Cytidine Repressor Participates In The Regulatory Pathway Of Indole In Pantoea Agglomerans YS19

Bacteria evolve a lot of signal regulatory mechanisms among individuals in confronting diverse environmental stresses, of which indole-based regulation attracts more attention. As a signal molecule regulating intricate metabolism, in our previous work, indole was found to be related to the formation of multicellular aggregate structures, bacterial motility and in planta colonization. However, the regulatory mechanism is still under investigantion. To capture the key proteins in the indole regulatory pathway is significant for understanding its regulatory mechanism and also the formation mechanisms of bacterial aggregation.This study selected endophyte Pantoea agglomerans YS19 which can form unique multicellular aggregate structure symplasmata as the model strain. A positive mutant down-regulated by indole was screened using the mTn5 transposon mutagenesis method and GusA activity method. By TAIL-PCR, the mutant gene was identified as cytR, encoding cytidine repressor. Many physiological behaviors were changed in the cytR mutant. The mutation of cytR promotes the growth in the early stationary growth phase, yet has no significant effect on the symplasmata formation. Indole promotes the symplasmata formation in both YS19 and △ cytR. The mutation of cytR enhances the swimming and swarming abilities of YS19. Besides, exogenous indole significantly inhibits the swimming and swarming abilities of widtype and △ cytR mutant. Meanwhile, the mutation of cytR decreased the growth promoting effects and colonization number of YS19, which, however, are increases by low concentrations of indole in both strains.A differential protein that was expressed in the cytR mutant was detected by SDS-PAGE and HPLC-MS. It was identified as the deoxyribose 5-phosphate aldolase(the molecular mass is 27.7 KDa) encoded by deoC. By real-time PCR, it was shown that indole promotes the expression of deoC, suggesting that the down-regulation effect of indole on cytR, as well as the deletion of cytR, relieves or partly relieves the inhibition of CytR on the deoC transcription, making the increased expression of deoC. In addition, under starvation condition, the expression of deoC increased, demonstrating that DeoC is one of regulatory proteins of YS19 in confronting starvation. Meanwhile, indole improves the viability of YS19 in starvation treatment. We infered that as a signal molecule, indole down-regulates the expression of cytR making an improved expression of deoC, which increases the amount of DeoC making an improved survival rate of YS19 under the starvation conditions. This study discussed the indole regulatory pathways from the point of DNA transcription regulation, which enriched our knowledges of cytidine repressor and expanded the research ideas of indole regulation.