Simultaneous profiling of native-state proteomes and transcriptomes of neural cell types using proximity labeling

Area of Science:

• Molecular Biology
• Cell Biology
• Neuroscience

Background:

• Cellular phenotyping requires understanding both transcriptomic and proteomic profiles.
• Transcriptome and proteome abundances show only modest correlation, necessitating complementary profiling.
• Existing methods often profile transcriptomes and proteomes separately, limiting comprehensive cellular analysis.

Purpose of the Study:

• To develop and validate a novel method for simultaneous, cell type-specific transcriptomic and proteomic profiling.
• To capture native-state proteomes and transcriptomes from specific cell types in vitro and in vivo.
• To investigate mRNA-protein concordance and discordance across different cell types and conditions.

Main Methods:

• Simultaneous Protein and RNA-omics (SPARO) method utilizing TurboID for biotinylation of RNA-interacting cytosolic proteins.
• Application in vitro using BV2 microglial and HEK293 cell lines.
• In vivo validation in astrocytic and neuronal Cre driver mice crossed with Rosa26-TurboID knock-in mice.

Main Results:

• SPARO successfully captured cell type-specific transcriptomes and proteomes simultaneously in vitro and in vivo.
• The method accurately reflected global proteomes and transcriptomes and captured neuroinflammatory responses.
• Analysis revealed insights into molecular processes with uniform or cell type-specific mRNA-protein concordance patterns.

Conclusions:

• SPARO provides a powerful approach for comprehensive cellular phenotyping by integrating transcriptomic and proteomic data.
• The method enables deeper understanding of molecular mechanisms underlying cellular functions, development, aging, and disease.
• SPARO facilitates the study of mRNA-protein relationships in a cell type-specific manner, offering new biological insights.