An AP2/ERF transcription factor controls generation of the twin-seedling rice.

Introduction: The floret of rice is a main component of the spikelet, and the ovule of pistil is a critical organ for successful reproduction and determines the number of seeds. However, the molecular mechanisms underlying the ovule development remain elusive.

Objective: Twin-seedling rice has great potential for application in rice production. The study was to isolate the gene that controls twin-seedling in rice and explore the molecular function of the gene in floret development.

Methods: We discovered a twin-seedling rice (tsr) mutant and constructed different segregating populations to clone TSR gene using map-based cloning method. To explore the molecular functions of TSR in determination of the ovary number and development, we applied molecular technologies such as yeast two-hybrid assay, electrophoretic mobility shift assay (EMSA), and dual-LUC transient expression assay to search for the TSR-interacting proteins and the target genes regulated by TSR.

Results: We report here the map-based cloning of TSR which encodes an AP2/ERF transcription factor. Mutations in TSR lead to occurrence of the twin-seedling rice. The tsr mutant showed open hulls of the spikelets and displayed changes in the number of stamens and ovules of the florets. The ovary of tsr mutant contained two conjugated ovules which developed into a mature seed with two viable embryos. Mechanistic studies revealed that TSR regulates the expression levels of genes related to spikelet determination and ovule identity by binding to their promoters. Furthermore, TSR interacted with OsMADS1 and this interaction allowed OsMADS1 to modulate the transcriptional activityy of TSR on gene expression. The molecular study of TSR provides new insights into the formation and development of rice floret and helps breeders generate twin-seedling rice in production.