In Rhizobium meliloti, nif and fix genes, involved in nitrogen fixation during symbiosis with alfalfa, are under the control of two transcriptional regulators encoded by nifA and fixK. Expression of nifA and fixK is under the control of FixL/J, a two-component regulatory system. We showed, using Escherichia coli as a heterologous host, that FixL/J controls nifA and fixK expression in response to microaerobiosis. Furthermore, expression of the sensor gene fixL and of the activator gene fixJ under the control of two different promoters allowed us to show that FixL mediates microaerobic induction of nifA when the level of FixJ is low and aerobic repression of nifA when the level of FixJ is high. Similarly, activation of fixK occurred in microaerobiosis when the FixJ level was low in the presence of FixL. In contrast to nifA, fixK expression was not affected by FixL in aerated cultures when the level of FixJ was high. We conclude that R. meliloti FixL senses oxygen in the heterologous host E. coli consistent with the microaerobic induction of nifA and fixK in R. meliloti and that nifA and fixK promoters are differentially activated by FixJ in response to the oxygen signal.