Regulation and Transcription Factors PDF

Good morning. Good afternoon. Good evening, or good night. This is part of the asynchronous material where we're going to finish up regulation and get into our next topic so that on Monday we can do some nice review of what we've done and move forward. I wanted to start with this particular slide be...

Good morning. Good afternoon. Good evening, or good night. This is part of the asynchronous material where we're going to finish up regulation and get into our next topic so that on Monday we can do some nice review of what we've done and move forward. I wanted to start with this particular slide because the image that is present here has a lot of very specific examples that actually correlate with human conditions. And so I just wanted to give you a little heads up on we're going to cover some of these, not all of them, but I just wanted you to see that the wide range of steps in this do in fact correlate with known human conditions. And so, for example, dysfunction in processing can correlate with dystrophy, mRNA export has correlates of each of these, including the ability to break down or turn over proteins, all of them, every step and dysregulation at any step is correlated with human disease. And so with this figure, I just wanted you to see that we've actually talked about a lot of this. So this is just a visualization of the different things we've already discussed. One of the things that was pointed out to me recently is that I'm not so great as telling you what figures you really need to know every step of versus those that don't matter as much. I'm going to try to do that a little bit more consistently. So for this example, this was just that concept of the function of a transcription factor where we have the transcription factor that leads to transcription. And so this is really an explanation of how this diagram over here works. You will have the factor in a node and then a line or an edge that indicates how that factor relates to the next molecule. And so in terms of if we've got a circle, that's going to be our factor or our molecule and all the lines connecting or direct paths of regulation. And so this figure here is just showing you the substrates, processes and disorders. It's just a nice circular impact statement for the idea of this RNA modification, specifically the pseudo-uridylation of RNA. Remember, the other example that we had was the base deamination of RNA as part of RNA processing and DNA damage. DNA damage. So if we move ahead to where we actually were, we are sort of on the attenuation piece. And so just a reminder with regards to the operon, the big takeaways are when are we on? big takeaways are when are we on? When are we off? So if we are in a situation where we are about breaking down something else, we are about utilizing another molecule, then it doesn't make sense to exist or be on when that molecule is not present. So in the case of the lac operon, we are not going to turn on the lac operon unless lactose is present to be used because the lac operon is all about the molecules necessary to break down lactose. And since lactose is not a preferred fuel, it is a fuel that we can only use under certain conditions, we are going to have situations where we want to keep it off even in the presence of lactose because there is a preferred fuel and there is a better plan for that. And so with glucose, we actually tie that to things like cyclic AMP. And I bring these up because I know you've seen them in physiology, but they'll also come back in biochem. So ultimately is when would we be on and why would we be on? So with TRIP, we have the opposite. Tryptophan is creating the molecules to create tryptophan. And so in this case, we don't want this to be turned off unless we have sufficient levels of tryptophan. And so the presence of tryptophan is going to actually bind a repressor and lead to repression of the operon. And so this one is on until you've made it and have a sufficient volume. This is a negative feedback regulation. And so we get into why we talk about the TRIP operon is because it has that second step, that translational regulation that has a lot to do with the shape and structure of the mRNA as it's being utilized. We call this attenuation. By changing the binding and physical structure of the mRNA molecule, you actually change the ability to translate the material. And so in terms of confirmations, one type of stem loop for the TRIP will shut it off and others will allow it to be translated. So keeping that in mind, this picture right here is sort of showing you that difference between when we have transcription enabling translation and when it's not. So remember, those are things that are coupled. So if we are in a prokaryote, those two things are in the same compartment, transcription and translation are in the same compartment, and so they can influence each other. And the shape of our transcript can influence whether translation can take place. And so this figure here on the left is not one that I am expecting you to fully engage with other than the big takeaway that not only does the mRNA itself have a role in this, the tryptophan have a role in this, but also tRNAs charged with tryptophan. Because we do not need high levels of transcription and translation when we have plenty of tryptophan to go around. That's specific to shutting off the TRIP operon. And so this takes us to where we really left off yesterday, which is that translational regulation. Changing the shape you take can change accessibility to the ribosome, just like changing the DNA structure could affect polymerase accessibility. We also have the reality of complementary base pairing from small interfering RNAs, SiRNAs, as being possible of inhibiting this process. And so antisense RNA, any sort of RNA that is complementary to a sequence can block that sequence. And in some ways, this can target it to being literally chopped up into pieces, literally digested. Or it can form a double stranded molecule or it can form confirmations to the mRNA that it prevents it from being associated with the ribosome and therefore being translated. And this is a really important concept if we think about it from the standpoint of accessibility equals possibility of an action taking place. Accessibility to the RNA polymerase enabled transcription. Accessibility of the ribosome enables translation. And having the ability to have changeable 3D confirmations gives us an extra level of control. So if we can think about this from the perspective of we turned off transcription and we wanted to produce X number of transcripts. And so we created those transcripts, but it turns out that we brought a bunch of stuff in from diet, for example, and we no longer need the same level of translation because we have now sort of an environmental provider. That gives us the ability to regulate translation, gives us the ability to have that read of the whole system, if you will. We can have that negative feedback to turn things off at the transcription level, but also control it so we don't have toxic levels by preventing translation from taking place. So we sort of link all these things together to ensure an overall cellular health. In eukaryotes, we actually do regulate translation. We regulate both the timing and the location. By preventing mRNA molecules from reaching the right compartment, we can prevent translation. By having that signal sequence and targeting material to the rough ER, we can actually control where that material is translated into. Any time we are doing this sort of regulation, we want to link it to a real cause and consequence, right? In terms of development, mRNA localization, availability of translation, localization of translation is actually really important to even the most foundational developmental concepts like spatial patterning, front versus back, left versus right, segmental patterning. That is absolutely essentially controlled at the level of translation after we've laid down the mRNA transcripts. Disregulating anything in relation to processing translation to any dysregulation is linked to human disease. Some of the areas that are of particular importance include tRNA dysfunction, myopathies of ribosomes, our ability to handle cellular environmental stresses. Stress responses can be dysregulated when translation is dysregulated. Of course, we also have to take into consideration the mTOR pathway, which we will talk about again. Any time that we see sort of tissue specificity to a disease, we can link that to transcription, but we can also link it more specifically depending on the change to a translational rate or a functional activity. So as we have the whole gambit of molecular genetics, any of those steps being dysregulated could be a direct cause. And especially an example of a ribosomopathy, we have a direct link to translation as well, functional outcomes. And so if we look at this figure, it's just showing you things like high concentrations of PTIN activity being associated with normal development versus low levels being associated with cancer. PI3 kinase activity, low levels being associated with immunodeficiencies, whereas high levels are associated with lymphomas. So it's a careful balance where we want the right amount, not too much, not too little. And if normal lies somewhere in the middle, then we can imagine that there might be a companion opposite for having too much as there is for having too little. So just keeping in mind that there is a direct consequence for protein expression and disorders. And so that's what these top figures are showing you. They're showing you systems where if you have high expression versus low and what are some of the consequences for that. So hopefully this gives you a nice tie into translation being directly correlated to human disease. And one of the key things that we want to talk about with that are things like our iron responsive elements. Regulation of RNA binding proteins to non-coding elements is a key aspect of controlling translation. So if we look at this first situation, we have a normal situation where we have an iron response element and translation. In the absence of iron, an IRP or an iron regulatory protein can bind to the response element and block translation. Once we have iron, that iron responsive protein cannot associate and we can get translation again. There is also the potential for the opposite to be true. Where in the absence of iron, we do not bind or I'm sorry, in the absence of iron, our iron responsive proteins can bind to the IRE and allow translation. And so it's really showing you the differential effects of an iron responsive element. This set of iron responsive elements not being bound means that the material gets targeted for degradation. It's not stable enough to actually be translated. Once the iron responsive proteins bind, it can be. But once iron levels increase, high concentrations of iron keep this iron responsive protein from being able to bond and that results in that degradation. So completely different effects from the same concept. And so this particular example here on the left really focuses on A, B and C and that allows. Sorry, it says E here and it should say C and I'm so sorry, high concentrations of iron binding. This entire example here is A, B and C. So I will correct that on the slide. So right here, this should say instead of E, this should say C. I apologize for that error. I should say C. So it's really showing you this example at the top. I apologize. Type of my finger went up instead of down when I was typing. I'm so sorry, I should have caught that anyway. So this is a differential effect of regulating translation using iron responsive proteins, iron responsive elements. There are also CAP independent mechanisms of regulating translation. So remember that when it comes to your carrier translation, we've got the CAP and that's how our ribosome associates. This is all about a CAP independent mechanism and it's really a stress response. And what we have are internal ribosome entry sites, IRES. And these are very much about selective cellular synthesis of proteins. Most of these genes that have these elements are going to be associated with stress, mitosis, apoptosis. And ultimately we get these key factors binding to the IRES and enabling translation, increasing it typically in the response to stress. And so here you have a normal situation, normal oxygen levels. And so we have our translational unit associating at that AUG. We're translating while we're all good. In the event of hypoxia, we can bypass the need for this spot and be internally activating translation. It's really interesting because this was actually first identified in poliovirus. And essentially what this is, is another set of sequences that directly recruit translational machinery. And so we have now eliminated the need for the same regulation because we are under a situation of extreme need. And it sort of enables this nice little switch between when everything's fine and an indicator that everything is not fine. It's a really cool concept. Remember that any time that you make a change to one of these processes, you have three outcomes. It could do nothing. It could leave everything exactly the same. Or it could increase activity or it could decrease activity. And the consequence of those three outcomes is different depending on what we're talking about. If we are talking about a loss of function that is different than a gain of function depending on the activity or role of the molecule that lost or gained function. Lack of binding something at an operator for an operon model equals no repression. A loss of function to a repressor means you can't shut off that operon. Lack of binding at a promoter, however, means you can't turn it on. Gain of function for an operator would mean that you would have excessive or constitutive repression. So you have to look at what is this molecule doing? What is the change that happened to the molecule? And therefore, what is the consequence of that molecule's change? So if we lose lactose, right, there's no lactose available. That's going to mean little or minimal activity of the lack operon. Do we have a requirement for iron to be present in order to be active, but there's no iron? That's going to be different than if iron is present. I really want to understand that we have to pay attention to those two things. What is the molecule I'm working with? What is its job? How is it regulated? And then if we dysregulate its activity or we dysregulate its control, did we make it go be more active? Did we make it be less active? And those will have different consequences. And so here is where we will pick up on Monday. I hope you have a lovely day and thank you so much for your time.