Highly Efficient SARS-CoV-2 Infection of Human Cardiomyocytes: Spike Protein-Mediated Cell Fusion and Its Inhibition

• Published in: Journal of virology 2021-11, Vol.95 (24), p.e0136821-e0136821
• Main Authors: Navaratnarajah, Chanakha K, Pease, David R, Halfmann, Peter J, Taye, Biruhalem, Barkhymer, Alison, Howell, Kyle G, Charlesworth, Jon E, Christensen, Trace A, Kawaoka, Yoshihiro, Cattaneo, Roberto, Schneider, Jay W
• Format: Article
• Language: English
• Subjects: Animals; Cathepsin B - metabolism; Cell Fusion; Chlorocebus aethiops; COVID-19 - virology; Embryonic Stem Cells - metabolism; Exocytosis; Humans; Induced Pluripotent Stem Cells - metabolism; Microscopy, Confocal; Myocytes, Cardiac - virology; SARS-CoV-2; Serine Endopeptidases - metabolism; Spike Glycoprotein, Coronavirus - metabolism; Spotlight; Spotlight Selection; Vero Cells; Viral Proteins - metabolism; Virology; Virus Internalization; Virus Replication; Virus-Cell Interactions
• ISSN: 0022-538X; 1098-5514
• DOI: 10.1128/JVI.01368-21

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 characte