Following our group's discovery (1), cloning (2), genetic characterization (3) and nucleotide sequence determination (4) of the E.coli proU operon, our interest has been in understanding its transcriptional regulation. proU transcription is induced 400-fold during growth at high osmolarity, which is the highest known for an osmoresponsive gene in any organism. Three cis elements for proU regulation have been identified (5), namely two promoters P1 and P2 and a negative regulatory element (NRE) of approximately 500 bp that is situated downstream of the promoters (overlapping the first structural gene). The P1 and P2 promoters are transcribed by RNA polymerase bearing the s^s- and s⁷⁰- subunits respectively (6,7). The nucleoid protein H-NS is involved in repressing proU expression at low osmolarity at least in part by interacting with the NRE (5,7), but this does not appear to be through a 'silencing' mechanism (8). Our current interests are in understanding (i) the contribution of the P1 promoter to proU regulation, particularly in light of the finding (9) that this promoter is cryptic in the closely related bacterium Salmonella typhimurium because of a novel phenomenon of transcription attenuation; and (ii) the mechanism by which the NRE mediates the repressive action of H-NS, and in particular the topological changes occurring at the NRE during growth at high osmolarity (5). We have recently shown that P1 promoter transcription is modulated by the transcription termination factor Rho as well as by H-NS, and that the promoter may be involved in proU expression during growth at low temperatures (10).

We have also developed and patented a proU promoter-based generic system for salt-induced overproduction of recombinant proteins in E.coli (11,12) and the strain developed for the purpose is sold by Life Technologies Inc., USA as BL21-SI (13). Studies are ongoing in our group to generate improvements to this process.

1. Gowrishankar, J. 1985. Identification of osmoresponsive genes in Escherichia coli: evidence for participation of potassium and proline transport systems in osmoregulation. J. Bacteriol. 164: 434-445.
2. Gowrishankar, J., P. Jayashree, and K. Rajkumari. 1986. Molecular cloning of an osmoregulatory locus in Escherichia coli: increased proU gene dosage results in enhanced osmotolerance. J. Bacteriol. 168: 1197-1204.
3. Dattananda, C.S., and J. Gowrishankar. 1989. Osmoregulation in Escherichia coli: complementation analysis and gene-protein relationships in the proU locus. J. Bacteriol. 171: 1915-1922.
4. Gowrishankar, J. 1989. Nucleotide sequence of the osmoregulatory proU operon of Escherichia coli. J. Bacteriol. 171: 1923-1931. Erratum. 172: 1165.
5. Dattananda, C.S., K. Rajkumari, and J. Gowrishankar. 1991. Multiple mechanisms contribute to osmotic inducibility of proU operon expression in Escherichia coli: demonstration of two osmoresponsive promoters and of a negative regulatory element within the first structural gene. J. Bacteriol. 173: 7481-7490.
6. Manna, D., and J. Gowrishankar. 1994. Evidence for involvement of proteins HU and RpoS in transcription of the osmoresponsive proU operon in Escherichia coli. J. Bacteriol. 176: 5378-5384.
7. Rajkumari, K., S. Kusano, A. Ishihama, T. Mizuno, and J. Gowrishankar. 1996. Effects of H-NS and potassium glutamate on s S- and s 70-directed transcription in vitro from osmotically regulated P1 and P2 promoters of proU in Escherichia coli. J. Bacteriol. 178: 4176-4181.
8. Gowrishankar, J., and A.J. Pittard. 1998. Superimposition of TyrR protein-mediated regulation on osmoresponsive transcription in vivo of proU in Escherichia coli. J. Bacteriol. 180: 6743-6748.
9. Rajkumari, K., A. Ishihama, and J. Gowrishankar. 1997. Evidence for transcription attenuation rendering cryptic a s s-dependent promoter of the osmotically regulated proU operon of Salmonella typhimurium. J. Bacteriol. 179:7169-7173.
10. Rajkumari, K., and J. Gowrishankar. 2001. In vivo expression from the RpoS-dependent P1 promoter of the osmotically regulated proU operon in Escherichia coli and Salmonella enterica serovar Typhimurium: activation by rho and hns mutations, and by cold stress. (submitted).
11. Bhandari, P., and J. Gowrishankar. 1997. An Escherichia coli host strain useful for efficient overproduction of cloned gene products with NaCl as the inducer. J. Bacteriol. 179: 4403-4406.
12. Gowrishankar, J., P. Bhandari, and K. Rajkumari.1998. Process for producing polypeptides. United States Patent 5,830,690.
13. Donahue R. A. Jr., and R. L. Bebee 1999. BL21-SI competent cells for protein expression in E.coli. Focus 21:49-51.