Multiplex live single-cell transcriptional analysis demarcates cellular functional heterogeneity

Author:

Atmanli Ayhan123ORCID,Hu Dongjian124,Deiman Frederik Ernst12,van de Vrugt Annebel Marjolein12,Cherbonneau François1,Black Lauren Deems35,Domian Ibrahim John126

Affiliation:

1. Cardiovascular Research Center, Massachusetts General Hospital, Boston, United States

2. Harvard Medical School, Boston, United States

3. Department of Biomedical Engineering, Tufts University, Medford, United States

4. Department of Biomedical Engineering, Boston University, Boston, United States

5. Sackler School of Graduate Biomedical Sciences, Tufts University School of Medicine, Boston, United States

6. Harvard Stem Cell Institute, Cambridge, United States

Abstract

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.

Funder

American Heart Association

National Heart, Lung, and Blood Institute

Publisher

eLife Sciences Publications, Ltd

Subject

General Immunology and Microbiology,General Biochemistry, Genetics and Molecular Biology,General Medicine,General Neuroscience

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