Transcription

Transcript

[0:00:00]
Have you ever cooked? I mean really cooked, not just toss a pop tart into a toaster. If you have, you know it’s a pretty messy process, especially if your chef that’s doing your cooking is a moron like my friend Galen over here. I’ve got a pretty expensive cook book here. If I hand that to him, he’s such a moron he’ll just start cooking anything, not essentially what I want him to cook. He’s pretty messy. He’s going to sit there and he’s going to start spilling stuff all over the book. It could get ruined.

Instead, if I just want one recipe made, what I should do, is I should hand him a photocopy of the recipe I’m interested in. Then he’s at least able to read. He can follow instructions. If he makes any damage to this, if he spills oil on it, sets it on fire, I don’t care because my cook book is still intact.

Your cells do a similar process when they’re trying to make proteins. The process of protein synthesis can actually occasionally create these things called free radicals, and other problematic chemical reactions. You don’t want your DNA, which is kind of like your cookbook, which is the one copy of the instructions you have on building and running the cell, you don’t want that damaged. So instead, what you do is you make a RNA copy of just the recipe that you’re interested. Then, you can send that photocopied transcript of your gene that you’re interested in, off to the idiot ribosomes that can’t follow the instructions. If that photocopy, that ‘messenger RNA’ transcript gets damaged, it’s not a big loss, because you can always make more.

So today what I’m going to do is, I’m going to go through the process of transcription. I’m going to go over how does it get started, and how does the enzymes involved in transcription know where they should be copying. Then I’m going to go into how does that RNA polymerase, the enzyme that is building the messenger RNA, how does it go ahead and build the RNA.

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Last I will describe what happens when the whole process comes to a halt, and how does the RNA get prepared to be sent out of the cell.

So before we get too far into this, I want to stop for a moment and make sure you understand why is this thing called Transcription? I really want to focus on this because, transcription is the first half of what’s called protein synthesis. The second half of protein synthesis is called translation. I’ve heard a lot of students go trans-. They don’t pay attention to the end part.

So let’s think, what is a transcript? Well, if you’ve ever paid attention, a transcript is actually a written record of usually something that’s spoken. For example, in court cases, you’ve got somebody called the stenographer who is typing down everything that anybody says in the court room. Now, that written record of what’s being spoken, is in this exact same language, just a slightly different version. Because you guys know that spoken English is a little bit different from written English.

For example, comma; very rarely do I use punctuation marks like comma, or period, comma or colon in my spoken English, period. The same thing with transcription. What we’re doing is we’re using the exact same language. We’re using the nucleic acid language, but we’re trying to copy one section of DNA. We’re going to do it in a slightly different version called RNA. They’re both nucleic acid sequences. So we’re going to be using RNA to make a photocopy of a section of DNA. We’re making a transcript, a copy of one small section.

So how does transcription begin? Let’s take a look.

Every gene, remember is the set of instructions on how to build a protein. It has at its beginning a section called the promoter. What is the promoter? The promoter is the section of DNA that has a sequence of As, Ts, Cs, and Gs that an enzyme will come along, and it will recognize this is the beginning of a gene.

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So the promoter helps attract that enzyme. Now how does the enzyme and the transcription factors that help it, how does it know that this is a promoter? Well, it has specific sequences in it. A common sequence in many promoters is the sequence TATA or it’s called the TATA box. It’s actually this sequence here. So that is one of the clues to the RNA polymerase, the molecule or enzyme that’s going to be building our transcript of our DNA. It’s one of the clues that it uses to recognize that this is a promoter.

Now to help you understand this, imagine I was trying to photocopy one recipe out of my cookbook. How do I know where a particular recipe begins? Well, I just keep flipping until I find the title of the recipe that I’m interested. Now, how did I recognize that this is the title of the recipe that I’m looking for? Well, notice it’s got different colors than the rest of it.

Now, you remember that genes are sequences of As, Ts, Cs, and Gs that are used to instruct ribosomes on how to build proteins. Each sequence of letters indicating which amino acid goes first, which amino acid goes second, third, fourth and fifth and so on and so forth. Well, what’s a recipe? It’s a sequence of instructions written not in A, T, C and G, but written in all these letters A through Z. And that, they tell the chef which ingredient to add first, which ingredient to add second, third, fourth, and so on and so forth.

So again, the RNA polymerase recognizes this sequence as a promoter. I recognize any of the colored sequences of letters as a title to a recipe. Slight differences in the promoter, just like slight differences in this title here, tell me which specific recipe it is that I’m looking at.

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So that way, I can read this recipe and say Espresso Beef Tenderloin Fillet. That’s very different than if I was reading something that said Angel Food Cake. So the promoter for one protein, will have slight differences that indicate which protein it is, versus a different protein. So if I wanted to build insulin or if I wanted to build something like myosin.

So the enzyme that does this like I said is RNA polymerase. So what happens is that it lands on the DNA. Let’s go ahead and take a look at that. Here is our promoter, here is the actual gene. So the promoter is before the gene itself. A lot of times they’ll call that up strand. That just means it’s before. So here this little purple blurb is our RNA polymerase. Now I’ve not drawn it in, because it gets pretty complicated, but there’s a bunch of little helper molecules. These are factors.

Remember in AP Biology, in Science in general, when scientists didn’t originally know what a molecule is, and they just don’t even have evidence for how many of them are, they just call them factors. Later on they’ll name them. Well, there’s a bunch of transcription factors that help identify this is a promoter for a specific gene.

Let’s suppose we want to make insulin. Well, there’s a promoter that is specific to insulin and transcription factors will help guide in the RNA polymerase when insulin is needed, to the promoter of insulin. That also tells it which of these two strands do we wish to copy. Do we want to copy the red one, or the blue one? It’s the promoter that tells you. Just like if I had opened up my recipe book, and it upside down, looking at the title oops I had it upside down, now it’s right side up. That’s how we begin transcription.

So now that the RNA polymerase has been brought to the promoter, by the transcription factors, and it recognized the correct promoter that it wants to begin copying. RNA polymerase does a lot of the jobs that you saw in DNA Replication. It does all those tasks pretty much by itself. It goes ahead and it opens up the helix. So let’s take a look at that.

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Now we’ve opened up the helix, but we’re only interested in copying one side of the DNA. We’re only interested in copying this strand. So what happens is that, now that we’ve got it open, we can have RNA nucleotides that are floating around inside the nucleus. They start matching up following the same basic Base Pairing rules that we’ve seen before. If we have an A here, normally, we’d be putting a DNA T. But you’ve got to remember, one of the differences between RNA and DNA, is DNA uses Thymine. In its place RNA uses a Pyrimidine called Uracil. That’s a Nitrogenous base called Uracil that is used in place of Thymine.

How do you remember that? Uracil is used by RNA? Well, how would you abbreviate Uracil? U. How would you abbreviate RNA even more? R. Just think U, R, correc,t and you will be.

So what happens is that we have, wherever there’s an A, we’ll put an RNA U. Wherever there’s a G, we’ll put an RNA C. Wherever there’s a T, we’ll put an RNA A. Wherever there’s a C, we’ll put an RNA G. So these RNA nucleotides start landing and that’s what we’ll see next.

RNA polymerase goes along and joins those sugars and phosphates together, building in the 5' to 3'direction. Now we’re beginning to run along the DNA and we’re building ourselves this green RNA copy of the DNA sequence. Again, it’s following the same basic Chargaff’s Base Pairing rules. Now, RNA polymerase hits this fork here. What does it do? It just keeps going. As it opens up more of the helix, the portion of RNA that’s already being copied a fair amount, starts to fall off. Because again, one of the differences between RNA and DNA, DNA is very stable in a double helical form, RNA is typically single stranded. Because it has additional oxygen in its ribose sugar, it tends to repel itself if we have long stretches of double stranded RNA.

So this messenger RNA that we’re making, starts to fall away from the DNA as the RNA polymerase continues. The portions of DNA that had previously been copied now begin to close up. That’s how we do it.

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So this just keep happening until the RNA polymerase reaches the end of the gene, and then it just stops. So how does it know what the end of the gene is? What happens is that, just like remember there is the promoter, tha was a sequence of As, Ts, Cs, and Gs that indicated this is the beginning. How does it know where there is the end? Well, we have what are called Termination Sequences. These are sequences of As, Ts, Cs, and Gs that indicate to the RNA polymerase you’ve reached the end.

Now, if this was a prokaryote, as the RNA polymerase falls off, because it’s reached the Termination Sequence. And the messenger RNA floats off, because it’s done being made, in a prokaryote or bacteria we could have ribosomes already landing on this and making our protein follow the instructions of the messenger RNA. But with eukaryotes, creatures like yourselves or plants or muskrats, there is a lot more complications. So let’s take a look at what happens.

Before the RNA is allowed out of the nucleus, a number of things happen. First, remember, viruses use RNA, some of them like HIV, viruses use RNA to take over your cells. So when they inject their RNA into your cytoplasm, you’ve got enzymes that are on the alert destroying any random RNA that’s floating around.

Now you want to protect your RNA from being chopped up. So what you is on the 5' beginning end, you put in a modified Guanine. This is called the 5'-cap. That’s what we see here. It’s a backward Guanine that’s put in here. That protects the enzymes. Normally the enzymes will come along and kind of like Pacman on Steroids. but it goes, and floats away disappointed.

On the other end, we put in a whole bunch of As. When I say a whole bunch of As, I mean several hundred perhaps. Now scientists don’t like to say a whole bunch of, so what they do is they call it a Poly A tail, because that sounds cooler.

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Now why not just make both ends of the messenger RNA invulnerable to our chopping up enzyme? Because our chopping of enzymes can attack from this end. Well, remember this messenger RNA is going to give instructions on how build a protein. If we go back to our analogy from the beginning, where I talked about how we’re copying a recipe for our moron chef, he reaches the end of the recipe. And he’ll just sit there and go,"Oh look a recipe," and he’ll keep going. And he’ll just keep making it. If you hand your chef this recipe, and say I need some cake. And he begins going okay and he starts adding in the things that you’ve told him to, well, we’ve got ourselves a problem. Because he’ll just keep making cake until he dies.

Say for example if I suddenly did something, I’ve just startled some of you at least. Right now, adrenaline is being dumped into your body from your adrenal glands. It’s going through your circulatory system and flooding through your body. Some of you even felt a little de frisson to use some French, a little shiver. That goes through your body as the adrenaline starts hitting your cells.

Well, in your adrenal glands, genetic signals are being activated right now. We’re starting to make messenger RNA to code for the proteins that are going to build the replacement adrenaline for what you just dumped into your body. Do you want that section of RNA, that instructions to build adrenaline in your body for the rest of your life? Do you want to be in the restaurant walking and twitching from adrenaline overload, because I startled you once so many years ago? No. So instead, if we go back to our recipe analogy, let’s go ahead and we’ll attach a fuse to the recipe. We’ll make it long enough so that when I light the fuse, it’ll take maybe 10 minutes or so for the fuse to burn until it hits our photocopied paper of the recipe.

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So the moron chef will sit and go, "Okay I add these things," Then oh it’s gone. So he builds enough of the proteins or builds enough of the cake that we want, but it’s destroyed. So this poly-A tail gets something for our enzymes to chew on, until finally they hit the actual messenger RNA that’s being used to build our protein. Then that destroys it. That means we make enough of the protein, but not too much.

Now you’ll also notice that between this and there, there are some things missing. That’s because while bacterial messenger RNA goes straight in the cytoplasm can be converted, our DNA has a lot of extra intervening regions inside the genes. These intervening regions are called introns. Those portions of the messenger RNA that are actually going to be expressed or expressed regions, those are called exons. So you have enzymes that recognize the introns and cut them out. So we cut out these intron, cut out these intron, and then splice together the remaining exons. The group of enzymes that do this is called a spliceosome which literally means a splicing body or object.

Why have these introns? Well, there’s a number of proposed reasons. Scientists used to be completely confused, and they’d just call it ‘Junk DNA'. But scientists discovered these introns can actually have some significant importance. Now sometimes, these introns may be left over viruses that added their DNA to ours. Hopefully, mutations have happened and deactivated this pro-viruses they’re called. But a lot of times it turns out that the introns may be involved in how to regulate the proteins, such that, sometimes you splice up the introns, sometimes you don’t. That gives you from one messenger RNA primary transcript.

If let’s suppose I want to keep this intron, but get rid of that. Other times I’ll keep this one, get rid of that one. A third time, I’ll get rid of both.

[0:16:00]
That gives me three possible outcomes from one original primary transcript. Another reason to have the introns, if we go back to the DNA, remember these represent sequences of As, Cs, Ts, and Gs. While the exons represent sequences of As, Cs, and Gs that are actually used, the introns are not. If you go back to meiosis, remember during meiosis one, Prophase one, there is the stuff called crossing over where pieces of mummy’s DNA are swapped with pieces of daddy’s DNA. These introns get places where it can break a gene without actually damaging the portions that are used to code for functional proteins. It allows more variation in the proteins that are being created in offspring.

Now, this splicing let’s go back and finish up with our metaphor of the chef. Now, your cookbook, instead of being just very simple basic cookbook like the DNA of a prokaryote or a bacteria, your cookbook is chock full of ads. If you just photocopy straight the recipe with the included ads, and you hand it to your moron chef who’s our representation of the ribosome, he’ll begin by saying, "Okay step one add flour. Step two, interest in refinancing your house okay." He’ll be on the phone, and you’ll be massively in debt. That’s not making your cake.

So instead you cut out and paste together. Just the sections of the photocopy that actually contain the instructions of the recipe that you want them to follow. Add in your fuse, your poly-A tail, put a 5'-cap on the other end, and now we’ve finished post transcriptional editing. That’s it for transcription.

Let’s run through this again, one last time just to make sure you’ve got it all down. So transcription’s all about making a messenger RNA copy of a single gene, so that that messenger RNA copy can go to the ribosome, and it can follow the instructions there to build ourselves a protein.

[0:18:00]
It begins with promoter sequences helping identify, along with transcription factors, help that RNA polymerase identify where the beginning of the gene is. That RNA polymerase opens up the DNA double helix, and begins adding RNA nucleotides in a long sequence until it hits the Termination Sequence, which identifies the end of the gene. Next that RNA copy leaves. If it’s a eukaryote, undergoes some editing to add in the 5'-cap and the poly-A tail also to get rid of all those intervening regions or introns. That then goes off to the ribosome where we can begin translation.

If you can remember that, and just vomit up what I just said, maybe 3 or 4 sentences, you’ll get a better score on a transcription essay than most people do nationwide. And you’re on your way to getting a good grade on the AP. I highly recommend that if you want to maximize your score however, most essay questions they involve both transcriptions and translations. So I highly recommend you either use your textbook to read about translation, or watch my video on Translation to give yourself a good solid score. But otherwise, I’m pretty sure you’ll do just fine.

Tags
Transcription mRNA promoters exons/introns protein synthesis