The effect of γ-irradiation on Mu DNA transposition and gene expression

Abstract

Mobile genetic elements are a ubiquitous presence in the genomes of a well-studied organisms. The effect of genomic stress on the status and transposition of these elements has not, as yet, been extensively characterized. We have been using temperate, transposable bacteriophage Mu as a model system to examine the behavior of mobile genetic elements and have previously shown that many DNA-damaging agents did not induce a Mu prophage to enter the lytic cycle of multiple rounds of DNA transposition. To extend these results and to examine the possibility that they were a reflection of damage to the DNA substrate for Mu transposition, we have constructed a mini-Mu plasmid, pMD12, which contains the early region of Mu, flanked by both extremities required for transposition in cis, and the beginning of the transposase gene A fused in frame to the lacZ gene. This A′-lacZ fusion protein maintains β-galactosidase enzymatic activity under the control of the expression of the Mu transposase A gene and thus, the capacity for Mu transposition can be easily monitored by assaying for β-galactosidase. By measuring the amount of β-galactosidase after various doses of γ-irradiation, we found that doses of up to 75 krad had no effect on the expression of the Mu transposase gene A. This was confirmed by the lack of induction of a Mu prophage in strains containing a chromosomally inserted Mu genome. Although the plaque-forming units per colony-forming unit of strain CSH67, containing a chromosomally inserted λ prophage, increased approximately 100-fold from 0 to 75 krad, no stimulation of induction of prophage Mu lytic growth was observed. We also found that plasmid pMD12 did not transpose and chromosomally associate upon γ-irradiation. This supports the assertion that DNA-damaging agents, including γ-rays, do not induce the transposition of prokaryotic mobile genetic elements.