RNA Modification and Processing - Online Tutor, Practice Problems & Exam Prep

mRNA Processing

Hi. In this video, we're going to be talking about mRNA modification and processing. So after transcription, RNA has to undergo through a few various processing steps before translation can occur. Because the mRNA, when it's transcribed, is not at all ready to be translated. It has to be perfected and sort of unified up so that the protein that is made from translation is correct. And so there's a few different things that happen. So the first one is that it gets this thing called a 5' cap, and this is a cap of a residue called a methylguanosine molecule. And you don't necessarily need to know what this is, but the 5' cap is just a molecule that gets added on to it, and that cap protects the RNA from degradation, and it will be important for translation, and we'll talk about how it's important for translation in the translation videos. But the first thing that happens is the 5' cap, super important. The second thing that happens is it gets a polyadenylation tail at the 3' end. What this is, it's around 150 to 200, it can be more than that, but generally it's around this many adenine nucleotides. So it's just 'A' 200 times at the very end of the transcript. And, what triggers this, there's an enzyme that adds this on. Right? And so what triggers it is there's actually a polyadenylation signal at the end of the transcript, and that's what it looks like, AAUAAA. And this is the signal that triggers the addition of the poly A tail after it. This poly tail is also super important because it allows it to be imported out of the nucleus so that it can be translated. And so here, if we have an example, so here's the coding sequence of the mRNA, we have the cap here, we have the poly A tail, and then we have these two regions that the UTR stands for untranslated regions, and we'll talk about these in other videos. But just know that here's the cap, here's the coding sequence, and here's the poly A tail. Now the third thing that happens is called splicing, and splicing removes the noncoding segments of the transcript called introns from the coding segment of the transcript called exons. Oh, I have a piece of hair. Let's get that out of there. Okay. So splicing, there's introns and this is noncoding and exons. And pretty much when you have an mRNA transcript, it looks kind of like this, where you have the coding sequences, the exons, intertwined with the introns, and that's what the entire transcript looks like. And so, splicing says, okay, we're going to cut you out, we're going to cut you out, we're going to cut you out, whoops, cut this out, and the 2 exons will come together and form a single transcript. That's splicing. So the enzyme that's responsible for this is called the spliceosome, and the spliceosome is a bunch of proteins and enzymes and RNA. It's not just one thing, and it cuts out the introns in what we call pre-mRNA, because it's not mRNA yet, it's not processed yet, to form mRNA. So the spliceosome is made up of RNAs called small nuclear RNAs. If you want to know the names, it's U1, U2, U4, U5, U6. Those are the RNAs, and it's also made up of proteins, and so we call the spliceosome, we give it a special fancy name called the small ribonuclear protein complex, or you may see it, snRNP. And this is the combination of the small nuclear RNAs and the proteins that make up the spliceosome. And so this spliceosome or the small ribonuclear protein complex comes together, and it has to recognize certain sequences in order to splice. So what it does is it recognizes a 5' splice site, which is always the GU nucleotide. It recognizes a 3' splice site, which is always AG, and you will see this as the GUAG rule, and it recognizes a third sequence called the branch point, and the branch point is just a single adenine nucleotide. So it's a single 'A' around 18 to 40 nucleotides upstream of the 3' splice site. So you have GU, you have the 'A', and you have AG. And these are 3 sequences that are found in every single splice site where the splice is always going to cut it out. So what happens actually is there's this structure called the lariat, which I'm about to show you, but it's a small circular structure that is formed through the intron excising. So what happens, you see here, so you have an exon, intron, and exon, and you can see our sites here. You have the GU, the AG, and the branch point. So what happens is the first thing is there's a cut at the GU and this forms this circular structure called a lariat. See how it's like kind of circle, it's folded back in on itself at the branch point? And eventually, this will get cut as well, this one gets cut here. And so the lariat goes off into Neverland essentially and gets degraded, and then you have this spliced mRNA. Now, so far we've been working with 2 exons, right, in my images, but you can have 100 of exons, and all of them have to be spliced. And so all of them can be spliced together, so you can have all a 100 splice, you know, one right after the other, or you can have various combinations of splicing, so you can have, you know, say 97 where 3 of them at some point have been removed and you have a new combination of exons, and that's called alternative splicing. When the exons are put together, but not necessarily in order and not necessarily all of them. So alternative splicing is a big thing that gives us a lot of different types of genetic diversity. And then another thing that happens is our final thing, it's called RNA editing. It doesn't necessarily happen that often in eukaryotes, but it's very common in prokaryotes and some lower eukaryotic organisms. And this is another form of post-transcriptional RNA processing, which is what we call anything that happens to the RNA after it's been transcribed post-transcriptional. So what happens is RNA editing is literally just changing the RNA sequence, and there's a bunch of different ways it can do that. Right? It can substitute, so a nucleotide is exchanged for another nucleotide. So if it was 'A', and now it's going to be 'U', it's substitutional. You have insertional, and that's a nucleotide added. You have the opposite, deletional, which is when it's deleted. And so all of these different things can take a sequence that may otherwise code something and change it in some way so that it's now a different sequence. It happens in bacteria very often. And so how it knows where to do this, it's not just random, it's not just random changing of nucleotides. Actually, there are RNAs called guide RNAs. These RNAs choose where the RNA editing will occur. So if we have an example like this, so here's an RNA sequence, which is here 5' to 3'. The guide RNA is on top of it, making it look like what we normally see, and this guide RNA is actually complementary to the mRNA that we're looking at changing. And this brings in proteins that will come in and it looks here, delete or add or substitute or whatever it's going to do, a nucleotide in this mRNA sequence. So that's super important, to editing that RNA. So with that, let's now move on.

Which of the following is NOT a method of mRNA modification?

• A. 5' cap
• B. 3' Poly-A tail
• C. Methylation
• D. Splicing

The spliceosome is made up of which of the following components?

• A. DNA and RNA
• B. RNA and Protein
• C. Only RNA
• D. Only Protein

Which of the following is not a sequence that the spliceosome recognizes?

• A. 5' GU
• B. 3' AG
• C. Branch Point
• D. Splice Point

After transcription the RNA sequence cannot be changed or modified before translation.

• A. True
• B. False

Here’s what students ask on this topic:

What is the purpose of the 5' cap in mRNA processing?

The 5' cap in mRNA processing is a modified guanine nucleotide, specifically 7-methylguanosine (m7G), added to the 5' end of the mRNA transcript. This cap serves several crucial functions: it protects the mRNA from degradation by exonucleases, aids in the export of the mRNA from the nucleus to the cytoplasm, and is essential for the initiation of translation. The cap structure is recognized by the ribosome and other translation initiation factors, ensuring that the mRNA is properly translated into protein. Without the 5' cap, the mRNA would be unstable and inefficiently translated.

What is the role of the poly-A tail in mRNA?

The poly-A tail is a stretch of adenine nucleotides, typically 150-200 bases long, added to the 3' end of the mRNA transcript. This tail is crucial for several reasons: it enhances the stability of the mRNA by protecting it from exonuclease degradation, aids in the export of the mRNA from the nucleus to the cytoplasm, and plays a role in the initiation of translation. The polyadenylation signal (AAUAAA) in the mRNA sequence triggers the addition of the poly-A tail. This modification ensures that the mRNA is stable and efficiently translated into protein.

What is alternative splicing and why is it important?

Alternative splicing is a process during mRNA processing where different combinations of exons are joined together, resulting in multiple mRNA variants from a single gene. This process is facilitated by the spliceosome, which recognizes specific splice sites and can include or exclude certain exons. Alternative splicing is important because it increases genetic diversity and allows a single gene to produce multiple protein isoforms with different functions. This diversity is crucial for the complexity of eukaryotic organisms and allows for more versatile and adaptable biological processes.

How does RNA editing differ from other forms of RNA processing?

RNA editing is a post-transcriptional process that alters the nucleotide sequence of an RNA molecule, distinct from other RNA processing steps like capping, polyadenylation, and splicing. RNA editing can involve substitutions, insertions, or deletions of nucleotides, and is guided by complementary RNAs known as guide RNAs. This process can change the coding potential of the mRNA, leading to the production of different protein variants. While RNA editing is common in prokaryotes and some lower eukaryotes, it is less frequent in higher eukaryotes. It adds another layer of regulation and diversity to gene expression.

What is the spliceosome and how does it function in mRNA splicing?

The spliceosome is a complex of small nuclear RNAs (snRNAs) and proteins, collectively known as small nuclear ribonucleoproteins (snRNPs). It is responsible for the removal of introns from pre-mRNA during splicing. The spliceosome recognizes specific sequences at the 5' splice site (GU), the 3' splice site (AG), and the branch point (a single adenine nucleotide). It then cuts the pre-mRNA at these sites, forming a lariat structure with the intron, which is subsequently degraded. The exons are then joined together to form the mature mRNA, ready for translation.