Spatially resolved mapping of proteome turnover dynamics with subcellular precision

Area of Science:

• Cellular Biology
• Proteomics
• Biochemistry

Background:

• Cellular functions rely on dynamic proteomic changes at the subcellular level.
• Existing methods for protein turnover lack spatial resolution within cells.
• Understanding subcellular protein dynamics is crucial for deciphering cellular mechanisms.

Purpose of the Study:

• To develop a novel method, prox-SILAC, for mapping newly synthesized proteins with subcellular resolution.
• To investigate proteome dynamics within the mitochondrial matrix and endoplasmic reticulum lumen.
• To explore protein turnover in response to cellular stress and differentiation.

Main Methods:

• Combined proximity-dependent protein labeling (APEX2/HRP) with metabolic stable isotope labeling (pulse-SILAC).
• Applied the prox-SILAC technique to analyze protein turnover in specific subcellular compartments.
• Investigated proteomic dynamics in mitochondrial and ER environments under various cellular conditions.

Main Results:

• Revealed heterogeneous protein turnover dynamics in mitochondrial ribosomes and respiratory complexes, suggesting hierarchical assembly.
• Identified distinct protein turnover patterns in the endoplasmic reticulum during stress or differentiation.
• Highlighted specific protein subsets with unique dynamics potentially involved in stress response and differentiation.

Conclusions:

• Prox-SILAC enables high-resolution mapping of protein synthesis and turnover at the subcellular level.
• The method provides insights into the assembly mechanisms of macromolecular machines and organelle dynamics.
• Prox-SILAC is a versatile tool for studying protein dynamics in various subcellular compartments under physiological and pathological conditions.