Strategies to increase rice productivity to meet the global demand have been the main concern of breeders around the world. Although a growing number of functional genes related to crop yield have been characterized, our understanding of its associated regulatory pathways is limited. Using rice as a model, we find that blocking miR396 greatly increases grain yield by modulating development of auxiliary branches and spikelets through direct induction of the growth regulating factor 6 (OsGRF6) gene. The upregulation of OsGRF6 results in the coordinated activation of several immediate downstream biological clades, including auxin (IAA) biosynthesis, auxin response factors, and branch and spikelet development-related transcription factors. This study describes a conserved microRNA (miRNA)-dependent regulatory module that integrates inflorescence development, auxin biosynthesis and signalling pathways, and could potentially be used in engineering high-yield crop plants.
As fundamental nutrients, amino acids are important for rice (Oryza sativa) growth and development. Here, we identified the amino acid permease 5 (OsAAP5), that regulates tiller number and grain yield in rice. The OsAAP5 promoter sequence differed between indica and japonica rice varieties. Lower expression of OsAAP5 in the young leaf blade in indica varieties than in japonica varieties was associated with more tillers in indica than in japonica. Down-regulation of OsAAP5 expression in japonica using RNA interference (RNAi) and clustered regularly interspaced short palindromic repeats led to increases in tiller number and grain yield, whereas OsAAP5 overexpression (OE) had the opposite effect. Both a protoplast amino acid uptake assay and HPLC analysis indicated that more basic (Lys, Arg) and neutral (Val, Ala) amino acids were transported and accumulated in the OE lines than in the wild type, but the opposite was observed in the RNAi lines. Furthermore, exogenous application of Lys, Arg, Val, and Ala in the OE lines substantially inhibited tiller bud elongation, but the effect was lost in the RNAi lines. Notably, concentrations of the cytokinins cis-zeatin and dihydrozeatin were much lower in the OE lines than in the wild type, whereas concentrations in the RNAi lines were higher. Thus, OsAAP5 could regulate tiller bud outgrowth by affecting cytokinin levels, and knockout of OsAAP5 could be valuable for japonica breeding programs seeking high yield and grain quality.
Fitness cost is a common phenomenon in rice blast disease-resistance breeding. MiR396 is a highly conserved microRNA (miRNA) family targeting Growth Regulating Factor (OsGRF) genes. Mutation at the target site of miR396 in certain OsGRF gene or blocking miR396 expression leads to increased grain yield. Here we demonstrated that fitness cost can be trade-off in miR396-OsGRFs module via balancing growth and immunity against the blast fungus. The accumulation of miR396 isoforms was significantly increased in a susceptible accession, but fluctuated in a resistant accession upon infection of Magnaporthe oryzae. The transgenic lines over-expressing different miR396 isoforms were highly susceptible to M. oryzae. In contrast, overexpressing target mimicry of miR396 to block its function led to enhanced resistance to M. oryzae in addition to improved yield traits. Moreover, transgenic plants overexpressing OsGRF6, OsGRF7, OsGRF8, and OsGRF9 exhibited enhanced resistance to M. oryzae, but showed different alteration of growth. While overexpression of OsGRF7 led to defects in growth, overexpression of OsGRF6, OsGRF8, and OsGRF9 resulted in better or no significant change of yield traits. Collectively, our results indicate that miR396 negatively regulates rice blast disease- resistance via suppressing multiple OsGRFs, which in turn differentially control growth and yield. Therefore, miR396-OsGRFs could be a potential module to demolish fitness cost in rice blast disease-resistance breeding.
High fluorescence quantum yield graphene quantum dots (GQDs) have showed up as a new generation for bioimaging. In this work, luminescent GQDs were prepared by an ameliorative photo-Fenton reaction and a subsequent hydrothermal process using graphene oxide sheets as the precursor. The as-prepared GQDs were nanomaterials with size ranging from 2.3 to 6.4 nm and emitted intense green luminescence in water. The fluorescence quantum yield was as high as 24.6% (excited at 340 nm) and the fluorescence was strongest at pH 7. Moreover, the influences of low-concentration (12.5, 25 μg/mL) GQDs on the morphology, viability, membrane integrity, internal cellular reactive oxygen species level and mortality of HeLa cells were relatively weak, and the in vitro imaging demonstrated GQDs were mainly in the cytoplasm region. More strikingly, zebrafish embryos were co-cultured with GQDs for in vivo imaging, and the results of heart rate test showed the intake of small amounts of GQDs brought little harm to the cardiovascular of zebrafish. GQDs with high quantum yield and strong photoluminescence show good biocompatibility, thus they show good promising for cell imaging, biolabeling and other biomedical applications.
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