Edward Nirenberg  🇺🇦

First though we need a bit of background. The central dogma of molecular biology says that DNA encodes RNA which encodes protein. DNA can be used as a template to make DNA. RNA can be used to make more RNA. But in general, you can't go backwards from RNA to DNA.

One basic reason for this is that RNA lives in a compartment (the cytosol) separate from our genome (the nucleus) and doesn't have any ready means to go backwards unless it's associated with a protein that can facilitate that process.

There is, however, an exception. Enzymes called reverse transcriptases can use RNA as a template to make DNA. Where does it occur? The most famous example is with retroviruses like HIV, which encode a reverse transcriptase into their genome.

The reverse transcriptase makes a DNA copy of the virus's genome and the virus's RNA genome is destroyed in the process. But this is not in fact what makes HIV so persistent. That requires a second enzyme that is encoded by HIV: integrase.

Integrases cut and attach DNA into our genome. In this way, the HIV genome can persist inside cells and any daughter cells they divide into for as long as those cells live. If not for integrase, over time, HIV's genome would be lost. So what does this have to do with people?

In general, our cells have very low (generally not even detectable) reverse transcriptase activity. The little bit we do have has a few sources. One is telomerase, which is an enzyme that adds caps to our chromosomes with the aid of an RNA template because with each...

replication, our chromosomes shrink a little bit. As our cells age, telomerase activity declines and really it's only very substantial in stem cells. Telomerase is also very sequence-specific. It doesn't pick up just any RNA.

The major one of relevance in this paper is something called LINE-1. LINEs are a major part of our genome, where they have the ability to perform copy-paste. Basically, they are transcribed into mRNA in our nucleus, and exported into the cytosol wherein they produce an enzyme.

That enzyme has reverse transcriptase and endonuclease activity, which allows it to make a DNA copy of the mRNA template and paste itself into the genome. However, most LINEs in our genome are not active at all- they don't make any functional enzyme.

LINE-1 elements will also preferentially target their own mRNA sequence for retrotransposition (that's what this process is called) and not any random mRNA floating around in the cell. And once again, this is rare in any normally behaving cell.

This study claims to show that the Pfizer/BioNTech mRNA vaccine is reverse transcribed in the cell, which alone isn't meaningful because it would be lost over time if only from the cell's own natural division (but as I'll explain, there's another important factor to this).

Right off the bat, you might notice an issue. The PCR amplicon does not include the entire sequence of the RNA in question, so it can't tell us what precisely is being reverse transcribed aside from that segment.

They do not, however, at any point, demonstrate integration of the sequence in question into chromosomal DNA, which would be a critical experiment to do so that's a key deficiency. But there's another piece to this.

Furthermore, while they show that the expression of L1 rises with treatment of the Huh7 cells with the Pfizer/BioNTech vaccine, they do not show proof that L1 is mediating the reverse transcription (which would just require KOing L1 and treating with the vaccine).

I still haven't gotten to the best part yet though. Cancer cell lines like Huh7 have really crazy genomes. Huh7 is a cell line taken from the hepatoma (liver cancer) of a 57-year-old Japanese male. https://aacrjournals.org/cancerres/article/42/9/3858/487013/Growth-of-Human-Hepatoma-Cell-Lines-with …

Normally, I would show you the karyotype of Huh7 cells to show you how bizarre they look compared to human cells. However, this actually wouldn't be informative because hepatocytes divide without cytokinesis which gives them unusual chromosome numbers. https://aasldpubs.onlinelibrary.wiley.com/doi/epdf/10.1002/hep.30286 …

However, the accumulation of extra chromosome copies is thought to slow hepatocyte division so you don't get tumors, which is precisely the opposite of what happens in cancer. The point being, cancer cell genomes do not behave like normal cell genomes.

So in short, this study started with a cell that behaves in a way that essentially engineers the results they expect/want to see to make misleading claims about it. The most I can conclude from this study is a cancer cell line might reverse transcribe at least part of Bnt162b2.

However, if all of the things in this experiment as claimed were reproduced, I would not be immediately concerned by any means. By the way, we have gone round and round with something similar with SARS-CoV-2, with claims that its genome (or fragments thereof) were...

We've given out hundreds of millions of doses of these vaccines. They're safe. They prevent severe COVID-19 amazingly well, they reduce transmission, they do all we ask of them and more. Don't believe the disinformation.

@threadreaderapp please unroll