Interplay between phosphorylation and palmitoylation mediates plasma membrane targeting and sorting of GAP43.
Bottom Line: Plasma membrane association decreased the diffusion constant fourfold in neuritic shafts.Simulations confirmed that a combination of diffusion, dynamic plasma membrane interaction and active transport of a small fraction of GAP43 suffices for efficient sorting to growth cones.Our data demonstrate a complex interplay between phosphorylation and lipidation in mediating the localization of GAP43 in neuronal cells.
Affiliation: Department of Neurobiology, University of Osnabrück, 49076 Osnabrück, Germany.Show MeSH
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Mentions: Our experimental data provided evidence that the distribution of GAP43 is guided by diffusion, phosphorylation-mediated plasma membrane association, and transport. To dissect the contribution of the different mechanisms in sorting of GAP43 to the growth cone, we developed a simulation based on three factors: 1) diffusion in the plasma membrane; 2) membrane reactions, that is, phosphorylation-mediated exchange between plasma membrane–bound and cytoplasmic protein; and 3) axonal transport of a fraction of the protein (Figure 8, A and B). The simulation showed that diffusion in the plasma membrane with a value that we had experimentally determined for PAGFP-F (∼0.4 μm2/s) as sole mechanism of distribution results in a high time constant for appearance in the growth cone. Such behavior is in agreement with the distribution of PAGFP-F as a plasma membrane–associated reference protein (red curve in Figure 8C and respective curve in Figure 6B). Inclusion of the occurrence of membrane reactions with a pseudoequilibrium constant similar to what we observed for GAP43wt (Kd* = 1 [50% bound]) and diffusion of the cytosolic component with a value that we had experimentally determined for 3×PAGFP resulted in the reduction of τ by a factor of approximately four (Figure 8C, dashed blue line). Addition of a transport component led to a further decrease of τ by a factor of two, resulting in kinetics similar to the behavior of GAP43wt (blue curve in Figure 8C and respective curve in Figure 6B). Remarkably, a fraction of only 10% of the molecules undergoing fast axonal transport was sufficient to simulate the experimental data for sorting of GAP43.
Affiliation: Department of Neurobiology, University of Osnabrück, 49076 Osnabrück, Germany.