Highly Efficient SARS-CoV-2 Infection of Human Cardiomyocytes: Spike Protein-Mediated Cell Fusion and Its Inhibition.
Chanakha K Navaratnarajah, David R Pease, Peter J Halfmann, Biruhalem Taye, Alison Barkhymer, Kyle G Howell, Jon E Charlesworth, Trace A Christensen, Yoshihiro Kawaoka, Roberto Cattaneo, Jay W Schneider, Wanek Family Program for HLHS-Stem Cell Pipeline
Author Information
Chanakha K Navaratnarajah: Department of Molecular Medicine, Mayo Clinicgrid.66875.3a, Rochester, Minnesota, USA. ORCID
David R Pease: Discovery Engine/Program for Hypoplastic Left Heart Syndrome, Mayo Clinicgrid.66875.3a, Rochester, Minnesota, USA.
Peter J Halfmann: Influenza Research Institute, Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin-Madison, Madison, Wisconsin, USA.
Biruhalem Taye: Department of Molecular Medicine, Mayo Clinicgrid.66875.3a, Rochester, Minnesota, USA.
Alison Barkhymer: Department of Molecular Medicine, Mayo Clinicgrid.66875.3a, Rochester, Minnesota, USA.
Kyle G Howell: Mayo Microscopy and Cell Analysis Core, Mayo Clinicgrid.66875.3a, Rochester, Minnesota, USA.
Jon E Charlesworth: Mayo Microscopy and Cell Analysis Core, Mayo Clinicgrid.66875.3a, Rochester, Minnesota, USA.
Trace A Christensen: Mayo Microscopy and Cell Analysis Core, Mayo Clinicgrid.66875.3a, Rochester, Minnesota, USA.
Yoshihiro Kawaoka: Influenza Research Institute, Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin-Madison, Madison, Wisconsin, USA.
Roberto Cattaneo: Department of Molecular Medicine, Mayo Clinicgrid.66875.3a, Rochester, Minnesota, USA. ORCID
Jay W Schneider: Discovery Engine/Program for Hypoplastic Left Heart Syndrome, Mayo Clinicgrid.66875.3a, Rochester, Minnesota, USA. ORCID
Severe cardiovascular complications can occur in coronavirus disease of 2019 (COVID-19) patients. Cardiac damage is attributed mostly to the aberrant host response to acute respiratory infection. However, direct infection of cardiac tissue by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) also occurs. We examined here the cardiac tropism of SARS-CoV-2 in spontaneously beating human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). These cardiomyocytes express the angiotensin-converting enzyme 2 (ACE2) receptor but not the transmembrane protease serine 2 (TMPRSS2) that mediates spike protein cleavage in the lungs. Nevertheless, SARS-CoV-2 infection of hiPSC-CMs was prolific; viral transcripts accounted for about 88% of total mRNA. In the cytoplasm of infected hiPSC-CMs, smooth-walled exocytic vesicles contained numerous 65- to 90-nm particles with canonical ribonucleocapsid structures, and virus-like particles with knob-like spikes covered the cell surface. To better understand how SARS-CoV-2 spreads in hiPSC-CMs, we engineered an expression vector coding for the spike protein with a monomeric emerald-green fluorescent protein fused to its cytoplasmic tail (S-mEm). Proteolytic processing of S-mEm and the parental spike were equivalent. Live cell imaging tracked spread of S-mEm cell-to-cell and documented formation of syncytia. A cell-permeable, peptide-based molecule that blocks the catalytic site of furin and furin-like proteases abolished cell fusion. A spike mutant with the single amino acid change R682S that disrupts the multibasic furin cleavage motif was fusion inactive. Thus, SARS-CoV-2 replicates efficiently in hiPSC-CMs and furin, and/or furin-like-protease activation of its spike protein is required for fusion-based cytopathology. This hiPSC-CM platform enables target-based drug discovery in cardiac COVID-19. Cardiac complications frequently observed in COVID-19 patients are tentatively attributed to systemic inflammation and thrombosis, but viral replication has occasionally been confirmed in cardiac tissue autopsy materials. We developed an model of SARS-CoV-2 spread in myocardium using induced pluripotent stem cell-derived cardiomyocytes. In these highly differentiated cells, viral transcription levels exceeded those previously documented in permissive transformed cell lines. To better understand the mechanisms of SARS-CoV-2 spread, we expressed a fluorescent version of its spike protein that allowed us to characterize a fusion-based cytopathic effect. A mutant of the spike protein with a single amino acid mutation in the furin/furin-like protease cleavage site lost cytopathic function. Of note, the fusion activities of the spike protein of other coronaviruses correlated with the level of cardiovascular complications observed in infections with the respective viruses. These data indicate that SARS-CoV-2 may cause cardiac damage by fusing cardiomyocytes.
