport sucrose, glucose, and fructose. Through the C. oleifera seeds in vitro culture, we found that the expression of CoSWEET10 can be induced by hexose and sucrose, and especially glucose. By generating the restoration lines of CoSWEET10 in Arabidopsis atsweet10, we found that CoSWEET10 restored the seed defect phenotype of the mutant by regulating soluble sugar accumulation and increased plant drought tolerance. Collectively, our study demonstrates that CoSWEET10 plays a dual role in promoting seed development and enhancing plant drought resistance as a sucrose and hexose transporter.

The NAC family of transcription factors (TFs) regulate plant development and abiotic stress. However, the specific function and response mechanism of NAC TFs that increase drought resistance in Picea wilsonii remain largely unknown. In this study, we functionally characterized a member of the PwNAC family known as PwNAC31. PwNAC31 is a nuclear-localized protein with transcriptional activation activity and contains an NAC domain that shows extensive homology with ANAC072 in Arabidopsis. The expression level of PwNAC31 is significantly upregulated under drought and ABA treatments. The heterologous expression of PwNAC31 in atnac072 Arabidopsis mutants enhances the seed vigor and germination rates and restores the hypersensitive phenotype of atnac072 under drought stress, accompanied by the up-regulated expression of drought-responsive genes such as DREB2A (DEHYDRATION-RESPONSIVE ELEMENT BINDING PROTEIN 2A) and ERD1 (EARLY RESPONSIVE TO DEHYDRATION STRESS 1). Yeast two-hybrid and bimolecular fluorescence complementation assays confirmed that PwNAC31 interacts with DREB2A and ABF3 (ABSCISIC ACID-RESPONSIVE ELEMENT-BINDING FACTOR 3). Yeast one-hybrid and dual-luciferase assays showed that PwNAC31, together with its interaction protein DREB2A, directly regulated the expression of ERD1 by binding to the DRE element of the ERD1 promoter. Collectively, our study provides evidence that PwNAC31 activates ERD1 by interacting with DREB2A to enhance drought tolerance in transgenic Arabidopsis.

Sucrose phloem unloading plays a vital role in photoassimilate distribution and storage in sink organs such as fruits and seeds. In most plants, the phloem unloading route was reported to shift between an apoplasmic and a symplasmic pattern with fruit development. However, the molecular transition mechanisms of the phloem unloading pathway still remain largely unknown. In this study, we applied RNA sequencing to profile the specific gene expression patterns for sucrose unloading in C. oleifera fruits in the apo- and symplasmic pathways that were discerned by CF fluoresce labelling. Several key structural genes were identified that participate in phloem unloading, such as PDBG11, PDBG14, SUT8, CWIN4, and CALS10. In particular, the key genes controlling the process were involved in callose metabolism, which was confirmed by callose staining. Based on the co-expression network analysis with key structural genes, a number of transcription factors belonging to the MYB, C2C2, NAC, WRKY, and AP2/ERF families were identified to be candidate regulators for the operation and transition of phloem unloading. KEGG enrichment analysis showed that some important metabolism pathways such as plant hormone metabolism, starch, and sucrose metabolism altered with the change of the sugar unloading pattern. Our study provides innovative insights into the different mechanisms responsible for apo- and symplasmic phloem unloading in oil tea fruit and represents an important step towards the omics delineation of sucrose phloem unloading transition in crops.

Sugar transporters of the SWEET family are essential for plant growth, development, yield formation, and stress responses by regulating sugar transport and distribution. This study characterizes the function and expression characteristics of CoSWEET2a, a Clade I SWEET gene in Camellia oleifera. We conducted subcellular localization, functional complementation in Arabidopsis, sugar response assays, drought tolerance tests, and hormone induction analysis. A key finding is CoSWEET2a, which that is localized on the vacuolar membrane in Camellia oleifera. Heterologous expression in Arabidopsis atsweet2 mutants revealed sugar-specific effects on root growth. Moreover, expression of CoSWEET2a increased soluble sugar content in Arabidopsis seeds. Additionally, CoSWEET2a overexpression enhanced drought stress tolerance by augmenting sugar content. The expression of CoSWEET2a is regulated by gibberellin (GA) and abscisic acid (ABA), and its promoter contains corresponding hormone response elements. In conclusion, CoSWEET2a functions as a “sugar buffer” on the vacuolar membrane, regulating sugar accumulation, root development, and drought stress responses. This discovery not only reveals that vacuolar SWEET plays an important role in maintaining cytoplasmic sugar homeostasis in plants but also provides a direct genetic target for engineering high-quality, drought-tolerant crops.