Lysis Genes of the Bacillus subtilis Defective Prophage PBSX

Abstract

Four genes identified within the late operon of PBSX show characteristics expected of a host cell lysis system; they arexepA, encoding an exported protein; xhlA, encoding a putative membrane-associated protein; xhlB, encoding a putative holin; and xlyA, encoding a putative endolysin. In this work, we have assessed the contribution of each gene to host cell lysis by expressing the four genes in different combinations under the control of their natural promoter located on the chromosome of Bacillus subtilis 168. The results show thatxepA is unlikely to be involved in host cell lysis. Expression of both xhlA and xhlB is necessary to effect host cell lysis of B. subtilis. Expression ofxhlB (encoding the putative holin) together withxlyA (encoding the endolysin) cannot effect cell lysis, indicating that the PBSX lysis system differs from those identified in the phages of gram-negative bacteria. Since host cell lysis can be achieved when xlyA is inactivated, it is probable that PBSX encodes a second endolysin activity which also uses XhlA and XhlB for export from the cell. The chromosome-based expression system developed in this study to investigate the functions of the PBSX lysis genes should be a valuable tool for the analysis of other host cell lysis systems and for expression and functional analysis of other lethal gene products in gram-positive bacteria.

Diversity in bacterial lysis systems: bacteriophages show the way

“…There are, however, some examples where such a pair of holin-like proteins seems in fact to form the actual holin 'functional unit', as mentioned above for Gp4 and Gp5 of mycobacteriophage Ms6 and XhlA and XhlB of B. subtilis phage PBSX (Figs 3 and 4). Krogh et al (1998) studied the role of these PBSX holin-like products in host cell lysis by expressing them in B. subtilis, in different combinations with the PBSX endolysin XlyA, under the control of native transcription signals. In contrast to what happens with phage k, the study revealed that neither XhlA nor XhlB in pairwise combinations with the endolysin could trigger host cell lysis efficiently.…”

Genetics of Phage Lysis

“…The regulation of the timing of lysis as the result of activity of a complex is not restricted to a phage that infects a mycobacterial host. Similarly, it was proposed that the holin function of the Bacillus subtilis prophage PBSX is a result of two holin-like proteins, XhlA and XhlB, that associate in the membrane to form a holin functional unit (54).…”

Characterization and complete genome sequence analysis of novel bacteriophage IME-EFm1 infecting Enterococcus faecium

“…Another characteristic of the phage IME-EFm1 genome is that it contains a protein toxin, haemolysin gene (ORF22) followed by holin and endolysin genes (ORF23 and 24). ORF22 possesses a XhlA (Pfam 10779), which encodes a putative membrane-associated protein (Krogh et al, 1998). This putative haemolysin protein can insert into cellular membranes and form pores.…”

“…Holin is a protein that perforates the membrane and inserts into the bacterial cell (Wang et al, 2000). Expression of both haemolysin and holin is necessary to effect host cell lysis (Krogh et al, 1998). The endolysin function was attributed to ORF24.…”