Thread by @concentricbygbw

Guess who’s back? 👀 That’s right, it’s Keith Robison (@OmicsOmicsBlog)! He’s taking the mic once again to share some lessons from SARS-CoV-2 #sequencing data.

@OmicsOmicsBlog 1/ Hi again! Keith here, with a few plots and visualizations based on @GISAID data. Here’s a 🧵 on an intriguing set of SARS-CoV-2 mutations that is catching some attention online: deletions within NSP1. Let’s dive in.

2/ Many have paid attention to viral mutations on the Spike protein, b/c it’s an important target for the immune response, the antigen used in many vaccines & the target of therapeutic monoclonal antibodies. Spike has been the most active protein in terms of mutations arising.

3/ But SARS-CoV-2 has many other proteins and these mutate too. NSP1 interacts with host ribosomes to simultaneously impede host mRNA from being translated (and even hasten their destruction) and allow viral mRNA to be translated. sciencedirect.com/science/articl…

4/ The NSPs (Non-Structural Proteins) are enzymes that perform different biochemical functions for the virus, in contrast to the structural proteins Spike, Envelope, Membrane and Nucleocapsid, which package the virus.

5/ NSPs 1 to 10 are encoded within the giant ORF1a/b proteins, which the virus makes and then chops up into individual functional proteins. ORF1b translates to over 7000 amino acids! upload.wikimedia.org/wikipedia/comm…

6/ ORF1b is also interesting because the virus “cheats” on the usual rules of translation - in one spot the ribosome slips back a base instead of moving three bases forward - a programmed ribosomal frameshift. (ORF1a results from standard translation.)

7/ The deletion which is gaining the most attention is NSP1 M85 (denoting methionine, the 85th amino acid in the protein). What’s interesting (and what makes me nervous) is that this deletion is showing up in all three major Omicron lineages – BA.1, BA1.1 and BA.2.

8/ Here’s the current data for the US, Canada, & several European nations which have reported many sequences w/ NSP1 M85del. I’m using a log scale on the Y-axis for plotting the frequency of M85del w/in a lineage, and I’m aggregating 3 recent BA.2 sublineages for these analyses.

9/ BTW, @concentricbygbw’s air travel COVID-19 genomic surveillance program, in partnership w/ @XpresCheck & @CDCgov, found the first Omicron BA.2 w/ M85del (EPI_ISL_7980710), four days before the next U.S. samples (EPI_ISL_9518849 & EPI_ISL_9326446) w/ this mutation signature!

10/ But these M85 mutations aren’t simple - there are actually a variety of different deletions or substitutions in this region, shown in separate plots of UK data below. What’s intriguing is that the different mutation signatures are showing different frequency trajectories!

11/ Here’s current data from Denmark; I’m only showing these high frequency mutations w/in BA.2 b/c it has a near monopoly there. There are some lower frequency ones to watch out for, like deletions of M85 & V86 with substitutions of E87 to Q or K, or deletions of only G82-H83.

12/ Data like this are sometimes called “natural experiments” - nature is exploring the effects of varying parts of the protein. At @Ginkgo we call our lab-based versions of these experiments “protein engineering” and we use that to tune the capabilities of proteins.

13/ Mutations in NSP1 aren’t new - we’ve seen them before. Take AY.78, a sublineage of Delta with a definitional mutation NSP1 E87D, but also some other deletions that showed up as a small fraction. But Omicron’s rise meant AY.98’s demise, and so these mutations disappeared too.

14/ NSP1 M85 deletions were also seen at very low levels in Alpha, though it seems that this “experiment” was ended by Delta pushing out Alpha.

15/ So we’re seeing Omicron repeatedly explore deletion mutations in this region of NSP1; Delta explored it less frequently and Alpha barely did at all. So what does it all mean? What can we learn from these trends?

16/ This data suggests that the NSP1 mutations interact with some other aspect of the biology of these variants, as the probability of just generating the mutations would be expected to be about the same over time.

17/ It isn’t known yet what biology this changes, but we do know that this region of the NSP1 structure is involved with binding to special sequences that are in the beginning of each SARS-CoV-2 mRNA (aka “subgenomic” RNAs).

18/ I’m not a structural biologist, but I can play w/ #PyMol! Here’s a ribbon diagram w/ the region prone to deletion (G82 to V86) rainbow-colored & labeled (@buildmodels 7K3N). These mutations chew into both a loop and a beta sheet that’s part of a “beta barrel” motif.

@buildmodels 19/ NSP1 has been explored as a drug target for SARS-CoV-2, with the general idea that blocking its interaction with viral RNAs eliminates the advantage NSP1 gives to them in terms of translation and stability.
pnas.org/doi/10.1073/pn…

20/ There’s a Cryo-EM structure of SARS-CoV-1 NSP1 in complex w/ the ribosome, but our region isn’t resolved & the key plugging up of the ribosome is done by the protein’s C-terminus. So I don’t have insight into how the deletions may alter the interaction.nature.com/articles/s4159…

21/ In addition to ribosomes, NSP1 binds to DNA primase, a key enzyme in the replication of our DNA. A structure for SARS-CoV-1 NSP1 and primase is available (7OPL). I’ve colored our interesting region in blue, the rest of NSP1 red and primase green—it doesn’t contact primase.

22/ SARS-CoV-2 mutates rapidly & in myriad ways that could affect how we detect, treat, and protect from it. That’s why it’s so important to keep doing viral genomic sequencing—a virus that likes to experiment w/ mutations calls for a dynamic “weather map” to track its evolution.