Abstract 4485: Identification and characterization of novel RACK1 binding partners that may modulate the elongation phase of protein synthesis
Authors: Malacrida Beatrice , Flygaard Rasmus K. , Dowling Catríona M. , Coffey John C. , Jenner Lasse B. , Kiely Patrick A. 
Published: July 2017

RACK1 (Receptor for Activated C Kinase 1) is a member of the tryptophan-aspartate (WD-repeat) family of proteins. By functioning as a scaffold protein, RACK1 is involved in regulating many different signalling pathways in cells, including cell proliferation and cell migration. Interestingly, RACK1 has been identified as a central component of the ribosomal machinery, specifically located on the 40S ribosomal subunit close to the mRNA exit channel where it makes contact with other ribosomal proteins. Importantly, it is believed that in this position, RACK1 can recruit a series of kinases and phosphatases as well as functioning to regulate the translation and the synthesis of particular cohorts of proteins. Moreover, RACK1 plays a central role in influencing the location of the translation machinery.

Using 2D and 3D models of colon cancer cell lines, together with Click-IT chemistry and Mass Spectrometry analysis, we were able to identify a series of proteins, including newly synthesized proteins, that interact with RACK1. We identified proteins involved in cell cycle control, metabolism and protein synthesis. As expected, we have found a number of proteins that are involved in cell migration and a number of proteins that are mediators of the cell adhesion process. Interestingly, some of the most highly scored interacting proteins were those involved in protein synthesis. Of those, we identified several elongation factors as being novel RACK1 binding partners. Elongation factors are a class of proteins directly involved in the elongation phase of protein synthesis. Along with this primary function, these factors have different important roles in cells, such as influencing cytoskeleton remodelling and activation of growth and proliferation pathways. Elongation factors have been found to be upregulated in many different types of cancer.

We have confirmed the interaction between RACK1 and several of these elongation factors and we have determined that many of these are dysregulated in colon cancer. We have established that the activity of these elongation factors is regulated by a combination of stress conditions and growth factor stimulations. We have determined that RACK1 plays a central role in scaffolding these proteins to promote proliferation and migration in our colon cancer cell models. Collectively, these results provide a novel and interesting role for RACK1 in the modulation of the elongation phase of protein synthesis.

We are now investigating the subcellular location of these factors in response to different stimuli and asking how is RACK1 involved in their localization downstream of IGF-I and integrin signalling. Characterizing and mapping the interaction between these proteins and RACK1, both in cells and tissue, may lead to the design of novel therapeutic approaches in colon cancer.

Citation Format: Beatrice Malacrida, Rasmus K. Flygaard, Catríona M. Dowling, John C. Coffey, Lasse B. Jenner, Patrick A. Kiely. Identification and characterization of novel RACK1 binding partners that may modulate the elongation phase of protein synthesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4485. doi:10.1158/1538-7445.AM2017-4485