OsTBP2.1, a TATA-Binding Protein, Alters the Ratio of OsNRT2.3b to OsNRT2.3a and Improves Rice Grain Yield.

The OsNRT2.3a and OsNRT2.3b isoforms play important roles in the uptake and transport of nitrate during rice growth. However, it is unclear which cis-acting element controls the transcription of OsNRT2.3 into these specific isoforms. In this study, we used a yeast one-hybrid assay to obtain the TATA-box binding protein OsTBP2.1, which binds to the TATA-box of OsNRT2.3, and verified its important role through transient expression and RNA-seq. We found that the TATA-box of OsNRT2.3 mutants and binding protein OsTBP2.1 together increased the transcription ratio of OsNRT2.3b to OsNRT2.3a. The overexpression of OsTBP2.1 promoted nitrogen uptake and increased rice yield compared with the wild-type; however, the OsTBP2.1 T-DNA mutant lines exhibited the opposite trend. Detailed analyses demonstrated that the TATA-box was the key cis-regulatory element for OsNRT2.3 to be transcribed into OsNRT2.3a and OsNRT2.3b. Additionally, this key cis-regulatory element, together with the binding protein OsTBP2.1, promoted the development of rice and increased grain yield.

OsLSD1.1 is involved in the photosystem II reaction and affects nitrogen allocation in rice.

Nitrogen (N) is an essential nutrient element for plants; however, high N accumulation often leads to a decrease in photosynthetic nitrogen use efficiency (PNUE). In rice (Oryza sativa L.), well-developed aerenchyma is formed to promote oxygen transport from the shoot to the root tips as an adaptation to submerged and oxygen-deficient environment. Total N concentrations were increased in the rice root by changes in O2 levels in the rhizosphere. However, few reports have focused on how aerenchyma formation-related genes participate in photosynthesis and affect nitrogen allocation in rice. In this study, we found that OsLSD1.1, located in the chloroplast, cell membrane, and nucleus, may be involved in the photosystem II reaction and affect chloroplast development. OsLSD1.1 knockout was found to significantly reduce the quantum efficiency of the PSII reaction center (ΦPSII). Furthermore, we observed that the nitrogen accumulation decreased in the grain of OsLSD1.1 mutants. RNA-Seq transcriptome analysis revealed that OsPEPC3, OsPsbR1, OsNRG2, OsNRT1.5A, OsNRT1.7, and OsAMT3;2 were downregulated in m12 compared with N-WT (wild-type Nipponbare), which may be a reason that photosynthesis and nitrogen transport were inhibited. Taken together, our findings demonstrated that OsLSD1.1 may be key in plant growth, photosynthesis, and nitrogen allocation in rice. Our results may provide theoretical support for the discovery of key genes for nitrogen physiological use efficiency.

Plant DNA methylation is sensitive to parent seed N content and influences the growth of rice.

BACKGROUND: Nitrogen (N) is an important nutrient for plant growth, development, and agricultural production. Nitrogen stress could induce epigenetic changes in plants. In our research, overexpression of the OsNAR2.1 line was used as a testing target in rice plants with high nitrogen-use efficiency to study the changes of rice methylation and growth in respond of the endogenous and external nitrogen stress. RESULTS: Our results showed that external N deficiency could decrease seed N content and plant growth of the overexpression line. During the filial growth, we found that the low parent seed nitrogen (LPSN) in the overexpression line could lead to a decrease in the filial seed nitrogen content, total plant nitrogen content, yield, and OsNAR2.1 expression (28, 35, 23, and 55%, respectively) compared with high parent seed nitrogen (HPSN) in high nitrogen external supply. However, such decreases were not observed in wild type. Furthermore, methylation sequencing results showed that LPSN caused massive gene methylation changes, which enriched in over 20 GO pathways in the filial overexpression line, and the expression of OsNAR2.1 in LPSN filial overexpression plants was significantly reduced compared to HPSN filial plants in high external N, which was not shown in wild type. CONCLUSIONS: We suggest that the parent seed nitrogen content decreased induced DNA methylation changes at the epigenetic level and significantly decreased the expression of OsNAR2.1, resulting in a heritable phenotype of N deficiency over two generations of the overexpression line.

Knockdown of a Novel Gene OsTBP2.2 Increases Sensitivity to Drought Stress in Rice.

Drought stress is a major environmental stress, which adversely affects the biological and molecular processes of plants, thereby impairing their growth and development. In the present study, we found that the expression level of OsTBP2.2 which encodes for a nucleus-localized protein member belonging to transcription factor IID (TFIID) family, was significantly induced by polyethylene glycol (PEG) treatment. Therefore, knockdown mutants of OsTBP2.2 gene were generated to investigate the role of OsTBP2.2 in rice response to drought stress. Under the condition of drought stress, the photosynthetic rate, transpiration rate, water use efficiency, and stomatal conductance were significantly reduced in ostbp2.2 lines compared with wild type, Dongjin (WT-DJ). Furthermore, the RNA-seq results showed that several main pathways involved in "MAPK (mitogen-activated protein kinase) signaling pathway", "phenylpropanoid biosynthesis", "defense response" and "ADP (adenosine diphosphate) binding" were altered significantly in ostbp2.2. We also found that OsPIP2;6, OsPAO and OsRCCR1 genes were down-regulated in ostbp2.2 compared with WT-DJ, which may be one of the reasons that inhibit photosynthesis. Our findings suggest that OsTBP2.2 may play a key role in rice growth and the regulation of photosynthesis under drought stress and it may possess high potential usefulness in molecular breeding of drought-tolerant rice.

pOsNAR2.1:OsNAR2.1 expression enhances nitrogen uptake efficiency and grain yield in transgenic rice plants.

The nitrate (NO3-) transporter has been selected as an important gene maker in the process of environmental adoption in rice cultivars. In this work, we transferred another native OsNAR2.1 promoter with driving OsNAR2.1 gene into rice plants. The transgenic lines with exogenous pOsNAR2.1:OsNAR2.1 constructs showed enhanced OsNAR2.1 expression level, compared with wild type (WT), and 15 N influx in roots increased 21%-32% in response to 0.2 mm and 2.5 mm 15NO3- and 1.25 mm 15 NH415 NO3 . Under these three N conditions, the biomass of the pOsNAR2.1:OsNAR2.1 transgenic lines increased 143%, 129% and 51%, and total N content increased 161%, 242% and 69%, respectively, compared to WT. Furthermore in field experiments we found the grain yield, agricultural nitrogen use efficiency (ANUE), and dry matter transfer of pOsNAR2.1:OsNAR2.1 plants increased by about 21%, 22% and 21%, compared to WT. We also compared the phenotypes of pOsNAR2.1:OsNAR2.1 and pOsNAR2.1:OsNRT2.1 transgenic lines in the field, found that postanthesis N uptake differed significantly between them, and in comparison with the WT. Postanthesis N uptake (PANU) increased approximately 39% and 85%, in the pOsNAR2.1:OsNAR2.1 and pOsNAR2.1:OsNRT2.1 transgenic lines, respectively, possibly because OsNRT2.1 expression was less in the pOsNAR2.1:OsNAR2.1 lines than in the pOsNAR2.1:OsNRT2.1 lines during the late growth stage. These results show that rice NO3- uptake, yield and NUE were improved by increased OsNAR2.1 expression via its native promoter.