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Inhibition of chromosome replication in Mycobacterium smegmatis: effect of the rpmH–dnaA promoter region

Laboratorio de Genética Molecular, Centro de Microbiología y Biología Celular, Instituto Venezolano de Investigaciones Científicas, Apartado 21827, Caracas 1020A, Venezuela¹

Tel: +58 2 504 1653. Fax: +58 2 504 1382. e-mail: lsalazar{at}pasteur.ivic.ve

In a previous study a functional mycobacterial origin of replication, oriC, was isolated on a plasmid. However, it was found that origin function was inhibited by the presence of the adjacent dnaA gene or its regulatory region, so that plasmids containing both of these regions next to the origin did not yield transformants. This inhibition could be due either to overexpression of dnaA on a plasmid being toxic, the transcription of dnaA into the downstream origin topologically inhibiting its function, or to the DnaA boxes upstream of dnaA somehow interacting with the DnaA boxes in the origin to prevent its function. To distinguish between these possibilities, plasmids were constructed lacking different parts of the dnaA gene: the promoter, the DnaA boxes, or both. Additionally, the putative dnaA promoter region was replaced by mycobacterial sequences that exhibit weaker or null promoter activity. The results indicate that the rpmH–dnaA promoter region, but not the DnaA boxes, is the principal cause of the incompatibility observed and suggest that this region could be playing a role in the inhibition of chromosome replication.

Keywords: oriC/replication, dnaA promoter, incompatibility, Mycobacterium

Abbreviations: FIS, factor for inversion stimulation; GFP, green fluorescent protein; IHF, integration host factor protein; OADC, oleic acid albumin dextrose complex

This article has been cited by other articles:

				L. Salazar, E. Guerrero, Y. Casart, L. Turcios, and F. Bartoli Transcription analysis of the dnaA gene and oriC region of the chromosome of Mycobacterium smegmatis and Mycobacterium bovis BCG, and its regulation by the DnaA protein Microbiology, March 1, 2003; 149(3): 773 - 784. [Abstract] [Full Text] [PDF]