Fig. 6: c-di-GMP reversed the inhibitory effects of MbpR.
a–d EMSA detection of MbpR (a, b) and MbpRR192A (c, d) binding to the mbpB or mbpA coding DNA sequence (CDS) probe in the presence of different amounts of c-di-GMP. The probe was incubated with protein at room temperature for 30 min. The experiments were performed three times with similar results. e Intracellular c-di-GMP level of the wild type, Δ3DGC, and Δ2PDE mutants. f Relative mbpA and mbpB transcription level in Δ3DGC and Δ2PDE strains analyzed by RT-qPCR. g, h Fluorescence intensity of EGFP, which was expressed from the reporting plasmid under the control of mbpA (g) and mbpB (h) promoter, in the wild-type, Δ3DGC, and Δ2PDE strains. The Δ3DGC strain was constructed by deleting dgcD, dgcC, and dgcB. The Δ2PDE strain mutant was generated by deleting pdeD and pdeA. i A pathway for c-di-GMP mediating L. plantarum colonization (Created in BioRender. Zhang, J. (2025) https://BioRender.com/g28k816). At low c-di-GMP level, MbpR binds to the mucin-binding protein (MucBP) CDS and inhibits its transcription. After sensing unknown signals, the activated diguanylate cyclases or inhibited c-di-GMP phosphodiesterases maintain the intracellular c-di-GMP level at a high level. The elevated c-di-GMP level results in MbpR releasing from the CDS of MucBP and promotes MucBP expression, promoting the colonization of L. plantarum. RNAP: RNA polymerase. Data in (e–h) are presented as mean values ± standard deviations (SD). Error bars indicate the SD. Dots represent individual data. Statistical significance in (e, g, h) was analyzed using one-way Analysis of Variance (ANOVA) with Tukey’s multiple-comparison test. In (e, f): n = 3 biological repeats, In (g, h): n = 5 biological repeats. Source data are provided as a Source Data file.
