Dynamic imaging of the intracellular trafficking of ERK suggests a novel mechanism at the basis of the functional differences between ERK1 and 2

In this thesis I studied the localization and trafficking in living cells of the Extracellular Regulated Kinase signaling by making visible ERK1 and ERK2 with a fluorescent tag. This approach allowed to identify different dynamical properties of the two kinases, posing the bases for the understanding of the functional differences between ERK1 and ERK2. The nucleo-cytoplasmic trafficking of tagged ERK1/2 has been measured by means of FRAP experiments. Surprisingly, I found that ERK1 shuttles at a much slower rate than ERK2. Moreover, I demonstrated that this difference is caused by an unique domain of ERK1 located at its N-terminus, since the progressive deletion of these residues converts the shuttling features of ERK1 into those of ERK2. Conversely, the fusion of this ERK1 sequence at the N-terminus of ERK2 slows down its shuttling to a similar value found for ERK1 and, when fused to small cargos such as a GFP monomer, it is capable of hampering their shuttling too. In addition, I identified some crucial aminoacids at ERK1 N-terminus, responsible in large part of this phenotype. Finally, I have demonstrated that the speed of nucleo-cytoplasmic shuttling critically affects the ERK capability of activating downstream effectors. In conclusion, I propose a novel biochemical model, in which the regulation of nucleo-cytoplasmic trafficking might provide a sensitive mechanisms through which cells modulate their response to extracellular stimulus. This mechanism significantly contributes to the differential ability of ERK1 and 2 to generate an overall signaling output.