Characterization of Germination Receptors of Bacillus cereus ATCC 14579

Abstract

Specific amino acids, purine ribonucleosides, or a combination of the two is required for efficient germination of endospores of Bacillus cereus ATCC 14579. A survey including 20 different amino acids showed that L-alanine, L-cysteine, L-threonine, and L-glutamine are capable of initiating the germination of endospores of B. cereus ATCC 14579. In addition, the purine ribonucleosides inosine and adenosine can trigger germination of the spores. Advanced annotation of the B. cereus ATCC 14579 genome revealed the presence of seven putative germination (ger) operons, termed gerG, gerI, gerK, gerL, gerQ, gerR, and gerS. To determine the role of the encoded putative receptors in nutrient-induced germination, disruption mutants were constructed by the insertion of pMUTIN4 into each of the seven operons. Four of the seven mutants were affected in the germination response to amino acids or purine ribonucleosides, whereas no phenotype could be attributed to the mutants with disrupted gerK, gerL, and gerS loci. The strain with a disrupted gerR operon was severely hampered in the ability to germinate: germination occurred in response to L-glutamine but not in the presence of any of the other amino acids tested. The gerG mutant showed significantly reduced L-glutamine-induced germination, which points to a role of this receptor in the L-glutamine germination signaling pathway. gerR, gerI, and gerQ mutants showed reduced germination rates in the presence of inosine, suggesting a role for these operons in ribonucleoside signaling. Efficient germination by the combination of L-glutamine and inosine was shown to involve the gerG and gerI operons, since the germination of mutants lacking either one of these receptors was significantly reduced. Germination triggered by the combination of L-phenylalanine and inosine was lost in the gerI mutant, indicating that both molecules are effective at the GerI receptor.

The Ger Receptor Family from Sporulating Bacteria

“…Interestingly, no single Ger receptor seemed to be essential for inosine-mediated germination. However, GerI, GerQ, and GerR mutants germinated at lower rates than wild-type spores when treated with inosine alone (Hornstra et al, 2006). This suggests that GerI, GerQ, and GerR are able to individually recognize inosine, but that their function is redundant.…”

“…The addition of inosine as a co-germinant with amino acids enhanced the germination response of B. cereus ATCC 14579 (Hornstra et al, 2006). The germination response to cogerminants was stronger with inosine/L-glutamine and inosine/L-phenylalanine.…”

“…The germination response to cogerminants was stronger with inosine/L-glutamine and inosine/L-phenylalanine. The combination of L-glutamine and inosine required the GerG and GerI receptors, while Lphenylalanine/inosine combinations required only the GerI receptor (Hornstra et al, 2006). This indicates that the GerI receptor interacts with L-phenylalanine and inosine and that both molecules contribute to germination.…”

“…Wild-type B. cereus ATCC 14579 spores germinated strongly with L-alanine, Lcysteine, L-threonine, and L-glutamine (Hornstra et al, 2006). Mutational analysis shows that the GerR receptor influences all germination responses to single amino acids, except Lglutamine (Hornstra et al, 2006).…”

“…Inosine is a known germinant of B. cereus spores (Barlass et al, 2002;Clements and Moir, 1998;Hornstra et al, 2006). Indeed, B. cereus ATCC 14579 spores are able to germinate with inosine as sole germinant (Hornstra et al, 2005).…”