Transcriptome-wide organization of subcellular microenvironments revealed by ATLAS-Seq

Area of Science:

• Molecular Biology
• Cell Biology
• Genomics

Background:

• Subcellular organization of RNAs and proteins is crucial for cellular function.
• Current understanding of molecular localization within cells and tissues remains incomplete.
• Global maps and conceptual frameworks for RNA and protein localization are needed.

Purpose of the Study:

• To introduce ATLAS-Seq, a novel method for mapping RNA and protein localization.
• To generate transcriptomes and proteomes from fractionated tissue lysates.
• To investigate the principles governing subcellular organization of RNA and protein molecules.

Main Methods:

• Utilized detergent-free tissue lysates fractionated via sucrose gradient centrifugation.
• Performed proteomic analysis to confirm separation of subcellular compartments.
• Generated transcriptomic and proteomic data from fractionated samples.

Main Results:

• Proteomic analysis validated the separation of subcellular compartments.
• RNAs unexpectedly co-sedimented with other RNAs, protein complexes, or functionally related molecules.
• Most RNAs showed different sedimentation profiles compared to their encoded proteins, except for those of secreted proteins.
• Hundreds of alternative RNA isoforms displayed distinct sedimentation patterns.
• Identified potential RNA binding proteins driving observed RNA localization patterns.
• Confirmed known RNA-protein interactions and predicted novel associations.

Conclusions:

• RNAs are organized into networks of co-segregating mRNAs encoding functionally related proteins.
• These findings provide new insights into the establishment and maintenance of subcellular organization.
• ATLAS-Seq offers a powerful tool for dissecting molecular organization within cells.