Phage-Borne Factors and Host LexA Regulate the Lytic Switch in Phage GIL01

Abstract

The Bacillus thuringiensis temperate phage GIL01 does not integrate into the host chromosome but exists stably as an independent linear replicon within the cell. Similar to that of the lambdoid prophages, the lytic cycle of GIL01 is induced as part of the cellular SOS response to DNA damage. However, no CI-like maintenance repressor has been detected in the phage genome, suggesting that GIL01 uses a novel mechanism to maintain lysogeny. To gain insights into the GIL01 regulatory circuit, we isolated and characterized a set of 17 clear plaque (cp) mutants that are unable to lysogenize. Two phage-encoded proteins, gp1 and gp7, are required for stable lysogen formation. Analysis of cp mutants also identified a 14-bp palindromic dinBox1 sequence within the P1-P2 promoter region that resembles the known LexA-binding site of Gram-positive bacteria. Mutations at conserved positions in dinBox1 result in a cp phenotype. Genomic analysis identified a total of three dinBox sites within GIL01 promoter regions. To investigate the possibility that the host LexA regulates GIL01, phage induction was measured in a host carrying a noncleavable lexA (Ind ؊ ) mutation. GIL01 formed stable lysogens in this host, but lytic growth could not be induced by treatment with mitomycin C. Also, mitomycin C induced ␤-galactosidase expression from GIL01-lacZ promoter fusions, and induction was similarly blocked in the lexA (Ind ؊ ) mutant host. These data support a model in which host LexA binds to dinBox sequences in GIL01, repressing phage gene expression during lysogeny and providing the switch necessary to enter lytic development.

Occurrence of Bacillus thuringiensis and their phages in Yemen soil

“…Results indicated that, phage active against Bt may be present in soil or integrated with host genome (lysogeny) or as independent linear replica within the cell. These results are in accordance with finding of [19,26,27,41,43,48]. Electron microscopic studies of the isolated phages (Fig.…”

Gene transcription from the linear plasmid pBClin15 leads to cell lysis and extracellular DNA-dependent aggregation of Bacillus cereus ATCC 14579 in response to quinolone-induced stress

“…4) and the genes of pBClin15 appear to be under similar transcriptional regulation to the genes of prophage GIL01 (Fig. 5) (Fornelos et al, 2011).…”

“…Functional tectiviral phages infecting B. thuringiensis as well as B. anthracis have previously been identified, which display similar genome organization and share relatively high sequence similarity at the DNA level to the B. cereus plasmid pBClin15 (Fornelos et al, 2011;Strömsten et al, 2003;Verheust et al, 2003Verheust et al, , 2005. Although it has never been shown that pBClin15 encodes a functional phage, it still exhibits properties of a prophage.…”

“…Regulation of lysogeny in the temperate B. thuringiensis phage GIL01 has been shown to require the host LexA transcriptional regulator as well as phage-specific factors (Fornelos et al, 2011). The transcriptional organization of GIL01 included two functional modules: a regulatory unit consisting of ORF1-8 that was controlled by the identified P1 and P2 promoters localized upstream of ORF1, and a module composed of structural and lytic genes ORF9-30 controlled by a third promoter P3.…”

“…ORF5 codes for a DNA polymerase, whereas ORF6 encodes a LexA-like transcriptional repressor. ORF15 has been predicted to be a capsid protein, whilst ORF9 and ORF12 may be involved in capsid assembly and DNA packaging, respectively (Fornelos et al, 2011;Strömsten et al, 2003;Verheust et al, 2003Verheust et al, , 2005.…”