In vivo AGO-APP for cell-type- and compartment-specific miRNA profiling in the mouse brain

TL;DR

Researchers developed two transgenic mouse lines to study microRNA (miRNA) profiles in specific brain cells and compartments, revealing distinct miRNA patterns in different cell types and synaptic regions.

Contribution

The creation of two transgenic mouse lines enables in vivo cell-type- and compartment-specific miRNA profiling using AGO-APP, offering a novel approach for studying miRNA function in the brain.

Findings

OB interneurons and cortical excitatory neurons have distinct miRNA signatures, including enrichment of miR-200 family and miR-183/96/182 cluster in OB interneurons.

PSD95-T6B fusion identifies postsynapse-enriched miRNAs that target synaptic function-related mRNAs.

Abstract

AGO-APP through the expression of the T6B peptide permits the isolation of Ago-bound microRNAs (miRNAs). Here, we present the generation and characterization of two transgenic mouse lines that enable AGO-APP to be performed in vivo. First, we generated mice for CRE-dependent T6B expression throughout the cell. Using this line, we performed AGO affinity purification (AGO-APP) in olfactory bulb (OB) inhibitory interneurons and cerebral cortex excitatory neurons. Bioinformatic analysis validated the high reproducibility of the approach. It also demonstrated that, despite global miRNome conservation between the two cell types, a set of miRNAs, including the miR-200 family and the miR-183/96/182 cluster, is massively enriched in OB interneurons, which aligns with previous observations. In the second mouse line, T6B is fused to the postsynaptic protein PSD95. Isolation of T6B-PSD95 fractions from OB and cortical neurons identified specific sets of postsynapse-enriched miRNAs. Gene ontology analyses confirmed that these miRNAs preferentially target mRNAs related to synaptic functions.

•We present two T6B-expressing transgenic mouse lines for in vivo AGO-APP studies•CRE-dependent T6B expression enables cell-type-specific Ago-miRNA isolation•OB interneurons and cortical excitatory neurons exhibit distinct miRNA signatures•PSD95-T6B fusion uncovers postsynaptic miRNA enrichment related to synaptic genes

We present two T6B-expressing transgenic mouse lines for in vivo AGO-APP studies

CRE-dependent T6B expression enables cell-type-specific Ago-miRNA isolation

OB interneurons and cortical excitatory neurons exhibit distinct miRNA signatures

PSD95-T6B fusion uncovers postsynaptic miRNA enrichment related to synaptic genes

Deciphering miRNA expression with cell-type and subcellular resolution remains a major challenge, as current analyses rely predominantly on in vitro approaches that fail to recapitulate physiological conditions. In this study, we present two transgenic mouse lines that enable the identification and quantification of Argonaute-bound miRNAs in defined cell types of the mouse brain. This approach further allows the characterization of miRNAs within specific subcellular compartments.

Kapoor et al. present two transgenic mouse lines enabling in vivo AGO affinity purification (AGO-APP) through expression of the T6B peptide. These models allow cell-type-specific and postsynapse-associated miRNA profiling, revealing distinct miRNA populations and synaptic enrichment patterns across olfactory bulb interneurons and cortical excitatory neurons.

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