BELL1-like homeobox genes regulate inflorescence architecture and meristem maintenance in rice.

Inflorescence architecture is diverse in angiosperms, and is mainly determined by the arrangement of the branches and flowers, known as phyllotaxy. In rice (Oryza sativa), the main inflorescence axis, called the rachis, generates primary branches in a spiral phyllotaxy, and flowers (spikelets) are formed on these branches. Here, we have studied a classical mutant, named verticillate rachis (ri), which produces branches in a partially whorled phyllotaxy. Gene isolation revealed that RI encodes a BELL1-type homeodomain transcription factor, similar to Arabidopsis PENNYWISE/BELLRINGER/REPLUMLESS, and is expressed in the specific regions within the inflorescence and branch meristems where their descendant meristems would soon initiate. Genetic combination of an ri homozygote and a mutant allele of RI-LIKE1 (RIL1) (designated ri ril1/+ plant), a close paralog of RI, enhanced the ri inflorescence phenotype, including the abnormalities in branch phyllotaxy and rachis internode patterning. During early inflorescence development, the timing and arrangement of primary branch meristem (pBM) initiation were disturbed in both ri and ri ril1/+ plants. These findings suggest that RI and RIL1 were involved in regulating the phyllotactic pattern of the pBMs to form normal inflorescences. In addition, both RI and RIL1 seem to be involved in meristem maintenance, because the ri ril1 double-mutant failed to establish or maintain the shoot apical meristem during embryogenesis.

Generation of artificial drooping leaf mutants by CRISPR-Cas9 technology in rice.

CRISPR-Cas9 technology, which uses an RNA-guided nuclease, has been developed as an efficient and versatile genome-editing method to induce mutations in genes of interest. To examine the feasibility of this method in developmental studies of a model monocot, rice (Oryza sativa), we introduced the construct gDL-1, which produced a guide RNA targeting the DROOPING LEAF (DL) gene. DL regulates midrib formation in the leaf and carpel specification in the flower. Because loss of function of DL causes the drooping leaf phenotype in regenerated seedlings, the effect of gene disruption should be easily detected. In transgenic plants carrying gDL-1, the DL gene was disrupted at high efficiency: seven out of nine plants examined had bi-allelic mutations. All transgenic plants with the bi-allelic mutation showed the drooping leaf phenotype. Observation of cross sections of the leaf blade clearly indicated that these transgenic plants failed to make midrib structures, and were comparable to the severe dl mutant dl-sup1. Thus, CRISPR-Cas9 technology can be a useful and efficient tool in developmental studies in rice.