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48 Cards in this Set

  • Front
  • Back
Give the differences between eukaryotes and prokaryotes in transcription.
Eukaryotes are not organized into operons and they aren’t transcribed together into one RNA molecule. Each gene has its own promoter and is transcribed separately. Chromatin structure affects gene expression, because DNA must unwind before transcription can happen. Also, repressors and activators are in both types of cells, but activators are more common in eukaryotes.
What must happen before DNA can be transcribed? What must be present?
For a gene to be transcribed, transcription factors, activators, and RNA polymerase must bind to the DNA. Before transcription, the chromatin structure changes and the DNA becomes more available to the transcription machinery.
What happens as genes become more active in transcription? Where are DNase hypersensitive sites found?
As genes become active in transcription, the regions around the genes become more sensitive to DNase I. These areas are called DNase hypersensitive sites, and they can develop 1000 nucleotides upstream of the start site of transcription -- the chromatin here loosens.
What are the three different processes that affect gene regulation?
Three different processes affect gene regulation by changing chromatin structure: the modification of histone proteins, chromatin remodeling, and DNA methylation.
What are the two domains of histones in the core of a nucleosome?
Histones in the octamer core of a nucleosome have two domains: a globular domain that associates with other histones and the DNA, and a positive tail domain that interacts with the negative phosphate groups on the DNA backbone.
What is the histone code?
The histone code is when the tails of histone proteins are modified by the addition or removal of phosphate groups, methyl groups, or acetyl groups. They code for information that affects how the genes are expressed.
What is one type of histone modification that has to do with the addition of groups? What is the effect?
One type of histone modification is the addition of methyl groups to the tails of histone proteins. These modifications can either activate or repress transcription; it depends on which amino acids in the histone tail get methylated.
What happens in the acetylation of histones?
In acetylation, acetyl groups (CH3CO) are added to histones. This stimulates transcription. The addition of one acetyl group to lysine 16 in the tail of histone H4 prevents 30-nm chromatin fibers from forming, so the chromatin ends up being open.
What do acetyl groups do to chromatin?
Acetyl groups normally destabilize the chromatin to let transcription take place.
What happens in chromatin remodeling?
In chromatin remodeling, chromatin-remodeling complexes (some transcription factors and regulatory proteins) alter chromatin structure, without altering the chemical structure of the histones.
Where do the chromatin-remodeling complexes bind, and why?
Chromatin-remodeling complexes bind directly to particular sites on DNA and reposition the nucleosomes, which allows transcription factors to bind to promoters and initiate transcription.
Describe the remodeling complex SWI-SNF.
The chromatin remodeling complex SWI-SNF is found in yeast, drosophila, humans, and others. The complex uses energy from ATP to reposition nucleosomes, which exposes promoters in the DNA to other regulatory proteins and RNA polymerase.
Give one of the two ways that a remodeling complex can reposition nucleosomes. (nucleosome slide!)
A remodeling complex repositions nucleosomes by (1), causing the nucleosome to slide along the DNA, so the DNA that was wrapped around the nucleosome occupies a position between nucleosomes so it is more open.
Give one of the two ways that a remodeling complex can reposition nucleosomes. (shape change!)
A remodeling complex repositions nucleosomes by (2), causing a conformational change in the DNA, the nucleosomes, or both so that the DNA that is bound to the nucleosome becomes more open.
Describe DNA methylation.
DNA methylation is the methylation of cytosine bases, which yields 5-methylcytosine. Heavily methylated DNA is associated with transcription repression, while normally active DNA is un-methylated in most eukaryotes.
Where is DNA methylation most common, and what happens?
DNA methylation is most common on cytosine bases adjacent to guanine nucleotides (CpG). DNA regions with many CpG sequences are called CpG islands, and they are found near transcription start sites. When transcription is stopped, they are methylated, but the methyl groups are removed before transcription initiation.
What are transcriptional activator proteins, and what do they do?
Transcriptional activator proteins stimulate and stabilize the basal transcription apparatus at the core promoter. The activators may interact directly with the basal transcription apparatus or indirectly though protein coactivators.
Give one function of a transcriptional activator protein. (Bind DNA . . . )
Transcriptional activators bind DNA at a specific base sequence (normally a consensus sequence in a regulatory promoter or enhancer); they contain one or more DNA-binding motifs like the helix-turn-helix, zinc finger, and leucine zipper.
Give one function of a transcriptional activator protein. (Interacting . . .)
Transcriptional activators can interact with other components of the transcriptional apparatus, and influence the rate of transcription.
Describe the GAL4 example of a transcriptional activator.
An example of a transcriptional activator protein is GAL4, which regulates the transcription of several yeast genes in a galactose metabolism. The genes that control galactose metabolism are inducible: when galactose is gone, the genes are not transcribed and the proteins that break down galactose don’t get produced.
What does the GAL4 do, and what is its general look?
GAL4 has several zinc fingers and binds to a DNA sequence UASg (the upstream activating sequence for GAL4, it is like an enhancer). When they are bound, GAL4 activates the transcription of the yeast genes that galactose needs to be metabolized.
Describe enhancers. What do they do?
Enhancers can affect transcription at distant promoters. Activator proteins bind to the enhancer and cause the DNA between the enhancer and the promoter to loop out, which brings the promoter and the enhancer close together, so the transcriptional activator proteins can directly interact with the basal transcription apparatus.
Describe insulators and their relation to enhancers.
Insulators (boundary elements) limit the effects of enhancers. They are DNA sequences that block or insulate the effect of enhancers in a position-dependent way. If there is an insulator between the enhancer and the promoter, the enhancer can’t affect the promoter anymore.
Describe the heat-shock protein example.
Several eukaryotic genes can be activated by the same stimulus. For example, many eukaryotic cells respond to extreme heat and stress by making heat-shock proteins that help to prevent damage.
Describe coordinately expressed genes, and how they are different in bacterial and eukaryotic cells.
Groups of bacterial genes are often coordinately expressed (turned on and off together), because they are physically clustered as an operon and they have the same promoter. However, coordinately expressed genes in eukaryotic cells are not clustered.
How do coordinately expressed genes respond to the same stimulus?
Coordinately expressed genes in eukaryotic cells can respond to the same stimulus because they have regulatory sequences in common in their promoters or enhancers.
Describe response elements in relation to heat-shock genes.
Different eukaryotic heat-shock genes, for example, have a common regulatory element upstream of their start sites. These sequences are called response elements, and they normally have short consensus sequences.
What are response elements?
Response elements are common regulatory elements in coordinately expressed eukaryotic genes, like heat-shock genes. They normally have short consensus sequences.
What is the metallothionein gene?
The metallothionein gene protects cells from the toxicity of heavy metals by encoding a protein that binds to heavy metals and removes them from cells. It is a single gene controlled by several response elements. Other response elements upstream of the metallothionein gene contribute to increasing its rate of transcription.
What are the MREs? What do they do?
Several copies of a metal response element (MRE) are upstream of the metallothionein gene. Heavy metals stimulate the binding of activator proteins to MREs, which makes the transcription rate higher for the metallothionein gene.
What happens to the transcription rates when there are multiple copies of MRE?
Since there are multiple MRE copies, high transcription rates are induced by metals. Two enhancers are located in the upstream region of the metallothionein gene: one contains a response element TRE, which stimulates transcription around the activated protein API.
What does alternative splicing allow mRNA to do?
Alternative splicing lets pre-mRNA be spliced in different ways, which makes different proteins in different tissues or at different development times. Many eukaryotic genes use alternative splicing to regulate genes.
What does alternative splicing do in Drosophila (overview)?
Alternative mRNA splicing regulates genes in Drosophila by deciding on how the gene is expressed to make a fly male or female. Sex in Drosophila comes from a “cascade” of gene regulation.
What happens in female Drosophila when the X:A ratio = 1?
When the X:haploid chromosome ratio is 1, a female-specific promoter is activated in development to stimulate the transcription of a sex-lethal gene (Sxl). The protein encoded by Sxl regulates the splicing of the pre-mRNA transcribed from another gene, a transformer (tra).
What happens in female Drosophila when the splicing of tra pre-mRNA makes a Tra protein?
The splicing of tra pre-mRNA ends up making a Tra protein. Together with Tra-2, Tra stimulates female splicing of the mRNA from the doublesex gene (dsx). This all produces a female-specific Dsx protein, which causes the embryo to have female characteristics.
What happens in male Drosophila embryos when the X:A ratio = 0.5?
In male Drosophila embryos, which have an X:A ratio of 0.5, the promoter that transcribes the Sxl gene in female is inactive, so no Sxl protein is produced, so the tra pre-mRNA is spliced at a different 3’ splice site to make a non-functional Tra protein form. This nonfunctional Tra protein in males causes dsx pre-mRNAs to be spliced differently from those in females, and a male-specific Dsx protein is made, which causes the embryo to have male characteristics.
What is RNA interference?
The expression of a number of eukaryotic genes in controlled with RNA interference, also known as RNA silencing or post-transcriptional gene silencing.
What triggers RNA interference, and what happens?
RNA interference is triggered by small RNA molecules known as microRNAs (miRNAs) and small interfering RNAs (siRNAs). The enzyme dicer cleaves and processes double-stranded RNA to make siRNAs or miRNAs that are 21-25 nucleotides in length.
What are the four ways that miRNAs and siRNAs regulate gene expression?
miRNAs and siRNAs regulate gene expression in four ways: (1) cleavage of mRNA, (2) inhibition of translation, (3) transcriptional silencing, and (4) degradation of mRNA.
Describe RNA cleavage.
RNA cleavage is when RISCs that contain an siRNA (some have an miRNA) pair with mRNA molecules and cleave the mRNA near the middle of the bound siRNA. After the cleavage, mRNA is further degraded.
Describe the inhibition of translation.
Inhibition of translation is when some miRNAs regulate genes by inhibiting the translation of their complementary mRNAs. They can inhibit both the initiation step of translation and steps after translation initiation, like those that cause premature termination.
Describe transcriptional silencing.
Transcriptional silencing is when other siRNAs silence transcription by altering chromatin structure. These siRNAs combine with protiens to form a “RITS complex” (for RNA transcriptional silencing). The siRNA part of RITS binds to its complementary sequence in DNA or RNA while being transcribed, and represses transcription by attracting enzymes that methylate the tails of histone proteins.
What happens when methyl groups are added to the histones during transcriptional silencing?
When methyl groups are added to the histones, they bind to DNA more tightly, and restrict the access of proteins and enzymes that are needed to carry out transcription.
Describe the degradation of mRNA.
Degradation of mRNA is a final way that miRNAs regulate gene expression. The miRNA triggers the decay of mRNA in a process without Slicer activity. The miRNA binds with a sequence complementary to the consensus sequence in the AU-rich element. Then the miRNA binds to the AU-rich element and degrades the mRNA in a process with Dicer and RISC.
How do ribosomes, aminoacyl tRNAs, initiation factors, and elongation factors affect the rate of translation and gene expression?
Since ribosomes, aminoacyl tRNAs, initiation factors, and elongation factors are all needed for translation, their availability affects the rate of translation, so they can influence gene expression.
Explain how insulin affects gene regulation.
Insulin stimulates the initiation of overall protein synthesis by increasing the availability of initiation factors.
Explain how translation can be regulated by specific mRNAs.
Translation can be regulated by specific mRNAs. The initiation of translation in some mRNAs is regulated by proteins that bind to an mRNA’s 5’ UTR and keep the ribosomes from binding, like how repressor proteins bind to operators and prevent structural genes from being transcribed.
How are many proteins modified after translation?
Many proteins are modified after translation with cleavage and trimming of amino acids, acetylation, or by adding phosphate, carboxyl, methyl groups or carbohydrates to the protein.