PRJNA765795: Highly efficient SARS-CoV-2 infection of human cardiomyocytes: spike protein-mediated cell fusion and its inhibition

Summary: 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 hiPSC33 CMs, smooth walled exocytic vesicles contained numerous 65-90 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, SARS42 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.

Overall Design: The hiPSC CMs were infected with mock or SARS-CoV-2/UW001/Human/2020/Wisconsin (UW-001) at MOI of 0.01 for 48hrs. Cells were lysed in Trizol and were kept at -80 ºC. Total RNA of the lysate was extracted using Direct-zol RNA Miniprep kit (R2050). Library preparation and sequencing was performed at Mayo Clinic Genome Analysis Core (GAC).