RecA expression and DNA damage in mycobacteria

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RecA expression and DNA damage in mycobacteria

Thomas, Nicola Alison; (1998) RecA expression and DNA damage in mycobacteria. Doctoral thesis (Ph.D), UCL (University College London). Green open access

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Abstract

Bacterial responses to DNA damage are highly conserved. One system, the SOS response, involves the co-ordinately induced expression of over 20 unlinked genes. The RecA protein plays a central role in the regulation of this response via its interaction with a repressor protein, LexA. The recA gene of Mycobacterium tuberculosis has previously been cloned and sequenced (Davis et al, 1991) and a LexA homologue has recently been identified (Movahedzadeh, 1996). The intracellular lifestyle of pathogenic mycobacteria constantly exposes them to hostile agents such as hydrogen peroxide. Consequently, the response of mycobacteria to DNA damage is of great interest. In this study the basal levels of RecA protein were shown to be elevated in M. microti and M. smegmatis compared to those found in M. tuberculosis, M. bovis BCG and a wide range of both saprophytic and pathogenic mycobacteria. Expression of this protein was shown to be inducible in both M. smegmatis and M. tuberculosis in response to DNA damaging agents, although the kinetics of induction were quite different. A reduction in the levels of LexA following treatment of M. smegmatis, M. tuberculosis and M. microti with a DNA damaging agent, probably as a result of LexA cleavage, provided further corroboratory evidence to suggest that the essential regulatory elements of the SOS response may have been conserved. There was no evidence of RecA induction in M. microti. Sequencing of the M. microti recA gene and 700bp of upstream sequence revealed a high degree of homology with equivalent sequences in M tuberculosis', including a putative LexA binding site. M. microti recA was shown to be inducible in M. smegmatis mc2 155 in response to DNA damage using a transcriptional fusion of the putative regulatory elements of this gene to the reporter gene chloramphenicol-acetyl transferase (CAT). Thus it appears that the kinetics of induction are controlled at the level of the host cell; this was further investigated by studying the interaction of purified mycobacterial LexA protein and M. microti cell-free extracts with the putative LexA binding site.

Type:	Thesis (Doctoral)
Qualification:	Ph.D
Title:	RecA expression and DNA damage in mycobacteria
Open access status:	An open access version is available from UCL Discovery
Language:	English
Additional information:	Thesis digitised by ProQuest.
Keywords:	Biological sciences; RecA protein
URI:	https://discovery.ucl.ac.uk/id/eprint/10102158