Executive Summary : | High salinity levels pose a significant challenge to agricultural productivity, as most crops cannot grow in saline soil. This leads to a gradual rise in saline lands and low salt tolerance in crops, which affects productivity. Understanding these interactions is crucial for improving salt tolerance mechanisms in crops. The proposed work aims to study the interaction between two conserved plasma membrane proteins of model plant Arabidopsis thaliana, SALT OVERLY SENSITIVE 1 (SOS1) and HIGH-AFFINITY K+ TRANSPORTER 1 (HKT1). The project uses yeasts as an expression system because HKT1 and SOS1 roles in salt tolerance are established by functional complementation in yeast mutants. A novel native-lipid nanodisc method is used to extract and isolate these yeast-expressed proteins. The method employs amphipathic Styrene Malic acid Copolymer (SMA) to encircle the protein and lipids, extracting the membrane protein in its native lipid environment. Gel electrophoresis and mass spectroscopy are used to identify lipids and proteins interacting with these proteins. There are no established protocols for isolating and studying plant proteins in yeast using the native-lipid nanodisc method. The project aims to apply this technique to isolate and study yeast-expressed SOS1 and HKT1, analyzing proteins in native-lipid nanodisc using gel electrophoresis and mass spectroscopy to identify interacting proteins and unique lipids. The protocol will also enable the identification of potential membrane protein and lipid interactions that can improve salinity tolerance mechanisms in crops. |