Nck Beta Adapter Protein Coordinates Bcr-Abl/Sam68 Intermolecular Interaction.
Authors: Bracco Enrico , Deklic Erika , Rosso Valentina , Mussino Stefano , Arruga Francesca , Carturan Sonia , Messa Francesca , Defilippi Ilaria , Panuzzo Cristina , Nicoli Paolo , Messa Emanuela , Rotolo Antonia , Pedrola Roberto , Morotti Alessandro , Balbo Alessandra , Iacobucci Ilaria , Martinelli Giovanni , Cilloni Daniela , Saglio Giuseppe 
Published: April 2020

Background: Philadelphia positive (Ph+) disorders, such as Chronic Myelogenous Leukemia (CML) are characterized by the presence of abnormal chromosome rising from translocation between chromosome 9 and 22 thus giving birth to a chimeric oncogenic protein named Bcr-Abl. This oncogenic kinase displays constitutive tyrosine kinase activity which leads to tyrosine residues autophosphorylation, in turn recruiting SH2 and/or PTB containing proteins. In the last decade Bcr-Abl targeted therapy has been successfully employed and, among currently available drugs inhibiting Bcr-Abl activity, Imatinb mesylate represents the most efficient. Despite the huge amount of data reporting the effects of Imatinib on signal transduction pathways (for example ERK1/2 and PI3K activation, CrkL phosphorylation etc…) in Ph+ leukemic cells rather few experimental evidences are available on the effects of Imatinib on adapter molecules. Therefore we are currently attempting to investigate such field.

Aims and Methods: The significance of interactions occurring between Bcr-Abl and adapter molecules is still matter of debate. Most of the interactions so far described (CrkL, Grb2, PI3K p85 regulatory subunit etc…) appear to play a role in coordinating and integrating a plethora of signals which in turn lead to proliferation, cell survival and/ or cytoskeletal organization. In the last years few, but very interestingly, data supporting the hypothesis that adapter molecules might also act as c-Abl catalytic regulators have been presented. By means of an interactomic approach, based on proteomic strategy using GST-Pull Down assay with an array of SH2 containing proteins, we attempted to gain insight into the role played by adapter molecules and Bcr-Abl interactions.

Results and Conclusions: The data herein presented aims to demonstrate the presence of quaternary complex involving the SH2-SH3 containing adapter protein Nck-beta, the oncogenic tyrosine kinase Bcr-Abl and the RNA binding protein Sam68. The experimental evidences we have collected support the hypothesis of an Imatinib-dependent interaction occurring between Nck-beta and Bcr-Abl. Furthermore, Pull Down experiments indicate an intermolecular interaction between Nck-beta and Sam68, supporting the idea of a novel complex Bcr-Abl/Nck-beta/Sam68. Interestingly, preliminary data carried-out using RNA Pull Down assay suggest that the quaternary complex Nck-beta/Sam68/ Bcr-Abl might modulates splicing process of a gene encoding for a protein capable of regulating apoptosis events, such as Bcl-X. Taken together these results represent the first experimental evidences showing an interaction between the oncogene Bcr-Abl and Sam- 68 leading to speculate a novel putative role played by Bcr-Abl in the intriguing and complex mRNA splicing scenario.