Fig 1.
The regulation of DivK and CtrA phosphorylation during the cell division cycle of Caulobacter crescentus.
(A) Influence diagram of two signal transduction pathways in C. crescentus. Barbed arrows indicate activation, while the blunt-headed line indicates inhibition. (B) Spatiotemporal distributions of DivK~P (red) and CtrA~P (light blue). PD = predivisional. Notice that both DivK and CtrA are phosphorylated in the PD cell.
Fig 2.
Two scenarios for the function of PleC (kinase or phosphatase) in the early predivisional cell.
Spatiotemporal dynamics of PleC (green) and DivL (dark blue) during the cell cycle under two scenarios for the functional transition of PleC. In scenario 1, PleC is a kinase in early PD cells, as suggested in [54–56]. In scenario 2, PleC is a phosphatase in early PD cells, as described in the protection by dephosphorylation model [45,53].
Fig 3.
Co-localization of PleC kinase and DivL in the early predivisional cell is required for DivL reactivation.
Spatiotemporal distributions of proteins during the cell cycle (prior to cytokinesis at t = 120 min). Color indicates concentration gradients from minimum (blue) to maximum (red). (A) DivJ is localized at the old pole (t = 30–120 min). The location of PleC is shifted from the old pole (t = 0–50 min) to the new pole of the predivisional cell (t = 90–150 min). Following DivJ localization, the function of PleC changes from a phosphatase to a kinase. (B) Upon phosphorylation, DivK localizes to the poles of the cell. Despite the presence of DivK~P at the new pole of the predivisional cell, DivL is present in the free form (unbound to DivK~P) because DivL co-localizes with PleC kinase and PleC kinase sequesters DivK~P, preventing it from binding to DivL. (C) CckA is uniformly distributed in the swarmer stage and localized at both poles in the predivisional stage. Reactivation of DivL at the new pole results in new-pole CckA becoming a kinase, while old-pole CckA remains a phosphatase. Consequently, the late predivisional cell establishes a gradient of CtrA~P along its length from high at the new pole to low at the old pole.
Fig 4.
Following cytokinesis, PleC reverts to the phosphatase form.
(A) The total concentration of CckA (blue curve) remains constant during the cell cycle. However, the proportions of phosphatase (red curve) and kinase (green curve) forms of CckA change for each stage of the cell cycle. After cytokinesis (compartmentalization), the concentrations of the kinase and phosphatase fractions of CckA in the swarmer and stalked compartments (t = 120–150 min) are similar to their concentration in the non-compartmentalized swarmer (t = 0–30 min) and stalked cell stages (t = 30–90 min), respectively. (B) Spatiotemporal distribution of PleC kinase, DivK~P and CtrA~P after compartmentalization (at t = 120 min). Color indicates concentration gradients from minimum (blue) to maximum (red).
Fig 5.
Overexpression of DivK may prevent the cell from progressing though different developmental stages.
For better comparison of mutant and wild-type distributions, the colors indicate concentration gradients from zero (blue) to maximum wild-type concentration, Cwt_max (red). For Cwt_max < Cmut_max, the red color represents all concentrations from Cwt_max to Cmut_max. For Cwt_max > Cmut_max, an additional tick between 0 and Cwt_max indicates maximum concentration in the mutant. (A) A four-fold increase in DivK synthesis (ksyn = 0.2 min-1) prevents competition between DivL and PleC kinase. DivL is not reactivated in the predivisional stage, leading to the loss of CckA kinase and CtrA~P asymmetry in the early predivisional stage. (B) An eight-fold increase in DivK synthesis (ksyn = 0.4 min-1) induces PleC to become a kinase independent of DivJ. Hence, the swarmer stage of the cell cycle is circumvented.
Fig 6.
Swarmer-to-stalked transition is uncoupled from G1-to-S transition in DivL mislocalization mutants.
Concentration gradients are color coded as in Fig 5. (A) DivL is uniformly distributed at all times in the cell cycle. Even though PleC transitions to a kinase, DivL remains deactivated (bound to DivK~P) in the predivisional cell, resulting in no gradient of CtrA~P. (B) DivL (and CckA) are localized at the old pole (t = 0–90 min), before switching to the new pole in the predivisional cell (t = 90–120 min). Even after PleC transitions to a kinase, DivL is not deactivated in stalked cells, delaying the dephosphorylation of CtrA~P until PleC delocalizes from the old pole (at t = 50 min). (C) DivL and PleC are co-localized at one of the poles during all stages of the cell cycle. Hence, CtrA is phosphorylated through all stages of the cell cycle (G1-arrest).
Fig 7.
PleC kinase conformation is required to establish replicative asymmetry.
Concentration gradients are color coded as in Fig 5. (A) In ΔpleC mutant cells (K-P-B-), free active form of DivL is lower than in wild-type cells, resulting in loss of CtrA~P in the predivisional cell. (B) In pleCH610A mutant cells (K-P-B+), an elevated level of DivK~P results in less active form of DivL and reduced CtrA~P. (C) In pleCF778L mutant cells (K-P+B+), inhibitor sequestration is retained, resulting in a normal CtrA~P gradient.
Fig 8.
The model simulates distribution patterns of CtrA~P and DivK~P in mutant cells.
(A) Model simulations predict the concentration gradient of CtrA~P prior to cytokinesis (t = 120 min) in mutant cells: pleCF778L (purple), divKD90G (blue), ΔpleC (red) and pleCH610A (yellow). The distribution for the wild-type cells is plotted in green for reference. (B) The model reproduces the inadequate dephosphorylation of DivK~P observed in the swarmer compartment of post-compartmentalized cells in the ΔpleC (red) and pleCH610A (yellow) mutant strains.
Fig 9.
Localization and function of the DivK-PleC-DivK and DivL-CckA-CtrA signaling networks, as suggested by the model calculations reported here.