Important functions of R-loop structures(RNA:DNA hybrids)had been revealed in many biological processes among diverse organisms.DNA Topoisomerase 1(TOP1),an essential manipulator for DNA topology in RNA transcription and DNA replication processes,can prevent R-loops accumulation by removing the positive and negative DNA supercoiling that are made by RNA polymerases during transcription.TOP1 is required for plant development,but little is known about its function in cotranscriptional R-loops resolving in different plant biological processes.Here we show that knockdown of OsTop1 has strong effects on rice development,including defects of root architecture and gravitropism which are due to genes misregulated in auxin signaling.We find that R-loops are naturally formed over rice auxin related gene loci like OsARF19 and OsMDR14,and accumulated by either knocking down OsTop1,or blocking its protein activity.OsTop1 therefore sets the accurate expression levels of auxin signaling gene network by limiting inherent R-loops over-accumulation,and thus controlling the root growth and gravitropism.Our data has revealed R-loops as the important components in polar auxin transport and plant root development,and highlighted that OsTop1 functions as the key to link transcriptional R-loops resolving and plant hormone signaling.This work extends our view on transcriptional regulation of plant hormone signaling.