Cell Biology | DNA Transcription

What's up, ninja nerds, in this video today? Let's talk about DNA

transcription

before we begin. If you like this video, hit the like button, comment in the comments section and also subscribe in the description box we have. links to our facebook instagram patreon that will all be there alright ninjas let's get into it alright ninja so with DNA

transcription

we have to have a basic understanding of the definition of what the heck transcription is and it's really a simple thing. take DNA, okay, double stranded DNA with eukaryotic

cell

s and even in prokaryotic

cell

s and turn it into RNA, so you take DNA and you make RNA, that's all transcription, but for transcription to occur, we need two particular types. of proteins or enzymes, if you will, to facilitate this process and I want to talk about them very quickly because they are very important now, the transcription can be a little different, that's okay and it is important to know the differences between prokaryotic cells, we will consider bacteria In this In the case of eukaryotic cells, human cells like you and me, in prokaryotic cells, there is a particular type of protein that is needed for transcription to take place.

What is that protein? So let's say we take this DNA strand here, we have this DNA strand. In this strand of DNA we have these blue portions that I have highlighted here as a box with some lines. This here for now. I want you to know what is called a promoter. This is called the promoter region. Now a promoting region is. a particular nucleotide sequence within the DNA and what that does is it allows particular proteins like RNA polymerases and transcription factors to bind to the DNA and then start moving through the DNA taking the DNA and making RNA, so that's the first thing you need to know.

Within the DNA there is a particular sequence of nucleotides that we will talk about a little later called the promoter region and that is the first thing we must identify. Let's say we take this particularly for prokaryotic cells, that is, prokaryotic cells and let's just say like a bacterial cell, okay, prokaryotic cells use a very particular type of enzyme, what is that enzyme? What is this and I will show you its structure. A basic structure of what the RNA polymerase holoenzyme is. It is made up of two things. One of the components of this enzyme is called the core enzyme and the core enzyme of this RNA polymerase holoenzyme.

It consists of multiple subunits that they love to ask you about on your exams and other exams, and these contain two alpha units, well, two alpha chain proteins, it contains two beta units, technically we say beta and main beta if you really want to be specific. and then one more called omega unit. Well, these are the main components of the core enzyme that makes up RNA polymerase. What's important to remember is that these are the ones that will actually read the DNA and produce RNA in that portion of the enzyme. reads DNA and produces RNA the next component of the RNA polymerase holoenzyme is the portion we need to attach the DNA to the promoter region without it we will not be able to allow this RNA polymerase to bind to the DNA and transcribe it, this is called right sigma or you can represent it as this subunit or factor if you accept these two components, the core enzyme which is made up of two alpha, beta and main beta and also the omega subunit.

Since the sigma subunit makes up the entire RNA polymerase, now let me show you, for example, here, let's say I represent the core enzyme as this kind of blue circle with lines and then we'll represent the sigma subunit as a kind of pink circle with some lines on the side. right, so let's imagine that here we have that central enzyme that we are going to represent like this and then the other component, which is the sigma subunit, which will represent like this to which the sigma subunit will then join. the promoter region, once it binds to the promoter region, this core enzyme of RNA polymerase will release from the sigma subunit and it will start moving down this DNA and as it goes down the DNA it will read the DNA from three to five and synthesize a strand of RNA from which we will talk in more detail later five to three to read the DNA and produce RNA.

This RNA that we make in prokaryotic cells with the RNA polymerase holoenzyme is very different from eukaryotic cells in prokaryotic cells that mRNA that we made from this RNA polymerase holoenzyme can make all the RNA that we need, whether it's RNA inside the prokaryotic cell , either RNA within the prokaryotic cell or RNA within prokaryotic cells. So, that's a very important thing that I really need you to know that prokaryotic cells use an RNA polymerase called a holoenzyme made up of two components, a core enzyme made of these subunits and a sigma subunit. The core is what is read.

DNA and produces RNA, the sigma subunit is what binds RNA polymerase to the promoter region, which allows it to transcribe DNA well and every time RNA is produced within a prokaryotic cell from this RNA polymerase, it is It produces all the RNA within that prokaryotic cell in a eukaryote. cells is a little bit different, so let's talk about that, let's say here we have three promoter regions that I want us to focus on and this is all within eukaryotic cells, in eukaryotic cells we need two different things to allow the transcription and This portion here on the right and this portion of prokaryotic cells we only need one enzyme that had two different components within the eukaryotic cells.

Each process requires a particular enzyme, an RNA polymerase and a transcription factor. We're going to write that, so let's say we take this. first promoter we want to read this gene this portion of the DNA and produce RNA and this is the RNA that we are actually going to synthesize right here, okay, from this gene, a particular enzyme, let's represent this in blue since we have been like In blue here there will be a particular enzyme that will read this DNA well and create this RNA. There is a particular enzyme, what is that enzyme called? It's called RNA polymerase, but this is the first promoter within eukaryotic cells that we're talking about.

Okay, so let's call it RNA polymerase. An RNA polymerase will read the DNA and produce a particular type of RNA, but in order for it to do this it needs a special protein that can bind to the promoter region which allows the RNA polymerase to bind to the DNA and read the DNA which is that particular protein. that protein let's represent it here in green, there is a particular protein that will bind here to the RNA polymerase and the promoter and will allow the RNA polymerase to bind to the DNA and start moving down reading the DNA and making this RNA.

What's it called? This is called transcription factor tf and there are many different types of transcription factors. What I need you to remember right now is that we call them transcription factors, which are used by RNA polymerases within eukaryotic cells, we call these general transcription factors. We'll talk about very specific types with a type 2 RNA polymerase a little later, but for now I need two things for this RNA polymerase to read. DNA and make this RNA RNA polymerase 1 needs a general transcription factor to bind to the promoter, allowing RNA polymerase 1 to then bind to the DNA, read it and produce RNA, what type of RNA does it produce?

I have all the RNAs inside the prokaryotic cells. from an RNA polymerase, but RNA polymerase 1 produces a very particular type of RNA and this is called rRNA. Now RNA is very important because it is incorporated into what are called ribosomes. Ribosomes and ribosomes are used in the translation process where we take the RNA and from there. produce proteins, so we'll talk about this later in another video, but for now the first thing I need you to know is RNA polymerase, one with transcription factors that reads DNA and makes RNA, that makes everything else pretty easy From this point, here is another promoter region. of a particular sequence of DNA within a eukaryotic cell, so this is the second promoter region, another enzyme binds to another RNA polymerase and not only that RNA polymerase here, but we also need a set of general transcription factors to bind to this promoter region, so general transcription factors that we need to bind to the promoter region, allowing this RNA polymerase to bind to the DNA, read it and then do what makes these particular types of RNA that we have here okay , this was the first promoter, this is the second, okay. so let's call this RNA polymerase 2.

RNA polymerase 2 will bind to this promoter through the transcription factor. Reads DNA and produces RNA. What type of RNA will it produce? The important thing I need you to remember is that it is producing m. rna mrna that you will see later again is the component. It will have to go through some very specific modifications that we will talk about in great detail and then eventually it will be translated with the help of rrna and another thing called trna into ribosomes and protein production, okay, the other thing that you guys can remember, if you want to be savants or ninja nerds, there is another RNA that is made here and we will talk about it a little later with what is called splicing and these are called small.

Nuclear RNAs and these are involved in what's called splicing and we'll get into that a little bit more in detail later, okay, but the great thing is that RNA polymerase II with the help of general transcription factors makes the mRNA and RNA polymerase are one with the help of the general RNA polymerase. Transcription factors produce RNA, when the heck do you think this last promoter region of this DNA sequence within this eukaryotic cell is going to produce RNA and is it the same process? What do I need here? I need general transcription factors to bind to the promoter region when it binds that facilitates or helps enable the type of RNA polymerase which three two bind to the DNA and then read the DNA and then synthesize which RNA which type of RNA is the type of RNA being synthesized from RNA polymerase III is primarily trna, but a little bit of snra is also made by type 3 RNA polymerase.

And if you guys really want to be more ninja nerdy, technically even a little bit of RNA is also made here, okay? trna, what the hell is this doing? You'll see later that this is also involved in the translation process, it carries a particular amino acid and an anticodon that will be involved in that process and we'll talk about that in a separate video, so I know that was a There's a lot of stuff to pull out and out of. this, but the big overarching theme that I really just took away from all of this what I want you to take away from this is this quick little thing here which is RNA polymerase 1 2 3 remember r m t RNA polymerase 1 gives way primarily to RNA polymerase 2 gives way mainly to RNA polymerase and then RNA polymerase 3 gives way mainly to RNA T these are the important things I want you to learn from all this if you want to go the extra mile, be extra ninja nerdy two and 3 can also give way to what small nuclear rna if you really want to go the extra mile, technically three can also give way to rna, but this is the basic thing to take away from what we just talked about and then the other The thing is that in prokaryotic cells we don't need all For these, we need an RNA polymerase holoenzyme to produce all RNAs.

One last thing is that you notice in eukaryotic cells that we have particular transcription factors that will be necessary for each RNA polymerase. The transcription factor in prokaryotes technically, if you want to be specific, is the sigma subunit because it is the portion that binds to the promoter to allow the core enzyme of RNA polymerase to read the DNA well, so it covers the basics of the two main ones. things that we need for this transcription process to happen now there's one other thing I want to talk about very quickly before we really start talking about the mRNA because that's going to be the main topic here that I want to have a quick little discussion about. how can we modulate the rate of transcription, either by speeding it up or by slowing it down, okay, the next thing I want to talk about is very, very briefly about the regulation of eukaryotic genes, so I want to have a quick little discussion about the regulation of genes, okay and The only reason I want to mention this is because it's very easy and it makes sense along with what we're talking about, but we're not going to talk about it in prokaryotic cells, we're going to talk mainly about this. genetic regulation and eukaryotic cells we are going to have a separate video because it is more complicated, we will talk about genetic regulation and prokaryotic cells with the lac operon and the tryptophan operon, we will get into that, but in eukaryotic cells there are two ways we can modulate and it is very easy one way we can modulate transcription is we have particular DNA sequences particular DNA sequences particularly palindromic sequences which are called enhancers and enhancers are basically DNA sequences and the most important thing I want you to learn from this is they can increase the rate. of transcription, therefore they increase the rate of transcription or the transcription process.For the fun of it, we'll say here's your RNA polymerase II, okay?

Here's your RNA polymerase two and it's reading the DNA. The DNA we already know has two strands. Let's call this strand top. This chain is also sometimes known as an antisense chain. This strand down here we'll call the coding strand now, when RNA polymerases read DNA, the strand they read is the template strand or the antisense strand, so that's the first thing I really need you to remember is that RNA polymerases, what chain they read, they read, we are going to put the template strand or it is also known as the antisense strand and that is the strand that they use to make the mRNA, they don't. we use the coding string, so we're going to put a little asterisk here to indicate that this is the string we're going to read now, when it reads it, it does it in a way that you, if you watch our DNA replication video, this should be very easy, let's say here that this end of the DNA is the leading end of the three, which means that this end is the leading five end and remember that a DNA strand on this side must have a complementary antiparallel strand on the other side, which means this is the three and here this has to be the five end on this side and this has to be the three ends on that side, what happens is that this RNA polymerase when it binds to the DNA does something very interesting: it binds to DNA through the initiation process and then opens DNA, who opened DNA before it was all that. in replication it was all that, like the replication complex, RNA polymerase does that, so the first thing we need to know is that RNA polymerase does what opens the DNA.

Now, in the replication, what else happened? You opened up DNA and you had those single strand binding proteins that kept it stable and kept it open, RNA polymerase does that on its own so it also stabilizes single stranded DNA molecules, so what was the enzyme in replication? that opened to unwind the DNA helicase? RNA polymerase has its intrinsic helicase activity so it also unwinds the DNA, after unwinding the DNA it starts reading the DNA, so let's say as you read the DNA in this three to five direction it will produce mRNA which will go in the opposite direction so you are going to read this 3 to direction 5 and as you do that you start synthesizing mrna correctly and this mrna will be synthesized in which direction what will be this starting point, the end of five and what would be this point of the three.

In the end, we know that the next thing that RNA polymerase does, whether in prokaryotic or eukaryotic cells, is it reads the DNA three to five and then it synthesizes the RNA in which direction, guys, five to three, very, very important, it last thing you guys do. I should ask, okay Zach, you also said that in replication DNA polymerases read the DNA and then if there was an accident or an error, they would check it and then cut the nucleotide. What about RNA polymerases? Do they do that too? It seems like they've done everything that was similar in DNA replication, that's the only thing that's controversial, so the only thing that's relatively controversial is whether there's a proof-reading function that we don't really know about, it's still subject to study. , so that's one thing.

To remember, if you want to compare this, the review function is a little bit uncertain at the moment, okay, so we have an idea now that we've read this DNA and created the RNA. I know we talk about this a lot in DNA replication. We're talking about that here and sometimes it can be really confusing when you say five and three and no, I don't understand what you're talking about, Zach, so I want to take a quick second and explain what the hell I mean when I say read it three to three. five and synthesizes it from five to three.

A diagram that I really think will clarify this for you. Let's take a second to understand what I mean by reading DNA from three to five and then synthesizing it from five to three. I think it's very important to understand that, so let's say here we have this DNA strand, so this will be our DNA template, if you agree, this is our DNA template. on this side the blue and then this is going to be the RNA that we are going to synthesize using RNA polymerase type 2 and the eukaryotes are the hollow enzyme RNA polymerase and the prokaryotes now when we are making this RNA we have to read the DNA in which direction the three ends with the five and what is the three and you remember the video about the structure of DNA is the oh so this will be the end three what is the end five is the phosphate group so the phosphate group is is going to be the five, so I have to read this starting at the o h portion towards the five end where the phosphate is, so RNA polymerase let's assume I'm RNA polymerase.

I'm walking to the right to do it. I find the three primes and I'm like, oh, okay, I'm going to go up, oh, I found the top three, the top five, let me fill this in, oh, I feel my nitrogenous base, the nitrogenous base that feels is adenine, so the collects in her small purse. of nucleotides okay, this is adenine, the complementary base is thymine uh oh no, that's not correct because you know that if we are taking DNA to make RNA, what is the only nucleotide that changes from the DNA and the RNA? Adenine is no longer complementary to thymine. in RNA it's uracil, so DNA, RNA polymerase, will come, read, find the three ends, read the nucleotide and say, oop, okay, this is an adenine, reach into your bag of a bunch of nucleotides and takes out the uracil when it takes it out. puts the nucleotide in a particular orientation what is the orientation we said reads it from three to five and synthesizes it from five to three what is the five end here is the nucleotide the five end is this phosphate group the three end is this group oh like this which is going to flip the nucleotide in the opposite direction and make sure that the nitrogenous base here is what is uracil, then when it does that, it's going to go to the next one, so it's going to continue, it's going to go to the next point, this is where the next one is. oh group, you'd be right, the three reads of the leading end, find, find the nucleotide, it says, oh, the nitrogenous base here won't let me look in my wallet for a bunch of different uh, good old nucleotides, I'm going to read it. a, I'm going to put my nucleotide and I'm going to rotate it where it has the five leading end facing down, three leading ends pointing up and then the nitrogenous base that is complementary to the t is a, when it does that, then it fuses the three prime ends and the five prime ends together form a bond.

What is that bond called a phosphodiester bond? and the same process happens, so it will do what we are going to fix these three primes there and then it will go to the next nucleotide here. the main three end where the group oh is read finds the nucleotide he says it is a g he puts his hand in his wallet he takes out a nucleotide with the cytosine when he does it he turns it to where the five end is on this side there is my phosphate the three the head end is facing up and it says oh, the nucleotide that goes with this is with the nitrogenous base c, then it says oh, I have my phi prime n situated near the three head ends of the nucleotide above, let me merge these two together and make my phosphate ester bond and just for the fun of it because repeating the repetition I guess is useful come on read this says well the next one here is my three prime end where the group reads oh find the nitrogenous base is a cytosine dig into its wallet and takes out the nucleotide guanosine sorry, the nitrogenous base of guanine then when you do that, you place it where the end five primes is located down, three primes n is located up in this case and then the nitrogenous bases in guanine so it says oh my five cousins ​​n, I can sew We join it to the three cousins ​​end of the previous nucleotide and we form my phosphodiester bond and that's how we make the RNA read it from three to five and synthesize it from five to three.

Damn, we're good now that we've done that, the last thing I need. What you need to understand is that RNA polymerase is a very important enzyme within eukaryotic and prokaryotic cells. A question that may arise and it is very silly and annoying, but you should know that in eukaryotic cells we can inhibit RNA polymerase using a kind of Amanitin toxin is for fungi and this can inhibit the internal RNA polymerase. We will put eukaryotic cells. Okay, there's another drug they also love to ask for at exams, called rifampicin. It is an antibiotic and this inhibits the internal RNA polymerase.

Yes it is an antibiotic. Good against bacteria, prokaryotic cells, so this will inhibit the RNA polymerase inside the prokaryotic cells, which would inhibit the initiation part, the elongation, basically making RNA. If you can't make RNA, you can't make proteins. If you can't produce protein. it can't perform the general functions of the cell, so this comes from a poisonous fungus, which is stupid to know, but they like to ask about it in their exams and then rifampicin is an antibiotic that they also love to ask about, okay? We've talked about lengthening. We created damn RNA. RNA polymerase is working very hard.

The last thing we have to do is finish it. We don't need more RNA. We have created the RNA we need. the protein called termination okay, so we talk about elongation, the next step, the last step that we really have to discuss here is termination, we have to finish this whole transcription process, so the last step is termination, Now, unfortunately, termination is probably one of them. Unfortunately, one of the most annoying and complicated and it's different in prokaryotes and eukaryotes, so it's a little frustrating, but termination is basically when we've already done our RNA transcription and we just need to separate or dissociate it. away from the DNA and prevent RNA polymerase from reading more DNA and making more RNA, so just stop transcription.

How do we do that? In prokaryotes there are two mechanisms. One of the ways this happens is through what is called road dependent termination. one is through this process called row-dependent termination and it's really simple, believe it or not, so let's say here we take the prokaryotes, we choose blue for our RNA polymerase, so the RNA polymerase here's our RNA polymerase, it's reading this DNA while reading the DNA. again, what are you doing from it? Remember it's making the RNA, in this case it could be any RNA, it could be the mRNA, the mRNA, whatever does this, there's a protein called rho and what rho does is this rho protein will start to move. the mRNA goes up and as the RNA that's being synthesized by the RNA polymerase goes up when it gets to this RNA polymerase, it basically says, hey, just take the RNA polymerase out of the DNA, if you take the RNA polymerase out of the DNA, is it going to be able to keep reproducing the DNA and make more RNA, no, so that ends the transcription process, so the most important thing I need you to know here is that with pathway-dependent termination, the rho protein makes the RNA polymerase separate. dissociate, if you want, okay, separate from the DNA, okay, beautiful, the next mechanism within prokaryotes is rho-independent termination, so we don't use the row protein, so we call this rho-independent termination. row, now with this process it is a little more complicated. and a little annoying, let's say here we have the correct DNA and inside the DNA we are going to mark them here, we are going to say this is our correct template strand, then this strand is the correct template strand or the antisense strand and then This will be our strand coding, so which one does RNA polymerase read?

Reads the template string or the antisense string. There is something similar in particular called inverted repeats that form within the DNA that is read by RNA polymerase. So what happens is this RNA. The polymerase will bind to that template strand and start reading it creating the RNA when it starts producing this RNA. If you find a particular sequence of DNA called inverted repeats, let's write these inverted repeats in a kind of nice little color. let's make orange and say here we have an inverted repeat where we have c c g g and then a bunch of nucleotides that we don't care about and then here we will have ggcc, okay, so we will just have this is the template.

Again, in the coding strain, it would just be the complementary base, so if this was cc, this would be gg cc, we don't really care about these nucleotides, cc gg, right, the RNA polymerase will read this template strand, what happens is correct. We're going to get this kind of strand here where you're going to have a bunch of nucleotides already formed here and then you get to this kind of inverted repeat area and what happens is it reads this and then basically everything you read inside the template strand should be the same as in the coding strand because it's the complementary base, so you're going to have g g c c which will produce a bunch of nucleotides that we don't care about and then c c g g what happens is when this RNA is kind. from coming and being transcribed from the RNA polymerase, something interesting happens where some of these Cs and some of these Gs in this portion have a strong affinity for some of the Cs and some of the Gs in this portion of the RNA and as they begin to having this affinity they begin to get closer and wantinteract with each other through these hydrogen bonds, so it creates this really interesting kind of hairpin loop, if you will, where there are a bunch of g's and c's inside this kind of hairpin loop that are friendly. of interacting with each other and what happens is that that hairpin loop is what causes the RNA polymerase to jump off the DNA and finish the transcription process because what happens is that once this hairpin loop is formed, what will happen is that there will be particular changes. enzymes that will bind to that portion and cleave the RNA from the RNA polymerase, so the most important thing I need you to know within the independent termination of the row is that you will reach this area where the RNA polymerase will transcribe the DNA reading forming RNA .

You will hit these areas of inverted reps when these inverted reps are performed, they will create something called a hairpin loop. This hairpin loop will activate particular cleavage enzymes to come and cleave a couple of nucleotides after that hairpin loop to separate it from the RNA polymerase and then here you have the RNA that you formed, so that's one of the ways that we have the independent termination pathway through prokaryotes. The last termination mechanism will be eukaryotic cells. Now how does this work? This one is actually relatively simple, so we had RNA polymerase in eukaryotes and this was orange, okay, it binds to DNA, it reads the DNA as it reads the DNA, it produces RNA when it starts to produce this RNA, it comes to a sequence particular where when you start reading DNA and produce RNA, you form a particular sequence of a u a a a okay, so what is the nucleotide sequence here, let's write it down? this type of nucleotide sequence occurs, so now we know what that nucleotide sequence is, let's put it this way, here is that nucleotide sequence, that polyadenylation signal that has been synthesized or formed by RNA polymerase with eukaryotes, once That happens, it is activated. particular enzymes and those enzymes will come to the area here and cleave the RNA from the RNA polymerase separating this RNA from the DNA and the RNA polymerase and then again, what will I have in this portion here as sort of a schematic portion? here this will be my polyadenylation signal.

This is important because we're going to talk about post-transcriptional modification in a second, so I know this was a bunch of crap, but I'll summarize very quickly because this is one of the most difficult parts of transcription: termination. prokaryotes there are two forms of independent pathway dependent row with this one you need a row protein to eliminate RNA polymerase if you don't have it, it can't make more RNA the other is row independent, you don't have row The protein, RNA polymerase, read the DNA that makes RNA and hits these areas of inverted repeats. These inverted repeats when made within the RNA create a hydrogen bond interaction between them that causes a hairpin loop to form that signals particular enzymes. to separate the RNA from the RNA polymerase and we have made our RNA there, the last one is in eukaryotes, the RNA polymerase is reading the DNA and it reaches a particular sequence of nucleotides where it reads and then produces a polyadenylation signal that activates the enzymes so that cleave the RNA from RNA polymerase, ending the transcription process that really impacts this home.

Now let's talk about post-transcriptional modification. At this point we know how to take the DNA. Do RNA well. We talk about the different types of RNA. using RNA polymerases using transcription factors we talked a little bit about gene regulation, we even went through all the stages of transcription taking the DNA and producing the mRNA to the point where we finally created the mRNA and separated it from the Unfortunately, DNA is not all for transcription. Now we have this correct mRNA, so basically, what have we covered up to this point? We take the DNA that we read, let's say here in this part, I'll just put here is our promoter, our RNA polymerase has read.

In this gene sequence we reach a termination sequence, let's say here is our termination sequence that we talked about here and once we reach that termination sequence, the RNA polymerase will fall off and then from this the RNA, so these were pretty much the basics. of the transcription, but now we have to modify this, here is what is actually a misnomer to say this is mrna, technically it is not mrna right now, so this piece of RNA that we made is okay and this is this process post-transcription. modification this only happens it's very important let me write this this only happens in eukaryotic cells so it's good all this stuff we're going to talk about here is only in eukaryotic cells it doesn't happen in prokaryotic cells so just make your RNA and that's it, technically this immature mRNA, so to speak, we will give it a very specific name, we call it heterogeneous nuclear RNA.

Now this heterogeneous nuclear RNA is a kind of immature mRNA that has to go through some modifications to actually mature. mRNA that can then be translated to produce proteins. What are these modifications? The first thing we need to do is put something on one of these ends, so now we know a little bit about the terminology of the ends of this immature. mrna or hn rna on this end let's call it the top five end what's on that top five end? Remember phosphate groups? What's on this end? something very interesting is at the five prime end of this heterogeneous nuclear RNA or the HRNA, you have a triphosphate that we are representing here with these orange circles, an enzyme comes to the rescue and cleaves one of those phosphate molecules.

What is that enzyme called? It's this cute little orange enzyme. This orange enzyme is called RNA triphosphatase and what it does is it comes and cleaves off the portion that it cleaves off of one of these phosphate groups. It's going to cleave one of the phosphate groups. so now I only have two phosphates on the end of this five prime end, then another enzyme comes in and says, "hey, there's only two phosphates here." Now I can add something here and I'm going to add what's called a gmp molecule. I'm going to add again, I'm going to add a gmp molecule which is guanosine monophosphate, so we're going to represent that here, what we add to the phosphate for guanosine monophosphate and then we're just going to represent this as guanosine, so this is our guanosine and that blue circle is the phosphate in guanosine, so what does it add technically?

You add to these little two phosphates, right, you add gtp, but when you do that, two phosphates are released into the form. of pyrophosphate which is then broken down into individual phosphates by pyrophosphatase, so if I took gtp and removed two phosphates, what am I left with gmp, so add this gmp group to that two phosphate end at the top five end, so this enzyme that adds that gmp in the form of gtp is called guanolyl uh transferase guanalyl transferase beautiful, so this last enzyme here, which is involved in this step here in the first five n, will add a methyl group to one of the components of the guanosine monophosphate. it's actually like one of the seventh components in that structure that adds a methyl group and at the end of this this enzyme that adds a methyl group to what you think is called methyltransferase at the end of this process where you took the raw at the end, which it had three phosphates, one was removed, the guanola transfers were added to the gmp, the methyl transferase was added and methyl group was added to that.

You formed this complex here and we call this whole complex that we just added in a group of seven methyl guanosine, okay and that's it. at that end of the top five, this is called limiting, this is called limiting, so everything we just did at this end of the top five is called limiting, what the hell are we doing all this for the purpose of limiting is to help to start the translation? this sequence this kind of five prime end with that seven methyl guanosine or that five prime constraint, if you will, is a kind of signal sequence that allows it to interact with the ribosome and undergo translation, the other thing it does is prevent degradation by nuclease enzymes that want to come and break down the RNA, so it helps prevent degradation, it helps start the translation process.

One more thing that is silly to know, but they love to ask, is that there is a particular molecule that this methyltransferase uses to add that methyl group and sometimes it is very important to know and this is called adenosyl methionine, also known as sam sam, it carries a methyl group, it's like a methyl donor, so to speak, it gives that methyl group to the methyl transferase. and the methyl transferase adds that methyl group to the guanosine monophosphate forming the 7-methylguanosine or that 5-prime cap, okay, that's the first thing that happens. Now we have to talk about the 3 prime end in the three prime end that we had. oh group, right, that's the ohn, but do you remember that in eukaryotes there was a particular signal that prevented that finished transcript from being generated?

What was that nucleotide signal? Do you remember? Test your knowledge guys, a a u triple a, that was that polyadenylation signal. Do you remember? that the polyadenylation segment that we talk about in eukaryotes that polyadenylation signal is recognizable by this cute little purple enzyme here this cute little purple enzyme is called poly polymerase it's called poly polymerase what it does is that on this side it has a lot of adenine nucleotides So you can eat a lot of nucleotides that contain the adenine nitrogenous base, you take one end and identify that polyadenylation signal, you take the other end and add all those adenine nucleotides, a bunch of them, sometimes up to 200 adenine nucleotides when he does this. it forms a tail at that three prime end with a bunch of adenine nucleotides and we call this a poly a tail so the poly a tail what is the purpose of this is exactly the same thing it helps start the translation process and it helps to slow down the degradation so kind of nuclease enzymes that will try to come and break down that end, okay, the other thing they do is help transport this hnrna, eventually they will help transport the hnrna that will become mrna out of the nucleus and into the cytosol. . so they also play a small role in transport out of the nucleus and into the cytosol.

Well, in this first step, what did we do? We made five main stops, we went over that part and the three main polys, a tail we made. Okay, now we have this, so after having done all this massive mess, we have reached this point. Well, in this part, what do we have? Let's just write this. We'll surround him here. These are our top five. cover with the 7-methylguanosine and at this end we have already formed our polyethylene glue, the next thing that happens is what is called splicing and this can sometimes be a little annoying, but it is not that bad, I promise, let's say here that this is the nucleotide sequence within this rna well, we are not in the rna we are still in this h in the rna we are still in this h in the rna at this point we have not done mrna yet within this hn rna there are particular nucleotides that will read translated and will in fact encode particular amino acids.

There are other nucleotides within this shRNA that will not be read and do not code for a particular amino acid. We give them those very specific names. I will highlight them in different colors. So let's say I highlight this one here and pink and then I'll highlight this one here in this kind of maroon color and then I'll choose a blue here and then we'll do another maroon color and then we'll do one more. color after this, here's another maroon and then we'll do it just for the fun of it, black, okay, these portions here, the pink, it's actually going to code for an amino acid, if it codes for an amino acid, we give it a very specific for that. and we call them exons, so exons code for an amino acid.

Well, particularly amino acids will make proteins. These other portions will be like this or we will mention which ones are exons and which ones are the next thing called. introns Introns are basically sequences of nucleotides that do not code for amino acids that will help make proteins very important. I'm going to call this pink portion of the h and RNA. I'm going to call it exon, but we have a bunch of them in this h and rna so I'm going to call this exon one, okay, that's going to code for some amino acids. I'm going to have this portion here that will be in the brown.

I'm going to call this an intron, but you can have multiple introns, so I'm going to call it intron 1. Same thing here, this is going to be coding, so if it's coding, that's what an exon is, well, we have multiple types, so let's go to call this exon. 2. then I am going to test them again, this one does not encode amino acids, so it will be an intron, but we already have the intron, once we will call it intron 2 and you already understand the The pattern that I am going to use here, this one encodes, for which will be one exon and we already have 1 2, so it will be three.

Well, let's do something called splicing, where we think about this if the introns don't encode any amino acids, we don't even need them, no, let's get rid of them, that's all splicing does, is get rid of these introns or also known as intermediate sequences and then join the exonswithin cardiac and skeletal muscle, okay engineers, I promise, I'm really sorry this is so long, but there's one last thing I want us to talk about. The last thing I want us to talk about is called RNA editing this. It's also mentioned a lot in your exams and the reason is because it's really interesting how this happens.

There are two different types of RNA editing. I just want to mention one of them because it is the most relevant to your usmles and in a way. in a clinical setting, so let's say here we have our correct mRNA, so this is an hn RNA. We already have at this moment for RNA editing. We have already formed our functional mRNA, so at this point this structure here is an mRNA. Well, this mRNA may have a particular nucleotide sequence that a special enzyme can read and sometimes change the nucleotides with what is that nucleotide sequence that you can see in this mRNA.

What we really want to know is c to a, let's talk about apoproteins, that's why I mention caa, so this is our signal, which is actually very important within this mRNA that will produce the april proteins, a particular protein called let's. Let's say this mRNA will code for a particular protein called apo b100. If you watch our video on lipoprotein metabolism, this will look familiar, but April B100, this will be Mr. enable that will code for that protein and here is a particular nucleotide. The sequence that we are going to modify in the hepatocytes, this nucleotide sequence is not altered in any way, it remains the same, so it will not be changed, it will still be c a a and as long as this mRNA is translated by the ribosomes, it will produce a particular protein. what we already talked about is called apob 100, but in enterocytes, okay, your gi cells, what are these cells called here?

They are called enterocytes. They have a very special enzyme where they can modify the same gene that produces april b100, but they produce a different protein. Damn, how do they do it? Let me explain to you that there is a nice little blue enzyme in enterocytes called cytidine d-aminase and what cytidine deaminase does is it deaminates the cytodine right here or the cytosine nitrogenous base and changes it to uracil, so now change it here where we go to have this like change c and put u a a if you know anything about your codons, there is a little trick to remember your stop codons, remember the little way to remember them, remember as you leave. you're far away, you're gone, these are the easy ways to remember your stop codons.

Do any of these look like stop codons? Yeah, uaa ua, that's a stop codon. So what will happen is when you have the ribosomes that will read this, let's say. here I put it as a little ribosome that will read this and produce a particular protein as it gets to this point where it will translate it, which is a stop codon, then it will read the rest of the RNA and translate it into a long protein no, like this that at this point the translation will stop, you will not read the rest of the mRNA and you will create the full protein, instead you will create a smaller protein and this small protein is called apo b-48, this is something that they love. ask in your exams because they're taking the same mRNA just modifying it a little bit to make a different protein that's a completely different size protein, so that's really cool, I definitely wanted you guys to know that and that wraps up our lecture on transcription. of DNA.

Alright nurse ninja, so in this video we talk a lot about DNA transcription. I hope that made sense and I hope you enjoyed it as always, ninja nerds, until next time.