阿拉伯芥RACK1A是否會與14-3-3蛋白有分子上的交互作用？

Abstract

Receptor for Activated C-Kinase 1 (RACK1) is a 36-kDa scaffold protein, which contains seven Trp-Asp 40 (WD40) repeats. RACK1 is highly conserved protein in both animals and plants. The WD40 repeats are involved in protein-protein interactions. There are three RACK1 genes in Arabidopsis thaliana, namely RACK1A (At1g18080), RACK1B (At1g48630) and RACK1C (At3g18130). RACK1 is involved in multiple hormone responses, developmental processes, and associated with 40S ribosomes in Arabidopsis. Notably, RACK1 is regarded as a versatile scaffold protein which serves as a nexus for multiple signal transduction pathways. Moreover, RACK1 plays a negative regulator under abscisic acid (ABA) responses, and rack1a mutants are more resistant to water stress. However, the specific interactions between RACK1 and its binding partners are still unclear. Previous immunopreciptation studies revealed that RACK1A may interact with 14-3-3. Here, we used yeast two-hybrid and bimolecular fluorescence complementation to study the interactions between these two proteins. Our results indicated that RACK1A did not interact with 14-3-3 omega directly. Furthermore, we observed that the RACK1A protein is localized to the cytoplasm in Arabidopsis protoplasts. Point mutation of a serine residue located in the C-terminue resulted in subcellular localization change. We use RACK1A overexpression lines to the possibility to identify RACK1A interacting proteins by immunoprecipitation.