1/ Here is day 3 of your twitter #SARSCoV2 genetics / translation course ;-)
On day 1 we saw how the virus used a polyprotein that can be cleaved into proper proteins, and on day 2 we saw how the virus can make our ribosome slip and read through a stop codon.
On day 1 we saw how the virus used a polyprotein that can be cleaved into proper proteins, and on day 2 we saw how the virus can make our ribosome slip and read through a stop codon.
2/ Today, we’ll see how all the open reading frames on the last third of the #SARS-CoV2 genome get translated which will create the important structural proteins such as the famous S spike and the essential E envelope, M Membrane and N Nucleoprotein. 
3/ These important structural proteins (&others) are encoded by the last 1/3 of the genome. Bec. translation always starts from 5’, these genes are “hidden” behind the final stop codon of ORF1ab, and they don’t get translated from the RNA that initially enters the cell.
4/ So how does the virus get these genes translated? (And, yes, I know I talk about the virus as if it has a will. It doesn’t, but it makes it easier to tell the story).
5/ Here’s what happens. Remember the RdRp, RNA-dependent RNA polymerase? RdRp is encoded by the ORF1b (after the ignored stop) and it can synthesize RNA from RNA. RdRp starts to create RNA using the original RNA as a template. It’ll do that starting from the 3’ end of the genome.
6/ Reminder: Synthesis of RNA (or DNA) is always done w a strand as a template, creating the complement of the template. If the original RNA had sequence 5’ ACUUGAC 3’ then the new strand of RNA will be 3’ UGAACUG 5’ (A complements U and C complements G).
7/ If you copy 3’ UGAACUG 5' again, you get the original 5’ ACUUGAC 3’ sequence back.
8/ So, RNA synthesis starts and all goes normal initially, it may even be that the RdRp just synthesizes the entire 30,000 nucleotide long genome (the complement, that is, a – strand) – but often, the RdRp at some point jumps from one part of the genome to another part.
9/ Specifically, just left (i.e., on the 5’ end) each of the ORFs like S, M and E, there is a sequence called a TRS (transcription regulatory sequence). This sequence is also present just right (i.e., on the 3’ end) of the Leader sequence.
10/ I didn’t tell you about the Leader sequence yet, but what happens with the jumping is that you get a chimera of the 3’ end of the genome glued to the Leader sequence from the 5’ end of the genome.
11/ Now there is a – strand RNA with the S next to the L. It is copied again to make a +strand of the same length. This "subgenomic" RNA can now be translated. The first gene after the Leader sequence will be translated (here S), until a stop codon is met.
12/ The leader sequence is like a signal for the ribosome to start translating. Because the jumping can happen anywhere where there is a TRS sequence, you get a whole series of “nested” subgenomic RNAs. From each of them the 5’ most ORF will be translated.
13/ So to recap: RNA-dependent RNA polymerase starts copying the RNA from the 3’ end of the genome but sometimes “jumps” to the leader sequence creating a set of chimeras with L next to different genes.
14/ Once this RNA is copied again to make + strand mRNA, whichever gene is next to L gets translated. Because the jumping can happen after each gene, each gene has a chance to end up next to L.
15/ What I find intriguing about the RdRp jumping, is that it should work some of the time, but not always. If the RdRp would ALWAYS jump just after S, then you’d never get a full length genome and no new infectious viral particles would ever be made. (That’d be good!)
16/ Thanks for reading! Now I just have to translate tweets into slides and I have my class for Thursday ready ;-)
And thank you all for staying home, it looks like some of the curves are slowly flattening!
And thank you all for staying home, it looks like some of the curves are slowly flattening!
17/ and Dr Britt Glaunsinger does a great job telling you the same story which took me 3 days in 10 minutes a video (minute 20-30)
