cTag-PAPERCLIP Reveals Alternative Polyadenylation Promotes Cell-Type Specific Protein Diversity and Shifts Araf Isoforms with Microglia Activation

• Published in: Neuron (Cambridge, Mass.) Mass.), 2017-09, Vol.95 (6), p.1334-1349.e5
• Main Authors: Hwang, Hun-Way, Saito, Yuhki, Park, Christopher Y., Blachère, Nathalie E., Tajima, Yoko, Fak, John J., Zucker-Scharff, Ilana, Darnell, Robert B.
• Format: Article
• Language: English
• Subjects: Activation; alternative polyadenylation; Animals; Antigens, Neoplasm - physiology; ARAF; Astrocytes; Astrocytes - metabolism; Brain - cytology; Brain - metabolism; Cells, Cultured; crosslinking immunoprecipitation; Cytokines; Female; Gene expression; Gene mapping; Genomes; Humans; Inflammation; Isoforms; Male; Mice; Microglia; Microglia - cytology; Microglia - metabolism; Nerve Tissue Proteins - physiology; neuron; Neurons; Neurons - metabolism; NOVA; Organ Specificity; Polyadenylation; Polypyrimidine Tract-Binding Protein - physiology; Protein Isoforms - metabolism; Protein-Serine-Threonine Kinases - metabolism; PTB; Ribonucleic acid; RNA; RNA-Binding Proteins - physiology; Rodents; single cell type; Studies
• ISSN: 0896-6273; 1097-4199
• DOI: 10.1016/j.neuron.2017.08.024

Alternative polyadenylation (APA) is increasingly recognized to regulate gene expression across different cell types, but obtaining APA maps from individual cell types typically requires prior purification, a stressful procedure that can itself alter cellular states. Here, we describe a new platform, cTag-PAPERCLIP, that generates APA profiles from single cell populations in intact tissues; cTag-PAPERCLIP requires no tissue dissociation and preserves transcripts in native states. Applying cTag-PAPERCLIP to profile four major cell types in the mouse brain revealed common APA preferences between excitatory and inhibitory neurons distinct from astrocytes and microglia, regulated in part by neuron-specific RNA-binding proteins NOVA2 and PTBP2. We further identified a role of APA in switching Araf protein isoforms during microglia activation, impacting production of downstream inflammatory cytokines. Our results demonstrate the broad applicability of cTag-PAPERCLIP and a previously undiscovered role of APA in contributing to protein diversity between different cell types and cellular states within the brain. •cTag-PAPERCLIP generates in vivo profiles of cell-type-specific polyadenylated mRNAs•cTag-PAPERCLIP preserves tissue integrity, capturing mRNAs in the cell’s native state•Distinct APA preference contributes to protein diversity between brain cell types•Activated brain microglia upregulate full-length kinase-active ARAF through APA Hwang et al. develop cTag-PAPERCLIP to profile mRNA 3′ ends in individual cell types from intact tissues in vivo. They use cTag-PAPERCLIP to show that mRNA alternative polyadenylation contributes to protein diversity between different cell types and cellular states within the brain.