PINOID and PIN-FORMED Paralogous Genes Are Required for Leaf Morphogenesis in Rice.

Auxin plays an essential role in modulating leaf development. However, its role in leaf development in rice (Oryza sativa L.) remains largely unknown. In this study, we found that PINOID (OsPID) and two Sister-of-PIN1s, termed PIN-FORMED1c (OsPIN1c) and OsPIN1d, are necessary for rice leaf development. The ospin1c ospin1d null mutant lines presented severe defects in leaf morphogenesis, including drooping and semi-drooping blades, an abnormally thickened sheath and lamina joint, and fused leaves with absent ligules and auricles. Loss-of-function ospid mutants displayed generally similar leaf morphology but lacked leaf fusion. Interestingly, misshaped leaf genesis displayed a preference for being ipsilateral. In addition, OsPIN1c and OsPID were commonly localized in the initiating leaf primordia. Furthermore, accompanied by the more severe organ morphogenesis in the ospin1c ospin1d ospid triple mutant, RNA sequencing analysis revealed that many genes essential for leaf development have an altered expression level. Together, this study furthers our understanding of the role auxin transport plays during leaf development in monocot rice.

SWI2/SNF2 chromatin remodeling ATPases SPLAYED and BRAHMA control embryo development in rice.

KEY MESSAGE: Chromatin remodeling ATPases OsSYD and OsBRM are involved in shoot establishment, and both affect OSH gene transcription. OsSYD protein interacts with RFL, but OsBRM does not. In plants, SPLAYED (SYD) and BRAHMA (BRM) encode chromatin remodeling ATPases that use the energy derived from ATP hydrolysis to restructure nucleosomes and render certain genomic regions available to transcription factors. However, the function of SYD and BRM on rice growth and development is unknown. Here, we constructed ossyd and osbrm mutants using CRISPR/Cas9 technology and analyzed the effects of mutations on rice embryo development. We discovered that the ossyd and osbrm mutants exhibited severe defects during embryonic development, whereas endosperm development was normal. These results indicated that the development of the embryo and endosperm is independent of each other. Consequently, the ossyd- and osbrm-null mutants did not germinate due to the abnormal embryos. Furthermore, we observed the embryos of ossyd- and osbrm-null mutants, and they indeed had distinct differentiation defects in shoot establishment, acquired during embryogenesis. To verify the function of OsSYD and OsBRM in embryogenesis, we measured the transcript levels of marker genes at different stages. Compared with wild type, the expression levels of multiple OSH genes were significantly reduced in the mutants, which was consistent with the defective shoot establishment phenotypes. The interaction between SYD and RICE FLORICAULA/LFY (RFL) was revealed using a yeast two-hybrid screening system, suggesting that the interaction between the LFY homolog and chromatin remodeling ATPases is ubiquitous in plants. Collectively, our findings provide the basis for elucidating the function of OsSYD and OsBRM during embryo development in rice.

OsmiR164-targeted OsNAM, a boundary gene, plays important roles in rice leaf and panicle development.

The CUP-SHAPED COTYLEDON (CUC) genes (CUC1, CUC2 and CUC3) regulate organ boundary formation in Arabidopsis. However, the functions of their homologous genes in rice (Oryza sativa) are still unknown. Here, we have identified an orthologous gene of CUC1 and CUC2 in rice, named OsNAM. Subcellular localization and yeast two-hybrid assay results have suggested that OsNAM encodes a conserved nuclear NAC (NAM/ATAF1/CUC2) protein with a transcriptional activator. The null mutant osnam-1 presented a fused leaf structure, small panicles, reduced branches and aberrant floral organ identities when compared with those of the wild type. Beta-glucuronidase staining and GFP reporter lines indicated that OsNAM was expressed in young tissues and that its boundary enrichment expression was regulated by OsmiR164. Loss-of-function mutants for OsCUC3 resulted in no obvious defects throughout rice development. The osnam oscuc3 double mutant, however, resulted in severe leaf fusion of the first two leaves, while the osnam single mutant showed a similar phenotype from the seventh leaf. These results indicated that OsNAM and OsCUC3 act redundantly for boundary specification during post-embryonic development. Overall, we describe the biological functions of OsNAM and OsCUC3 in rice development and the expression characteristics of OsNAM. This work reveals the important role of CUC genes in rice.