Fig. 2: The Pf6 KKPMP module controls Pf lysogeny via MvaU phosphorylation.
From: Control of lysogeny and antiphage defense by a prophage-encoded kinase-phosphatase module
a Differentially-expressed genes in biofilms versus planktonic cells in MPAO1 and PAO1. Statistically significantly changed Pf6 and Pf4 genes are labeled in red and blue, respectively. NS, not significant. b RNA-seq read coverage of Pf4 and Pf6 transcripts in MPAO1 biofilm and planktonic cells. Pf6 KKPMP genes (pfkA, pfkB and pfpC) are boxed. c Relative mRNA expression levels of pfkA, pfkB and pfpC in MPAO1 planktonic and biofilm cells (day 2–8). The (pfkA+pfkB)/pfpC ratio is shown as a blue line. d Pf phage titer kinetics in biofilms from MPAO1, MPAO1 ΔpfkA, and MPAO1 ΔKKMP using methods established in Supplementary Fig. 3c–e. e Virus to microbe ratios (VMR) in biofilms of MPAO1, MPAO1 ΔpfkA and MPAO1 ΔKKMP were determined by qPCR using primers to amplify an identical fragment of Pf4 and Pf6 and to amplify chromosomal gene gyrB. f Top: MvaU domain schematic. Left: Phosphorylation sites in native MvaU identified by phosphoproteomics in PAO1 with expression of indicated KKPMP components. Right: Phosphorylation sites in native MvaU-His purified from MPAO1 with indicated KKPMP components. g Effects of chromosomal expression of MvaU WT or MvaUS67D on the production of Pf4 and Pf6 during biofilm formation at day 2 and day 6. Data are shown as the mean ± SD. Two-sided Student’s t Test was used for comparisons of phage titers between MvaU and MvaUS67D at each day (n = 3), and P < 0.05 was considered statistically significant. h Model for KKPMP control of Pf lysogeny via MvaU phosphorylation. In planktonic (free-living) cells, where there is relatively minimal expression of PfpC, PfkA and PfkB lead to MvaU phosphorylation and maintain Pf lysogeny. In biofilms, elevated PfpC expression inhibits kinase activity, leading to MvaU dephosphorylation, elevated Pf gene expression and phage propagation. The size of each labeled shape corresponds to relative expression levels for each protein in each condition. At least two independent cultures were used (n = 3 for a, c, d, f, g; n = 2 for e), and data are shown as the mean ± SD in (c–e, g).
