Study of viral-host interactions underlying LINE1-mediated viral RNA retrotransposition using RNA-seq

SARS-CoV-2 sequences can be reverse-transcribed and integrated into the genomes of virus-infected cells by a LINE1 mediated retrotransposition mechanism. Whole genome sequencing (WGS) methods detected retrotransposed SARS-CoV-2 subgenomic sequences in virus-infected cells overexpressing LINE1, while an enrichment method (TagMap) identified retrotranspositions in cells that did not overexpress LINE1. LINE1 overexpression increased retrotranspositions about 1,000-fold as compared to non-overexpressing cells. Nanopore WGS can directly recover retrotransposed viral and flanking host sequences but its sensitivity depends on the depth of sequencing (a typical 20-fold WGS depth would only examine 10 diploid cell equivalents). In contrast, TagMap enriches for the host-virus junctions and can interrogate up to 20,000 cells and therefore is able to detect rare viral retrotranspositions in LINE1 non-overexpressing cells. Although Nanopore WGS is 10 - 20-fold more sensitive per tested cell, TagMap can interrogate 1,000 - 2,000-fold more cells and thus can detect rare retrotranspositions. When comparing SARS-CoV-2 infection and viral nucleocapsid mRNA transfection by TagMap, retrotransposed SARS-CoV-2 sequences were only detected in infected cells but not in transfected cells. This correlated with a significantly higher viral RNA level and LINE1-inducing cellular stresses in virus-infected as compared to RNA-transfected cells, which may facilitate LINE1-mediated retrotranspositions in virus-infected cells.