MicroRNAs are 19- to 22-nucleotide small noncoding RNAs that have been implicated in abiotic stress responses. In this study, we found that knockdown of microRNA166, using the Short Tandem Target Mimic (STTM) system, resulted in morphological changes that confer drought resistance in rice (). From a large-scale screen for miRNA knockdown lines in rice, we identified miR166 knockdown lines (STTM166); these plants exhibit a rolled-leaf phenotype, which is normally displayed by rice plants under drought stress. The leaves of STTM166 rice plants had smaller bulliform cells and abnormal sclerenchymatous cells, likely causing the rolled-leaf phenotype. The STTM166 plants had reduced stomatal conductance and showed decreased transpiration rates. The STTM166 lines also exhibited altered stem xylem and decreased hydraulic conductivity, likely due to the reduced diameter of the xylem vessels. Molecular analyses identified rice (), a member of HD-Zip III gene family, as a major target of miR166; moreover, rice plants overexpressing a miR166-resistant form of resembled the STTM166 plants, including leaf rolling and higher drought resistance. The genes downstream of miR166- consisted of polysaccharide synthesis-related genes that may contribute to cell wall formation and vascular development. Our results suggest that drought resistance in rice can be increased by manipulating miRNAs, which leads to developmental changes, such as leaf rolling and reduced diameter of the xylem, that mimic plants' natural responses to water-deficit stress.

Leafy heads of cabbage (Brassica oleracea), Chinese cabbage (Brassica rapa), and lettuce (Lactuca sativa) are composed of extremely incurved leaves. The shape of these heads often dictates the quality, and thus the commercial value, of these crops. Using quantitative trait locus mapping of head traits within a population of 150 recombinant inbred lines of Chinese cabbage, we investigated the relationship between expression levels of microRNA-targeted Brassica rapa ssp. pekinensis TEOSINTE BRANCHED1, cycloidea, and PCF transcription factor4 (BrpTCP4) genes and head shape. Here, we demonstrate that a cylindrical head shape is associated with relatively low BrpTCP4-1 expression, whereas a round head shape is associated with high BrpTCP4-1 expression. In the round-type Chinese cabbage, microRNA319 (miR319) accumulation and BrpTCP4-1 expression decrease from the apical to central regions of leaves. Overexpression of BrpMIR319a2 reduced the expression levels of BrpTCP4 and resulted in an even distribution of BrpTCP4 transcripts within all leaf regions. Changes in temporal and spatial patterns of BrpTCP4 expression appear to be associated with excess growth of both apical and interveinal regions, straightened leaf tips, and a transition from the round to the cylindrical head shape. These results suggest that the miR319a-targeted BrpTCP gene regulates the round shape of leafy heads via differential cell division arrest in leaf regions. Therefore, the manipulation of miR319a and BrpTCP4 genes is a potentially important tool for use in the genetic improvement of head shape in these crops.Chinese cabbage (Brassica rapa), cabbage (Brassica oleracea), and lettuce (Lactuca sativa) have uniform, tightly formed heads composed of yellow-green, crinkly leaves. These leaves serve as an important source of mineral nutrients, crude fiber, and vitamins in the human diet. Head shape can vary depending on the cultivar; the two most common shapes are round and cylindrical, with others often being oblong or cone like. Chinese cabbage is commonly sold by head shape and color rather than by individual varieties. The favorable shape often changes according to current consumer preference.

HYL1 (HYPONASTIC LEAVES 1) in Arabidopsis thaliana encodes a double-stranded RNA-binding protein needed for proper miRNA maturation, and its null mutant hyl1 shows a typical leaf-incurvature phenotype. In Chinese cabbage, BcpLH (Brassica rapa ssp. pekinensis LEAFY HEADS), a close homolog of HYL1, is differentially expressed in juvenile leaves, which are flat, and in adult leaves, which display extreme incurvature. BcpLH lacks protein–protein interaction domains and is much shorter than HYL1. To test whether BcpLH is associated with defects in microRNA (miRNA) biogenesis and leaf flatness, we enhanced and repressed the activity of BcpLH by transgenics and investigated BcpLH-dependent miRNAs and plant morphology. BcpLH promoted miRNA biogenesis by the proper processing of primary miRNAs. BcpLH downregulation via antisense decreased a specific subset of miRNAs and increased the activities of their target genes, causing upward curvature of rosette leaves and early leaf incurvature, concurrent with the enlargement, earliness, and round-to-oval shape transition of leafy heads. Moreover, BcpLH-dependent miRNAs in Chinese cabbage are not the same as HYL1-dependent miRNAs in Arabidopsis. We suggest that BcpLH controls a specific subset of miRNAs in Chinese cabbage and coordinates the direction, extent, and timing of leaf curvature during head formation in Brassica rapa.