The CI repressor of bacteriophage is a model for the role of cooperativity in the efficient functioning of genetic switches. Pairs of CI dimers interact to cooperatively occupy adjacent operator sites at O R and at O L . These CI tetramers repress the lytic promoters and activate transcription of the cI gene from P RM . CI is also able to octamerize, forming a large DNA loop between O R and O L , but the physiological role of this is unclear. Another puzzle is that, although a dimer of CI is able to repress P RM by binding to the third operator at O R , O R 3, this binding seems too weak to affect CI production in the lysogenic state. Here we show that repression of P RM at lysogenic CI concentrations is absolutely dependent on O L , in this case 3.8 kb away. A mutant defective in this CI negative autoregulation forms a lysogen with elevated CI levels that cannot efficiently switch from lysogeny to lytic development. Our results invalidate previous evidence that Cro binding to O R 3 is important in prophage induction. We propose the octameric CI:O R -O L complex increases the affinity of CI for O R 3 by allowing a CI tetramer to link O R 3 and the third operator at O L , O L 3.