Multiplex live single-cell transcriptional analysis demarcates cellular functional heterogeneity.
Ayhan AtmanliDongjian HuFrederik Ernst DeimanAnnebel Marjolein van de VrugtFrançois CherbonneauLauren Deems BlackIbrahim John DomianPublished in: eLife (2019)
A fundamental goal in the biological sciences is to determine how individual cells with varied gene expression profiles and diverse functional characteristics contribute to development, physiology, and disease. Here, we report a novel strategy to assess gene expression and cell physiology in single living cells. Our approach utilizes fluorescently labeled mRNA-specific anti-sense RNA probes and dsRNA-binding protein to identify the expression of specific genes in real-time at single-cell resolution via FRET. We use this technology to identify distinct myocardial subpopulations expressing the structural proteins myosin heavy chain α and myosin light chain 2a in real-time during early differentiation of human pluripotent stem cells. We combine this live-cell gene expression analysis with detailed physiologic phenotyping to capture the functional evolution of these early myocardial subpopulations during lineage specification and diversification. This live-cell mRNA imaging approach will have wide ranging application wherever heterogeneity plays an important biological role.
Keyphrases
- single cell
- binding protein
- living cells
- high throughput
- rna seq
- pluripotent stem cells
- gene expression
- single molecule
- fluorescent probe
- genome wide identification
- genome wide
- transcription factor
- copy number
- endothelial cells
- dna methylation
- stem cells
- oxidative stress
- poor prognosis
- fluorescence imaging
- heart failure
- cell cycle arrest
- genome wide analysis
- induced pluripotent stem cells
- cell therapy
- mesenchymal stem cells
- pet imaging
- cell death
- atrial fibrillation
- nucleic acid
- data analysis