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80 Cards in this Set
- Front
- Back
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Gene Expression |
the process of converting information archived in DNA into molecules that actually do things in the cell.
It occurs when a protein or other gene product is synthesized and active |
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E. Coli |
bacteria that lives in your body (it vastly outnumbers your own cells)
They use a wide array of carbohydrates to supply the carbon and energy they need. As your diet changes from day to day the availability of different sugars in your intestine varies. Each type of nutrient requires a different membrane transport protein to bring the molecule into the cell and a different suite of enzymes to process it.
Precise control of gene expression allows E. Coli to use the available sugars efficiently.
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Why is precise control over gene expression important? |
because bacterial cells from an array of species can be densely packed along your intestinal walls. All of these organisms are competing for space and nutrients. In this type of environment a cell has to use resources efficiently if it's going to be able to survive and reproduce.
If an individual synthesizes proteins it doesn't need then it is depleting its resources. Therefore they compete less successfully for the resources that are required to reproduce offspring. |
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Mechanisms for regulation of gene expression (in bacteria) |
1. Transcription control 2. Translation control 3. Post-translation control |
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Transcription control |
DNA -x-> mRNA--> Protein --> activated protein
The cell can avoid making the mRNAs for particular enzymes. If there is no mRNA then ribosomes cannot make the gene product. Transcriptional control occurs when regulatory proteins affect RNA polymerase's ability to bind to a promoter and initiate transcription |
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Translation Control |
DNA --> mRNA-x-> Protein --> activated protein
If the mRNA for an enzyme has been made, the cell could prevent the mRNA from being translated into protein. Translation control occurs when regulatory molecules alter the length of time mRNA survives, or affects translation initiation or elongation |
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Post-Translation Control |
DNA --> mRNA--> Protein -x-> activated protein
Many proteins have to be activated by chemical modification, such as te addition of a phosphate group.
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Methods of Regulation of Gene Expression in Bacteria |
1. Transcription control - saves the most energy for the cell as it stops it earliest
2. Translation control - allows the cell to make rapid changes in the amount of different proteins because the mRNA is already present and available for translation
3. Post-translation control- provides the most rapid response of all three mechanisms because only one step is needed to activate an existing protein (uses the most energy)
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Constitutively |
genes that are transcribed all the time
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Gene Expresion is a spectrum |
Gene expression is not an all-or-none proposition. Genes are not just "on" or "off" - instead, the level of expression can vary between these extremes
The ability to regulate gene expression allows cells to respond to changes in their environment |
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Metabolizing Lactose in E. Coli |
E. coli can use a wide variety of sugars for ATP production via cellular respiration or fermentation. Glucose is the preferred carbon sources.
It can use lactose but it has to break it down into glucose and galactose. The enzyme B-galactosidase catalyzes the reaction to break it down. And glucose is absorbed in the glycolytic pathway while other enzymes convert galactose to a substance that can be processed in the glycolytic pathway.
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Inducer |
A small molecule that triggers transcription of a specific gene |
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Inducer in E. coli for Lactose |
lactose itself regulates the gene for B-galactosidase - meaning that lactose acts an inducer
B-galactosidase is only produced when lactose is present and glucose isn't |
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How they identified regulating genes in E. coli |
1. generate a large number of individuals with mutations at random locations in their genomes. (exposing E. Coli to mutagens)
2. Screen the treated individuals for mutants with defects in the process or biochemical pathway in question- in this case, defects in lactose metabolism
They were looking for cells that cannot grow in an environment that contains only lactose as an energy source |
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Replica Plating |
1. Grow a master plate - Containing only glucose 2. Press velvet-covered block against master plate - transfer some cells 3. Press block against a replica plate - containing only lactose as an energy source 4. compare plates -cells that can use lactose as an energy sources grow into colonies, cells hat cannot are unable to metabolize lactose |
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Genes involved in lactose metabolism |
1. cells that cannot cleave lactose, even in the presences of inducer - Means there is no B-galactosidase ( the gene for it is defective lacZ-)
2. Cells cannot accumulate lactose - No membrane protein (galactoside permease) to import lactose; the gene for galactoside permease is defective (lacY-)
3. Cells cleave lactose even if lactose is absent as an inducer - Constitutive expression of lacZ and lacY; gene for regulatory protein that shuts down lacZ and lacY is defective - it does not need to e induced by lactose (lacI-)
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lacZ- Mutant in E. Coli
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1. cells that cannot cleave lactose, even in the presences of inducer - Means there is no B-galactosidase ( the gene for it is defective lacZ-) |
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lacY- Mutant in E. Coli
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Cells cannot accumulate lactose - No membrane protein (galactoside permease) to import lactose; the gene for galactoside permease is defective (lacY-) |
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lacI- Mutant in E. Coli |
Cells cleave lactose even if lactose is absent as an inducer - Constitutive expression of lacZ and lacY; gene for regulatory protein that shuts down lacZ and lacY is defective - it does not need to e induced by lactose (lacI-)
The hypothesis concluded was the that the normal lacI gene prevents the transcription of lacZ and lacY when lactose is absent. |
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Three Genes involved in lactose metabolism |
lacZ, lacY, lacI
lacZ and lacY both code for proteins which are required for the metabolism and import of lactose, while lacI is responsible for some sort of regulatory function. when lactose is absent the lacI gene or gene product shuts down the expression of lacZ and lacY. But when it is present they are produced. |
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Mapping of the three genes involved in lactose metabolism revealed what? |
They discovered that the genes are close together. This may suggest that lacZ and lacY might be transcribed together |
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Negative Control |
this occurs when a regulatory protein called a repressor binds to DNA and shuts down transcription
lacZ and lacY are under a negative control, and lacI acts as the repressor until lactose interacts with it as an inducer releasing the 'break' and allowing synthesis of lacZ and lacY to begin
Acts like a break on a car |
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Positive Control |
this occurs when a regulatory protein called an activator binds to DNA and triggers transcription
Acts like a gas pedel on a car |
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operon |
a set of coordinately regulated bacterial genes that are transcribed together into one mRNA. |
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lac operon |
the group of genes involved in lactose metabolism that are transcribed together into one mRNA |
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enzyme transacetylase |
the lacA gene codes for this enzyme and it is found adjacent to lacY and lacZ.
This enzyme catalyzes reactions that allow certain types of sugars to be exported from the cell when they are to abundant and could harm the cell. |
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Jacob-Monod model of lac operon regulation |
1. the lacZ, lacY, and lacA genes are adjacent are transcribed into one mRNA initiated from the single promoter of the lac operator. (cotranscribed)
2. the repressor is a protein encoded by lacI that binds to DNA and prevents transcription of the lac operon genes (lacX, lacY, and lacA). The repressor binds to a section of DNA in the lac operon called the operator
3. The inducer (lactose) binds to the repressor. When it does it changes the shape. This causes the repressor to come of the DNA. This is called the allosteric regulation where a small molecule binds to a protein and causes it to change its shape and activity. When the inducer binds to the repressor, the repressor can no longer bind to DNA and transcription can proceed. |
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Allosteric regulation |
where a small molecule binds to a protein and causes it to change its shape and activity. When the inducer binds to the repressor, the repressor can no longer bind to DNA and transcription can proceed. |
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How does Glucose inhibit the expersion of lac operon |
When both glucose and lactose are present in the environment the transport of lactose into the cell is inhibited, Because lactose does not accumulate in the cytoplasm the repressor remains bound to the operator. However when glucose levels outside the cell are low, galactoside permease is activate and lactose is transported into the cell inducing the lac operon expression. |
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inducer exlusion |
the mechanism of glucose preventing the transport of inducer
this is what allows E. coli to preferentially use glucose even when there are other sugar outside of the cell. |
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Why has the lac Operon been so important? |
1. it identified that gene expression is regulated by physical contact between regulatory proteins and specific regulatory sites in DNA.
2. It suggested that many bacterial genes and operons are under negative control by repressor proteins. |
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ara operon |
it is an example of a positive control where an activator protein binds to a regulatory sequence in DNA when genes are turned on. When bound to DNA the activator interacts with RNA polymerase to increase the rate of initiating transcription.
contains three genes that allow E. Coli to use the sugar arabnose. |
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AraC |
When arabinose sugar is present the transcription of the ara operon is turned on by an activator protein called AraC.
It codes for araB, araA, and araD which are required for arabinose metabolism
It also is a repressor as when there isn't any arabinose the two copies of AraC protien remain together, while one araC copy remains bound to the initiator the other copy now binds to a different regulatory site in the ara operon and it works to repress the transcription of both the ara operon and the araC gene. |
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Global Gene Regulation |
the coordinated regulation of many genes. |
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Regulon |
grouping genes into a regulon- a set of separate genes or operons that contain the same regulatory sequence and that are controlled by a single type of regulatory protein.
They allow bacteria to respond to challenges taht include shortages of nutrients, sudden changes in temperature, expose to radiaton or shifts in habitat. |
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Regulon with positive and negative controls |
Positive control- when an environmental change triggers the removal of the repressor protein from all the operators every gene in the regulon is transcribed
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Gene expression within eukaryotes |
your cells respond to their environment, just as bacteria and unicellular eukaryotes however the cells in a multicellular eukaryote express different genes in response to changes in the internal environment - specifically to signals from other cells. |
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Differential gene expression |
This is responsible for creating different cell types arranging them into tissues, and coordinating their activity to form the multicellular society we call an individual |
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Ways gene regulation occurs in Eukaryotes |
1. chromatin remodeling 2. transcription 3. RNA processing 4. mRNA stability 5. Translation 6. Post-translational modification |
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chromatin |
in eukaryotes DNA is wrapped around proteins to create a structure called chromatin
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Chromatin Remodeling |
the stretch of DNA containing promoter must be released from tight interactions with proteins, so that RNA polymerase can make contact with the promotor |
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RNA processing |
The steps required to produce a mature, processed mRNA from a primary RNA transcript.
Ex: splicing- if different cells use differnt splicing patterns, different gene products result |
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mRNA Stability |
the mRNA life span is regulated in eukaryotes: mRNA that remain in the cell for a long time tend to be translated more than mRNAs that have a short life span |
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why does Chromatin Remodeling happen? |
for a molecular signal to trigger the transcript of a particular gene, the chromatin around the target gene must be remodeled. This must happen as DNA is so tightly packed that the RNA polymerase cannot access it |
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Chromatin Basic Structure |
1. chromatin consist of DNA complexed with histones and other proteins.
2. Each Nucleosomes has DNA wrapped almost twice around it with a core of 8 histone proteins. Between each pair of nucleosomes there is a "linker" stretch of DNA.
This structure occurs because DNA is negatively charged (They have phosphate groups) and histones are positively charged (they have two positively charged amino acids).
2. the 30 nm fibers are attached at intervals along their length to proteins that form a scaffold or framework inside the nucleus.
3. When the chromosomes condense the scaffold proteins and the 30-nm proteins fold into larger more tightly packed structures |
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Nucleosomes |
Any of the subunits that repeat in chromatin; a coil of DNA surrounding a histone core |
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Histones |
Any of various simple water soluble proteins that are rich in the basic amino acids lysine and arginine and are complexed with DNA in the nucleosomes of eukaryotic chromatin. |
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Chromatin Altering |
The chromatin must be decondensed to expose the promoter so RNA polymerase can bind to it.
Evidence of Decondensed Chromatin
1. DNases are enzymes that cut DNA. Some DNases cleave DNA at random locations and these cannot cut efficiently if DNA is tightly wrapped.
2. Histone mutants- the lack of histone proteins prevent the assembly of normal chromatin. When chromatins are in their normal state eukaryotic genes are turned off - when DNA is wrapped into a 30-nm fiber the parking brake is on |
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How is chromatin Altered |
1. enzymes add methyl groups to DNA 2. Enzymes chemically modify histones 3. Macromolecular machines actively reshape chromatin |
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DNA methylation |
DNA methyltransferases add methyl groups (-CH3) to cytosine residues in DNA
methylated CpG sequences are recognized by proteins that trigger chromatin condensation. Actively transcribed genes usually have low levels of methylated CpG near their promoters, and non-transcribed genes usually have high levels o methylated CpG. |
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Histone Modification |
A large set of enzymes adds a variety of chemical groups to specific amino acids of histone proteins. These include phosphate groups, methyl- groups, short polypeptide chains, and acetyl groups (-COCH3)
Addition of these groups to histone promotes condensed or decondensed chromatin depending on the specific set of modifications. |
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Hisdone code |
this postulates that particular combinations of histone modifications set the state of chromatin condensation for a particular gene |
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Histone acetyltransferases (HATs) |
they add acetyl groups to the positively charged lysine residues in histones
On switch for transcription |
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Histone deacetylass (HDACs) |
remove acetyl groups from positively charged lysine residues in histones
Off switch for transcription |
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Acetylation |
this usually results in decondensed chromatin, a state associated with active transcription.
When HATs add acetyl groups they neutralize the positive charge on lysine and loosen the close association of nucleosomes with DNA allowing g binding sites for other proteins that help open the chromatin |
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chromatin-remodeling complexes |
These are enzymes that harness the energy in ATP to reshape chromatin. It causes nucleosomes to slide along the DNA or knock the histines completely off the DNA to open up stretches of chromatin and allow gene transcription |
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epigenetic inheritance |
pattern of inheritance that are due to something other than differences in DNA sequences. It implies another level of inheritance that adds to standard DNA-based mechanisms to explain how different phenotypes are transmitted |
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promoter |
the site in DNA where RNA polymerase binds t initiate transcription |
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TATA binding protein (TBP) |
after the TATA box has been exposed by chromatin remodeling the first step in initiating transcription is binding of the TATA binding protein.
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Promoter-proximal elements |
regulatory sequence that are located close to the promoter and bind regulatory proteins
they have sequences that are unique to specific sets of genes. They create a mechanism for eukaryotic cells to express certain genes but not others. |
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Enhancer |
Regulatory sequences that are far from the promoter and activate transcription
1. They can be more than 100,000 base pairs away or located within an intron
2. There are many types of enhancers that exist
3. Most genes have more than one enhancer
4. They usually have binding sites for more than one protein
5. Enhancers can work even if their normal 5-3 orientation is flipped or if they are moved to a new location
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Transcriptional activators |
when regulatory proteins bind to enhancers and transcription occurs |
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Silencers |
Eukaryotes that possess regulatory sequences that are similar in structure and share key characteristics with enhancers but work to inhibit transcription |
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Repressors |
When regulatory proteins called repressors bind to silencers transcription s shut down. |
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Regulatory Transcription Factors |
enhancers are binding sites for activators and repressors that regulate transcription. All of these proteins are termed regulatory transcription factors.
Different cells express different genes because they have different transcription factors. |
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How do transcription factors recognize specific DNA sequences |
There are major and minor groves with differences in composition and shape that can be recognized by transcription factors |
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Basal transcription facots |
these are proteins that interact with the promoter and are not restricted to particular genes or cell types. They are necessary for transcription to occur but they do not provide much in the way of regulation.
Ex: TATA-binding protein |
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Mediator |
A large complex of proteins called mediator acts as a bridge between regulatory transcription factors, basal transcription factors, and RNA polymerase II |
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Model for Transcription initiation in Eukaryotes |
1. transcriptional activators bind to DNA and recruit chromatin remodeling complexes and histone acetyltransferases (HATs)
2. the chromatin remodeling complexes and HATs open the chromatin which includes the promoter, promoter proximal elements, and enhancers
3. other transcriptional activators bind to the newly exposed enhancers and promoter-proximal elements;basal transcription factors bind to the promoter and recruit RNA polymerase II
4. Mediator connects the transcriptional activators and basal transcription factors that are bound to DNA. this step is made possible through DNA looping. RNA polymerase II can now being transcription. |
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Post-Transcriptional Control |
1. splicing RNAs in various ways 2. Modifying the life span of mRNAs 3. Altering the rate at which translation is initiated 4. Activating or inactivating proteins after translation has occured |
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Alternative splicing of mRNAs |
introns are spliced out in the nucleus as the primary RNA is transcribed. This is done through spliceosomes. During splicing gene expression is regulated when selected exons are removed from the primary transcript along with the introns. As a result this can yield different mature mRNA. |
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mRNA stability and RNA interference |
RNA interference occurs when a tiny single-stranded RNA held by a protein complex binds to a complementary sequence in an mRNA. this event unleashes either the destruction of mRNA or a block to the mRNA's translation |
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RNA interference process |
1. Transcription of a mRNA gene - it forms a hair pin 2. The RNA is trimmed by enzymes n nucleus creating a double-stranded hairpin that is exported to the cytoplasm 3. The enzyme cuts out the hair pin to form a double-stranded RNA molecule with only 22 nucleotides 4. One of the strands is taken up by a group of proteins called the RNA-induced silencing complex (RISC). This strand is called the microRNA (miRNA) 5. Once its part of the RISC, the miRNA binds to its complementary sequence 6. IF the match is perfect and enzyme in the RISC destroys the mRNA by cutting it in two. If it isnt perfect than the translation is inhibited
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Activation or inactivating a protein |
Instead of waiting for transcription, RNA processing, and translation to occur the cell can keep an existing but inactive protien waiting in the wings and then quickly activate it in response to altered conditions.
Speed is gained at the expense of efficiency |
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Proteasome |
When a protein needs to be destroyed enzymes mark it by adding many small copies of a polypeptide called ubiquitin and a proteasome recognizes the tag and cuts them into short segments. |
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What causes Cancer |
1. Genes that stop or slow the cell cycle 2. Genes that trigger cell growth and division by initiating specific phases in the cell cycle |
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Tumor suppressor |
proteins that stop or slow the cell cycle when conditions are unfavorable for cell division
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proto-oncogenes |
genes that stimulate cell division |
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oncogene |
an allele that promotes cancer development |
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p53 tumor suppressor |
it codes for a regulatory transcription factor that serves as a master brake on the cell cycle.
In mutant cells that lack the p53 that can bind to enhancers, DNA damage cannot arrest the cell cycle, and the cell cannnot kill itself, and damage DNA is replicated
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