Protein kinase A governs a RhoA–RhoGDI protrusion–retraction pacemaker in migrating cells

Abstract

The cyclical protrusion and retraction of the leading edge is a hallmark of many migrating cells involved in processes such as development, inflammation, and tumorigenesis. The molecular identity of signaling mechanisms that control these cycles has remained unknown. Here, we used live cell imaging of biosensors to monitor spontaneous morphodynamic and signaling activities, and employed correlative image analysis to examine the role of cAMP-activated Protein Kinase A (PKA) in protrusion regulation. PKA activity at the leading edge is closely synchronized with rapid protrusion and with the activity of RhoA. Ensuing PKA phosphorylation of RhoA and the resulting increased interaction between RhoA and RhoGDI establishes a negative feedback that controls the cycling of RhoA activity at the leading edge. Thus, cooperation between PKA, RhoA, and a RhoGDI forms a pacemaker that governs the morphodynamic behavior of migrating cells.

Control of the G-protein cascade dynamics by GDP dissociation inhibitors

“…[30][31][32][33] Since in the cytoplasm GTPases rapidly bind to GDIs, in the model we consider the overall reactions where the dissociation of a GTPase from the membrane is combined with the binding to a GDI, and the dissociation of a GTPase from the GDI is combined with the subsequent membrane association. In terms of the overall reactions, the data show that the GTP-bound RhoGTPase binds GDIs with much lower affinity than the GDP-bound RhoGTPase 28 (in line with the observations that the latter dissociates much faster from the membrane than the former). 30,31 While this difference in the equilibrium affinity constants is included in our model, it is unknown if the affinities of RhoGDI1 for RhoA and Rac1 are similar or significantly different.…”

“…For instance, phosphorylation of RhoA by protein kinase A (PKA) facilitates RhoA binding to RhoGDI, thereby promoting RhoGDI to associate faster to RhoA and sequester the complex into the cytoplasm. 28 Here we controlled the binding affinity by changing the rate constant for binding RhoGDI to RhoA (denoted b 1 , see Eq. 4 in Methods).…”

PKA-induced dimerization of the RhoGAP DLC1 promotes its inhibition of tumorigenesis and metastasis

“…The cAMP/PKA pathway has a profound role in the regulation of cell motility and cytoskeletal reorganization 35 . Furthermore, PKA has been shown to hinder multiple components of Rho signalling, which are crucial to cytoskeletal dynamics and cell movement 36,37 . PKA directly phosphorylates RhoA at Ser188 and inhibits its function by enhancing the interaction between RhoA and the Rho guanine-dissociation inhibitor 38 .…”

A dual role model for active Rac1 in cell migration

“…32 Quasi-periodical protrusions of the leading edge segments have been associated with RhoA activity, and a pacemaker cycle has been postulated that additionally involves protein kinase A and RhoGDI. 33 Correlated fluctuations of lamellipodial segments in large cells occur on a spatial scale of ten microns and a temporal scale of a minute. 32,33 Shape fluctuations of Dictyostelium cells typically occur on similar spatial and temporal scales during random motility.…”