Fluorescence spectroscopy of low-level endogenous β-adrenergic receptor expression at the plasma membrane of differentiating human iPSC-derived cardiomyocytes

Philipp Gmach, Marc Bathe-Peters, Narasimha Telugu, Duncan C. Miller, Paolo Annibale*

*Corresponding author for this work

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Abstract

The potential of human-induced pluripotent stem cells (hiPSCs) to be differentiated into cardiomyocytes (CMs) mimicking adult CMs functional morphology, marker genes and signaling characteristics has been investigated since over a decade. The evolution of the membrane localization of CM-specific G protein-coupled receptors throughout differentiation has received, however, only limited attention to date. We employ here advanced fluorescent spectroscopy, namely linescan Fluorescence Correlation Spectroscopy (FCS), to observe how the plasma membrane abundance of the β1- and β2-adrenergic receptors (β1/2-ARs), labelled using a bright and photostable fluorescent antagonist, evolves during the long-term monolayer culture of hiPSC-derived CMs. We compare it to the kinetics of observed mRNA levels in wildtype (WT) hiPSCs and in two CRISPR/Cas9 knock-in clones. We conduct these observations against the backdrop of our recent report of cell-to-cell expression variability, as well as of the subcellular localization heterogeneity of β-ARs in adult CMs.
Original languageEnglish
Article number10405
Number of pages15
JournalInternational Journal of Molecular Sciences
Volume23
Issue number18
DOIs
Publication statusPublished - 8 Sept 2022

Keywords

  • GPCR
  • β-adrenergic receptors
  • hiPSC-CM
  • Cardiomyocyte
  • Fluorescence correlation spectroscopy
  • FCS
  • Fluorescence
  • CRISPR/Cas9
  • Differentiation

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