Polynucleotide phosphorylase is necessary for competence development in Bacillus subtilis

Abstract

comR (pnpA) is a newly identified gene in Bacillus subtilis that is necessary for the expression of late competence genes. Transformability of a comR (pnpA) mutant is 1-5% of that seen in comR+ strains. Cloning and sequencing identified ComR as polynucleotide phosphorylase (PNPase). The PNPase amino acid sequence has 50% identity and 67% similarity with the Escherichia coli enzyme. Enzymatic assays show that this is the only PNPase activity in B. subtilis. comR (pnpA) is necessary for comG-lacZ and comK-lacZ expression, but this requirement is bypassed by a mecA disruption. In B. subtilis, the loss of PNPase has little effect on expression from a fusion of the srfA promoter directly to lacZ, but is necessary for normal expression from certain srfA-lacZ fusions that include portions of the normal srfA transcript. When a srfA-lacZ translational fusion is tested in isogenic pnpA+ and pnpA derivatives of E. coli, lower expression is seen in the pnpA mutant. Since expression from lacZ fusions to comA, sinR, and mecA appeared similar in the B. subtilis pnpA and pnpA+ strains, the loss of PNPase does not have a strong general effect on gene expression. These results suggest that PNPase may be necessary for modification of the srfA transcript in order to activate translation or stabilize the transcript, and that this may be necessary for competence development. This is the first evidence of post-transcriptional effects on the development of competence in B. subtilis.

Modulation of Yersinia Type Three Secretion System by the S1 Domain of Polynucleotide Phosphorylase

“…This observation prompted us to test whether the phenomenon of PNPase enhancing growth at low temperatures was also independent of its ribonuclease activity, an issue that to the best of our knowledge has not been addressed. Similar to what has been previously reported for E. coli, B. subtilis, and Y. enterocolitica (3)(4)(5), PNPase is clearly required for YPT and YP to grow at low temperatures ( Fig. 9A; data for YP not shown).…”

“…For example, the cold shock response of Escherichia coli involves the induction of several genes including cold shock proteins (CSP) 1 as well as exoribonucleases PNPase and RNaseR involved in RNA metabolism (1,2). In addition to E. coli, Bacillus subtilis and Yersinia enterocolitica have been shown to require PNPase for growth in the cold (3)(4)(5). It is believed that PNPase is required for the restart of growth following cold shock by degrading the remaining CSP mRNA molecules (5,6).…”

Regulation of a Bacillus subtilis mobile genetic element by intercellular signaling and the global DNA damage response

“…Transfer into recipients that contained ICEBs1 occurred with Ϸ50-fold lower frequency ( Table 1), indicating that ICEBs1 encodes at least one mechanism that inhibits acquisition of a second element. Acquisition of ICEBs1 by recipients was not due to natural transformation, because the recipients were comK mutants incapable of transformation (32). Transfer of ICEBs1 from nonactivated donor cells [⌬(rapI phrI)::kan, IRN342] was not detected under these conditions (Ͻ2 ϫ 10 Ϫ8 transconjugants per donor).…”

Overexpression of the polynucleotide phosphorylase gene (pnp) of Streptomyces antibioticus affects mRNA stability and poly(A) tail length but not ppGpp levels

“…Some 20 ribonucleases (RNases) have been identified in Escherichia coli (Regnier & Arraiano, 2000), and several of these are known to be involved in the decay and processing of stable and messenger RNAs. The complement of RNases identified thus far in Bacillus is smaller than that of E. coli (Farr et al, 1999), but several of these enzymes function analogously to those of E. coli (Luttinger et al, 1996;Herskovitz & Bechhofer, 2000) and it is clear that in both species gene expression can be regulated at the level of RNA stability (Luttinger et al, 1996;Grunberg-Manago, 1999).…”