Reverse-transcribed SARS-CoV-2 RNA can integrate into the genome of cultured human cells and can be expressed in patient-derived tissues

The study “Reverse-transcribed SARS-CoV-2 RNA can integrate into the genome of cultured human cells and can be expressed in patient-derived tissues” provides significant insights into the behavior of SARS-CoV-2, particularly concerning its RNA’s potential to integrate into the human genome.

Here’s a detailed summary:

Background: The study addresses the prolonged detection of SARS-CoV-2 RNA in patients after COVID-19 recovery, with some cases showing recurrent positive PCR tests without shedding infectious virus. This led to the investigation of whether SARS-CoV-2 RNA could integrate into human DNA and contribute to these positive tests​​.

Methods and Findings:

  • Researchers used techniques like Nanopore long-read sequencing, Illumina paired-end whole genomic sequencing, and Tn5 tagmentation-based DNA integration site enrichment sequencing. These methods provided evidence that SARS-CoV-2 sequences can integrate into the host cell genome.
  • The study involved transfecting HEK293T cells with LINE1 expression plasmids before infecting them with SARS-CoV-2. DNA from these cells was isolated and analyzed, revealing DNA copies of SARS-CoV-2 sequences in the infected cells, suggesting that the virus’s RNA can be reverse-transcribed and integrated into the host cell genome​​.

Expression of Viral Sequences:

  • Researchers analyzed published RNA-seq data from SARS-CoV-2-infected cells and found human-viral chimeric reads, indicating the expression of integrated viral sequences. These chimeric reads were present in various sample types, including cultured cells and organoids from different tissues. The abundance of these reads correlated with viral RNA levels​​.

Discussion and Implications:

  • The study presents evidence that SARS-CoV-2 sequences can be reverse-transcribed and integrated into human DNA. This was supported by the detection of “human-viral-human” chimeric reads and LINE1 endonuclease recognition sequences at the integration sites, indicative of LINE1 retrotransposon-mediated integration.
  • Approximately 30% of the viral integrants analyzed in cultured cells did not have a recognizable LINE1 endonuclease recognition site nearby, suggesting the possibility of alternative integration mechanisms.
  • The study noted the challenge in demonstrating the presence of integrated SARS-CoV-2 sequences in patient tissues due to the expected low frequency of cells carrying such integrations. However, the global scale of SARS-CoV-2 infections implies that even rare events could be clinically relevant.
  • The integration and expression of viral sequences in the form of chimeric RNAs raise questions about their potential role in the disease course and their impact on the immune response. The study speculates that cells with integrated and expressed SARS-CoV-2 sequences might continuously stimulate the immune system, potentially triggering protective responses or conditions like autoimmunity.
  • This research could have implications for antiviral therapies and PCR tests, as viral transcripts derived from integrated viral DNA sequences might be detected by highly sensitive PCR assays, not necessarily indicating active viral replication​​.

In summary, this study sheds light on a potential mechanism by which SARS-CoV-2 can integrate its RNA into the human genome, contributing to prolonged viral RNA detection and possibly influencing the clinical outcomes and immune responses in patients post-COVID-19 recovery.

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