Expression of bioluminescence in the marine bacterium Vibrio fischeri is controlled by a unique cell density-dependent regulatory mechanism called auto-induction. The genes required for bioluminescence (the lux genes) are organized in two divergently transcribed operons (luxR-luxICDABEG). One operon (luxICDABEG) contains the genes required for light production (luxCDABE) and the synthesis of a diffusible signal molecule called autoinducer (luxI). The other operon contains the luxR gene which encodes a transcriptional regulatory protein that activates transcription of both lux operons in the presence of autoinducer. This bidirectional stimulatory mechanism leads to a positive feedback circuit that results in a rapid increase in light production at a particular culture cell density which is characteristic of autoinduction. Transcriptional positive feedback is apparently limited by a negative autoregulatory circuit through which LuxR acts to inhibit its own synthesis. Transcriptional negative autoregulation requires autoinducer, the lux operator located in the control region (which is the binding site for LuxR), and negative acting DNA sequences in the luxICDABEG operon. Deletion analysis of the luxICDABEG operon demonstrated that a negative acting element is located in the luxD gene at a position 2.0 kilobases from the lux operator. The nucleotide sequence of this luxD element is similar to the lux operator (11 of 20 base pairs identical) and can function as a LuxR-binding site when it replaces the lux operator in the control region. These results suggest that the luxD element functions as a low affinity binding site for LuxR and that occupancy of this site is required to achieve transcriptional negative autoregulation of luxR.