Gibberellins (GAs) are phytohormones that regulate growth and development throughout the life cycle of plants. RSG (REPRESSION OF SHOOT GROWTH) is a tobacco (Nicotiana tabacum) transcriptional activator with a basic leucine zipper domain that regulates the endogenous amount of GAs by control of GA biosynthetic enzymes. Negative feedback contributes to homeostasis of the GA levels. Previous studies suggested that RSG is directly or indirectly involved in the GA negative feedback of NtGA20ox1 encoding GA 20-oxidase. Using transgenic tobacco plants, we have identified a cis-acting region that is responsible for the feedback regulation of NtGA20ox1. This region contains an RSG-binding sequence. A mutation in the RSG-binding sequence abolished negative feedback of NtGA20ox1 in transgenic plants. Chromatin immunoprecipitation (ChIP) assays showed that RSG binds to the NtGA20ox1 promoter in vivo in response to a decrease in GA levels, and that this binding is abolished within 3 h after GA treatment. Furthermore, decreases in GA levels promote modifications of active histone marks in the promoter of NtGA20ox1. Our results suggest that RSG plays a role in the homeostasis of GAs through direct binding to the NtGA20ox1 promoter.