TRANSCRIPTIONAL CONTROL OF THE GLND GENE IS NOT DEPENDENT ON NITROGENAVAILABILITY IN ESCHERICHIA-COLI

Glutamine synthetase (GS) is one of the most important enzymes in the assimilation of nitrogenous compounds in Escherichia coli and related bacteria. For the control of its activity and biosynthesis, tricyclic cascades of uridylylation/deuridylylation of PII protein, adenylylatio n/deadenylylation of glutamine synthetase, and phosphorylation/dephosp horylation of Ntr1 are operating, where the regulation of uridylylatio n/deuridylylation by uridylyl transferase-uridylyl removing enzyme (UT -UR) (the product of the glnD gene) would play the ultimate nitrogen s ensing role. However, the possible nitrogen-regulatable element in the upstream of the glnD gene has been debated. In the present experiment , we have cloned and sequenced the four minute regions of the Escheric hia coli chromosome, where rpsB, map, glnD, and dapD genes have been i dentified in sequence. We could localize the transcriptional start sit e at seven nucleotides upstream of the translation initiation codon by primer extension analysis. The nitrogen dependency of the glnD gene h as been analyzed by Northern blot, RNase protection, and promoter-luci ferase activity assays. These data suggested a constitutive expression of the glnD gene independent of nitrogen availability, From these res ults, it could be concluded that the ultimate nitrogen sensing device for the bacteria should be the UT-UR itself, through modulation of its activity in response to the nitrogen status rather than its biosynthe tic mechanism.