Gene Regulation
Look over the last slide for the Lecture!! Very helpful
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How much CAP and cAMP are present when glucose and lactose are present?
CAP = catabolite active protein. When lactose is present and gulcose is scare, the level of cAMP is high: lots of lac mRNA is synthesized When both lactose and glucose are present, the level of cAMP is low: little lac mRNA synthesized.
Eukaryotic Gene Expression
Chromatin Structure, transcriptional Regulation, Post-Transcriptional Regualtion (RNA processing, mRNA stability, Initiation of translation, protein processing, protein degradation). DNA is in nucleus, can have chromatin modification (heterochromatic [very condensed chromatic] to euchromatin [can now have it bind to RNA polymerase] for example), Then transcription [turn on or not], make primary transcript [pre-RNA molecule with exons and introns], Turn it into mature RNA molecule [the tail, cap, splicing etc.], then transport to cytoplasm from the nucleous. In the cytopalsm you can initiate translation. When ribosome binds and starts to make the protein. Eukaryotic cells may or may not initiate translation, they could degrade the mRNA or make the polypeptide from the mRNA. Then you have protein processing, you might degrade the protein or make an active protein. Then there is the transport to cellular destination for cellular function. Shows how precisely how proteins can be regulated.
Eukaryotes Gene Expression Summary:
Eukaryotic genes are NOT organized in operons, Eukaryotes use a variety of post-transcriptional mechanisms to fine-tune gene expression. The 5' cap and 3' poly-A tail protect mRNA from degradation, The longer the mRNA lasts in the cytoplasm, the more protein molecules it can make.
Differential Gene Expression in Multicellular Organisms
Every cell has the same genes (DNA), cells express different subset of genes, using many different types of regulation helps eukaryotes fine-tune gene regulation (Not gametes or red-blood cells)
Initiation of transcription
General Transcription Factors: Bind promoters. Specific Transcription Factors: Bind enhancers and silencers far upstream or downstream of the transcription start site. Enhancers: Increase the RATE of transcription Silencers: Decrease the RATE of transcription Need everything to come together in the correct order for RNA polymerase to bind
Operons:
Genes that work together, live together. Genes are next to each other in the DNA, one mRNA molecule makes several proteins, genes turned ON or OFF at the same time.
Chromatin Structure
Heteronchromatin NOT expressed-so tight that RNA polymerase can't get in, other protein cannot get it Euchromatic: CAN BE expressed, RNA polymerase could bind. Protein modifications can change chromatic structure Histone tails-they can be modified-acytl groups added to them, phosphate groups added etc. Acetylated histones: Actylation of histone tails promotes loose chomatic structure that permits transcription. Unacetylated histones, this is more like heteronchromatin, this is all dependent on what can bind to these histone tails.
E. coli trp operon is repressible
Promoter, region in DNA where the RNA polymerase binds. Operator: sequence of DNA. Genes of operon (used to make the various enzymes for tryptophan synthesis), trpA. So promotor can bind, it can make the mRNA into the proteins (start codon and stop condon, right next to each other). Proteins fold, interact with each other to make tryptophan. All the information is very condensed Trp Repressor: Also has a promoter. It constantly makes this at the same time, always there. It used the trpR + promoter, mRNA, it makes a protein that is an inactive repressor. Tryptophan is a corepressor because it binds to the inactive repressor, making it active. When there is no tryptophan the repressor in inactive, the operon is on, and there is a constant, low level of transcription occurring.
Operon Regulation
Repressible: ON unless turned off, tend to be anabolic pathways (synthesize essential molecules, only stop when enough product is available) Ex Trp operon (Tryptophan) Inducible: OFF unless turned on. Tend to be catabolic pathway (break down nutrients, don't need enzymes when nutrient not present). Example: Lac operon (Lacotse) Cells only make when you drink milk, you don't always drink milk.
Tryptophan is a corepressor
The extra tryptophan that is hanging around after your big turkey dinner. It is called a corepressor because it binds to the repressor protein. It goes from the inactive repressor form to an active repressor. There has to be extra around to do this. It then binds to the operator region, the operator is blocked by the repressor so the RNA polymerase cannot bind to make the mRNA, so more tryptophan will not be produced. (Tryptophan present, repressor active, operon off).
Summary of lac operon regulation
The lac repressor (turns transcription ON/OFF, NO lactose operon is off, lactose present--->operon is on, CAP & cAMP: Control the RATE of transcription. (When glucose and lactose are both present, the operon works slowly, when there is no glucose the operon works quickly
General Rule for Bacterial Operons
These are examples of Negative Feedback Repressible Operons ---> Pathways (synthesize essential molecules, only stop when enough product is available from environment or from metabolic activity). Inducible Operons ----> Catabolic pathways (break down nutrients, don't need enzymes when nutrient not present).
Tissue-and Gene-Specific Regulation
To promoters, to different enhancer regions. What to be able to express these in their specific tissues. Different gene sequences for the liver and the eye. There are specific available activators to bind to the specific transcription factors to initiate transcription. Decides which genes get turned on. Albumin is a gene made in the liver, binds lots of things in blood makes things circulating. Sweep of specific transcription factors. Crystallin: Expressed in the eyes
Prokaryotes
Transcription initiation, operons. They have rapid response by turning transcription on (change to environmental conditions rapidly).
Gene and Enzyme Regulation (metabolic pathway)
Tryptophan is an amino acid, something the cells are really going to need, you can't make the proteins requiring tryptophan if you don't have it. When there is plenty, you don't need to make more of it. Example: When you have been fasting for thanksgiving dinner your body has been making lots of tryptophan to compensate. After turkey dinner, environment has lots of tryptophan. It would not make sense to use all this energy to make typotophan when it is readily in the environment. Stops making it, there is a negative feedback for the first enzyme. The precursor cannot do anything. Presence of typtophan can bind to the first enzyme and block it. So first disable the enzymes and then stop making them. The tryptophan can also turn off the mRNA that make this particular gene to create the enzyme (regulation of enzyme production). trpB and trpA gene make enzyme 3: Genes are arranged in operons, they are arranged right next to each other. In eukaryotes several genes that are related to a metabolic pathway can be on different chromosomes, scattered everywhere. Prokaryotes can turn on transcription once and make all the proteins they need, and then when they don't need them they can turn them all off. mRNA can make several proteins if they want to, not just one.
Bacteria also use Positive feedback to regulate gene expression
When E. coli has both glucose and lactose in the environment, it uses the glucose and NOT lactose. The cells sense the presence of glucose in the environment via cAMP
The system with cAMP and CAP
When lactose is present and gucose is scarce: cAMP binds to CAP to positively regulate the lac operon. Then the activated CAP binds next to the promoter. This stimulates the production of mRNA. While this is happening the allolactose is bound to the inactive lac repressor to ensure that lots of mRNA is produced. When both lactose and glucose are present: Don't have anything to bind to the CAP protein, doesn't bind to next to the promoter, doesn't help mRNA to come in. Some lac proteins, but not at a high rate. Mostly using glucose, a small amount of lactose used. RNA polymerase less likely to bind, the inactive lac repressor is bound. Glucose determines how fast you make the proteins, lactose is the ON/OFF switch, glucose is like the dimmer switch.
E. coli lac operon is inducible:
When there is no lactose, there is no need to make enzymes that digest lactose. Active repressor (protein) binds to the operator of the gene. Active repressor ensures that RNA Pol cannot bind the promoter when the repressor is bound to DNA. When there is lactose available there is also allolactose (inducer). This allolactose binds to the active repressor and inactivates it. This is not a corepresser, it is an inducer. Then the RNA polymerase can bind and make RNA for making proteins for lactose.