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41 Cards in this Set
- Front
- Back
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What does it Gene expression mean |
Means that genes have to be expressed or silenced |
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How does cell differentiation occur |
Same nuclear genome can lead to different protein patterns based on which genes are expressed THUS Depends on change in gene expression rather than changes in nucleotide sequence of cell’s genome |
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2-dimensional gel electrophoresis |
Is a method of characterizing differences in proteins expressed by two different tissues --> Proteins are separated by charge in first dimension(x-axis) and molecular weight in the second dimension(y-axis)--> Left side = acidic Right side = basic Top = high molecular weight Bottom = low molecular weight |
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Somatic cell reprogramming |
Gives evidence that a differentiated cell contains all the genetic instructions necessary to direct the formation of a complete organism Mechanism Nucleus of unfertilized egg (non-differentiated cells) destroyed by UV light Skin cell (differentiated cells) taken from a frog Nucleus of skin cell placed in unfertilized egg → normal embryo forms |
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What are the 6 steps that eukaryotic gene expression can be controlled |
Transcriptional control RNA processing control Cytosolic gene expression control steps mRNA degradation control Translation control Protein activity control |
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What are the steps at which gene expression can be controlled in nucleus |
In the nucleus--> Gene expression is controlled by transcriptional control(DNA creating RNA transcripts) and gene expression is controlled by RNA processing(RNA transcript splicing into mRNA) |
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What are the steps at which gene expression can be controlled in cytosol |
In the cytosol gene expression is controlled differently; 1) RNA transport and localization control 2)mRNA degradation control (mRNA → inactive mRNA) Occurs especially when mRNA is damaged 3) Translation control (mRNA → protein) 4) Last step; Protein activity control (Protein → inactive protein or active protein) Protein modification can lead to protein being active or inactive |
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In transcription, Promoters and gene regulatory sequences are for? |
Important in determining if gene is turned on or off |
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In transcription, transcription factors are for |
Binding to specific promoter and gene regulatory sequences--> turning genes on and off |
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In transcription, RNA polymerases are for ? |
Producing the RNA transcripts aka mRNA used to build proteins |
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In transcription, open reading frames**(dont get) |
Encodes the proteins |
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DNA binding proteins such as transcription factors |
Bind to the surface of DNA without opening the DNA Double helix--> These transcription factors(Tf) bind DNA ionicly(non covalently) |
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Transcription factors and genetic switches |
Transcription factors recognize genetic switches Short DNA sequences are fundamental components of genetic switches |
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Helix-turn-helix motif |
One of the simplest and most common DNA binding motifs Consists of an α-helix, a turn, and another α-helix |
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Recognition helix |
Binds to the major groove Important in recognizing specific sequences in DNA |
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Second helix |
Stabilizes the interaction of recognition helix with the DNA major groove |
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Helix-turn-helix proteins bind DNA as a dimer |
The two copies of the recognition helix (red cylinder) bind DNA separated by exactly one turn of the DNA helix (3.4 nm) The other helix of the helix-turn-helix motif (blue color) stabilizes the recognition helix |
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Helix-loop-helix (HLH) motif |
d |
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What is used to indicate where DNA-binding proteins bind? |
Gel mobility shift assay--> Indicates that a protein binds to a specific DNA sequence DNA footprinting |
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How does Gel mobility shift assay work? |
Tube 1 DNA fragment is radioactively labeled DNA is negatively charged, migrates to positive electrode--> DNA forms single band Tube 2 DNA fragment is radioactively labeled Proteins such as nuclear protein extracts are mixed with labeled DNA molecules in the assay Free DNA -->Migrates fast through the poly-acryl-amide gel --> Forms furthest band corresponding to single band in tube 1 DNA covered with proteins --> Migrates slowly --> Stratified based on weight; heaviest moving shortest distance |
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How does DNA footprinting work |
Mechanism Family of single-stranded DNA molecules labeled at the 5’ end A specific region of DNA is protected from cleavage by a DNA-binding protein DNA undergoes random cleavage by nuclease or chemical --> Protein is removed from the protected region and the cleaved DNA strands are separated --------> Separation is performed via gel electrophoresis DNA “Footprint” occurs where no cleavage is observed in the DNA strand --> Footprint = region protected by protein attached to DNA Gel electrophoresis is conducted both with and without the DNA binding protein --> The two runs are compared |
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What technique is used to select and purify against specific transcription factors |
DNA affinity chromatagraphy |
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Polycistronic messenger RNAs |
A single mRNA encoding several different proteins |
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polycistronic mRNAs |
A feature of bacteria and prokaryotic systems--> Expression of genes regulated by specific proteins that bind to the promoter --> due to a single mRNA encoding several proteins In contrast, Eukaryotic systems have monocistronic mRNAs |
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Control of Prokaryotic gene expression is through |
Negative regulation by end products |
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Tryptophan and Prokaryotic gene expression |
The synthesis of tryptophan depends on the level of tryptophan present Excess tryptophan (Genes are off) --> End-product tryptophan binds to repressor → active repressor --> Active repressor binds to DNA Prevents binding of RNA polymerase → prevents transcription Lack of tryptophan (Genes are on) --> RNA polymerase remains bound to DNA --> Transcription of tryptophan continues |
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What are the mechanisms by which specific gene regulatory proteins control gene transcription in prokaryotes |
1) Negative regulation + Ligand binds --> Switches gene on by removing repressor protein 2) Negative regulation w Ligand bounded leads to repressor protein preventing transcription--> Removal of ligand switches gene on by removing repressor protein 3) Positive regulation + ligand --> Switches gene off by removing activator protein 4) Positive regulation w ligand allows bound activator protein to promote transcription--> removal of ligand leads to switching off of activator protein |
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How does the Dual control of the Lac Operon operate? |
Lac operon controlled by both glucose and Lactose concentrations In the presence of Glucose, CAP does not bind to the CAP-binding site In the absence of Lactose, Lac repressor blocks RNA synthesis start site Thus: + Glucose, + Lactose = Operon OFF: CAP not bound + Glucose, - Lactose = Operon OFF: CAP not bound, Lac repressor bound - Glucose, - Lactose = Operon OFF: Lac repressor bound - Glucose, + Lactose = Operon ON |
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What is the purpose of the Mediator protein complex |
Helps establish interactions between components within the promoter region Required to activate transcription |
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What is transcriptional synergy? |
Transcriptional synergy means that multiple transcription factors bind at once to yield an exponential increase in transcription |
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Eukaryotic gene regulatory proteins often...? |
Often assemble into complexes on DNA--> Tf tend to bind to DNA in groups while in solution factors will remain seperated |
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What allows multiple inputs to integrate at a promoter? |
DNA Looping--> Can contribute to gene activation at a distance |
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What are the ways Gene regulatory proteins can be activated? |
Protein synthesis – regulatory protein synthesized only when needed Ligand binding Covalent modification – phosphorylation activates regulatory protein Addition of second subunit – DNA-binding subunit activates regulatory protein Unmasking – inhibitory protein removed by phosphorylation Stimulation of nuclear entryRelease from membrane |
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A single gene regulatory protein can coordinate the expression |
Of several different genes--> A single gene regulatory protein interacts with different protein complexes on different genes to help regulate expression of those genes Example: glucocorticoid receptor( with glucocorticoids--> leads to high levels of gene expression |
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Condensed DNA-histone complex is |
chromatin |
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Basic unit of DNA packaging in eukaryotes |
Is the nucleosome |
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A nucleosome consists of |
A segment of DNA wound in sequence around eight histones |
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Ways to open chromatin structure in order to access DNA and stimulate transcription |
Nucleosome remodeling, Histone removal, Histone replacement, Specific pattern of histone modification |
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Specific pattern of histone modification |
acetylation, methylation of histones |
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Histone removal |
Histone chaperones remove histones leaving DNA |
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Nucleosome remodeling |
Chromatin remodeling complex moves nucleosomes around and loosens DNA-Histone complex |
