Functional Analysis Of HANABA TARANU In Arabidopsis And Cucumber

All the aerial organs are originated from the shoot apical meristem (SAM) in plant. Organ primordia are initiated from the peripheral zone of the SAM and develop into leaves during the vegetative stage, and into flowers during the reproductive phase. Between the meristem and the newly formed organ primordia, a boundary with specialized cells is formed that separates meristematic activity from determinate organ growth. Arabidopsis HANABA TARANU(HAN) encodes a GATA3 family transcription factor that is specifically expressed at the boundaries between meristem and floral organ primordia and at the boundaries of different floral whorls. HAN functions in floral organ development, SAM organization and embryo development in Arabidopsis, whereas HAN homologs in the grass species such as rice and maize are involved in repressing bract outgrowth and promoting branching.Despite interactions that have been found between boundary genes with regulators controlling meristem maintenance or primordia development, most researches were performed during embryogenesis or vegetative growth, little is known about whether and how boundaries communicate with meristem and organ primordia during the reproductive phase. Combining genetic, molecular and biochemical tools, we explored interactions between the boundary-expressing HAN and two meristem regulators BREVIPEDICELLUS(BP) and PINHEAD(PNH), and three primordia-specific genes PETAL LOSS(PTL), JAGGED(JAG) and BLADE-ON-PETIOLE(BOP) during flower development. Our data demonstrated the key role of HAN in determining petal number. HAN and PNH transcriptionally promote each other, and biochemically combine to regulate meristem organization. HAN directly stimulates the expression of JAG and BOP2, and physically interacts with JAG to regulate floral organ development. Besides, HAN directly binds to the promoter and intron of CYTOKININ OXIDASE 3 (CKX3) to mediate cytokinin homeostasis in the boundary. Our data suggest that boundary-expressing HAN communicates with the meristem through the PNH, regulates floral organ development via JAG and BOP2, and maintains boundary morphology through CKX3 during flower development in Arabidopsis.Cucumber(Cucumis sativus L.) is a globally cultivated vegetable that is of important economic and nutritional value. Cucumber cultivation area and production in China rank first in the world. Through homology cloning, we isolated the HAN homolog CsHAN1 in cucumber. CsHAN1 is predominantly expressed at the junction of SAM and the stem, and can partially rescue the han-2 floral organ phenotype in Arabidopsis. Overexpression and RNAi of CsHANl transgenic cucumber displayed retarded growth early since embryogenesis and produced highly lobed leaves. In situ and qRT-PCR data show that the expression of meristem genes WUSCHEL(WUS) and SHOOT MERISTEMLESS(STM) were greatly reduced in the meirstem of CsHAN1 RNAi lines, and multiple leaf developmental genes were altered in the lobed leaves in CsHAN1 transgenic cucumber. Therefore, CsHAN1 may regulate SAM development through regulating the WUS and STM pathways, and modulate leaf development through a complex gene regulatory network incucumber.As such, this study uncovered the different biological functions and molecular mechanisms of boundary gene HAN in Arabidopsis and cucumber, which enriched the functional charicterization of boundary genes and boundary regions in plant, and provided the molecular guidance for breeding of new cucumber varieties with optimum plant architecture.