A genome wide screening approach for membrane-targeted proteins

Area of Science:

• Proteomics
• Cell Biology
• Molecular Biology

Background:

• Membrane-associated proteins are crucial for cellular communication.
• Identifying these proteins comprehensively is essential for understanding cellular functions.
• Current methods may not fully capture the diversity of membrane-associated proteins.

Purpose of the Study:

• To develop a novel method for rapid and comprehensive identification of membrane-targeted gene products.
• To apply this method to a model organism lacking a sequenced genome.
• To assess the accuracy of current genome annotation tools for predicting membrane proteins.

Main Methods:

• Modification of the yeast Ras recruitment system to create the Ras membrane trap (RMT).
• Pilot screen of the central nervous system of the mollusk Lymnaea stagnalis using RMT.
• Bioinformatic analysis of identified gene products and their homologs.
• Experimental verification of membrane association in mammalian cells.

Main Results:

• Identified 112 gene products, with 79 lacking assignable homologs in existing databases.
• Current annotation tools predicted membrane association for only 45% of identified proteins, while experimental verification confirmed it for all tested.
• RMT identified intracellularly targeted proteins (endoplasmic reticulum, nucleus), expanding its utility beyond plasma membrane proteomes.
• Discovered two sequences representing gene families previously unknown in invertebrate genomes.

Conclusions:

• The Ras membrane trap (RMT) is an effective tool for identifying membrane-associated proteins, including intracellular ones.
• Current genome annotation tools likely underestimate the prevalence of membrane-associated proteins.
• The findings highlight the need for genome sequencing in mollusks and other major invertebrate phyla to fully understand their proteomes.