Single-cell analysis is revealing increasing diversity in gene expression profiles among brain cells. Traditional promotor-based viral gene expression techniques, however, cannot capture the growing variety among single cells. We demonstrate a novel viral gene expression strategy to target cells with specific miRNA expression using miRNA-guided neuron tags (mAGNET). We designed mAGNET viral vectors containing a CaMKIIα promoter and microRNA-128 (miR-128) binding sites, and labeled CaMKIIα cells with naturally low expression of miR-128 (Lm128C cells) in male and female mice. Although CaMKIIα has traditionally been considered as an excitatory neuron marker, our single-cell sequencing results reveal that Lm128C cells are CaMKIIα inhibitory neurons of parvalbumin or somatostatin subtypes. Further evaluation of the physiological properties of Lm128C cell in brain slices showed that Lm128C cells exhibit elevated membrane excitability, with biophysical properties closely resembling those of fast-spiking interneurons, consistent with previous transcriptomic findings of miR-128 in regulating gene networks that govern membrane excitability. To further demonstrate the utility of this new viral expression strategy, we expressed GCaMP6f in Lm128C cells in the superficial layers of the motor cortex and performed calcium imaging in mice during locomotion. We found that Lm128C cells exhibit elevated calcium event rates and greater intrapopulation correlation than the overall CaMKIIα cells during movement. In summary, the miRNA-based viral gene targeting strategy described here allows us to label a sparse population of CaMKIIα interneurons for functional studies, providing new capabilities to investigate the relationship between gene expression and physiological properties in the brain. We report the discovery of a class of CaMKIIα cortical interneurons, labeled via a novel miRNA-based viral gene targeting strategy, combinatorial to traditional promoter-based strategies. The fact that we found a small, yet distinct, population of cortical inhibitory neurons that express CaMKIIα demonstrates that CaMKIIα is not as specific for excitatory neurons as commonly believed. As single-cell sequencing tools are providing increasing insights into the gene expression diversity of neurons, including miRNA profile data, we expect that the miRNA-based gene targeting strategy presented here can help delineate many neuron populations whose physiological properties can be readily related to the miRNA gene regulatory networks.

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7726537PMC
http://dx.doi.org/10.1523/JNEUROSCI.2570-19.2020DOI Listing

Publication Analysis

Top Keywords

viral gene
20
gene expression
20
lm128c cells
20
gene targeting
16
targeting strategy
16
physiological properties
12
gene
11
cells
10
camkiiα
9
expression
8

Similar Publications

Microbial eukaryotes (aka protists) are known for their important roles in nutrient cycling across different ecosystems. However, the composition and function of protist-associated microbiomes remains largely elusive. Here, we employ cultivation-independent single-cell isolation and genome-resolved metagenomics to provide detailed insights into underexplored microbiomes and viromes of over 100 currently uncultivable ciliates and amoebae isolated from diverse environments.

View Article and Find Full Text PDF

Unlabelled: -methyladenosine (m A) is the most prevalent cellular mRNA modification and plays a critical role in regulating RNA stability, localization, and gene expression. m A modification plays a vital role in modulating the expression of viral and cellular genes during HIV-1 infection. HIV-1 infection increases cellular RNA m A levels in many cell types, which facilitates HIV-1 replication and infectivity in target cells.

View Article and Find Full Text PDF

Gammaherpesviruses are oncogenic pathogens that establish lifelong infections. There are no FDA-approved vaccines against Epstein-Barr virus or Kaposi sarcoma herpesvirus. Murine gammaherpesvirus-68 (MHV68) infection of mice provides a system for investigating of gammaherpesvirus pathogenesis and testing vaccine strategies.

View Article and Find Full Text PDF

Temporally and Spatially Controlled Age-Related Prostate Cancer Model in Mice.

Bio Protoc

January 2025

Department of Structural and Cellular Biology, Tulane University, New Orleans, LA, USA.

The initiation and progression of prostate cancer (PCa) are associated with aging. In the history of age-related PCa research, mice have become a more popular animal model option than any other species due to their short lifespan and rapid reproduction. However, PCa in mice is usually induced at a relatively young age, while it spontaneously develops in humans at an older age.

View Article and Find Full Text PDF

Modeling coding sequence design for virus-based expression in tobacco.

Synth Syst Biotechnol

June 2025

Department of Biomedical Engineering, The Iby and Aladar Fleischman Faculty of Engineering, Tel Aviv, Israel.

Transient expression in Tobacco is a popular way to produce recombinant proteins in plants. The design of various expression vectors, delivered into the plant by , has enabled high production levels of some proteins. To further enhance expression, researchers often adapt the coding sequence of heterologous genes to the host, but this strategy has produced mixed results in Tobacco.

View Article and Find Full Text PDF

Want AI Summaries of new PubMed Abstracts delivered to your In-box?

Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!