Quiz 9 Flashcards
(73 cards)
1
Q
- Understand the importance of DNA between enhancers and promoters.
A
- often looped out to allow binding between gene specific transcription factors and general transcription factors.
2
Q
- Understand the importance of enhancers
A
- modular
contain a constellation of binding sites for various transcription factors that integrate information from the environment. - Modular enhancers contain multiple binding sites for different transcription factors that can work independently
3
Q
- Understand the importance of multiple enhancers.
A
- multiple enhancers provide fine control over gene expression.
4
Q
- Understand the importance of homeotic genes in flies
A
- Homeotic genes in flies are used as examples of how enhancers and transcription factors work together to regulate gene expression.
- These genes are arranged in order of their expression
- Regions correspond to regions that will develop into adult structures
5
Q
- Be familiar with the homeotic genes and their function.
A
- Homeotic selector genes determines developmental fate of each segment
6
Q
Understand the conservation of homeotic genes
A
- conserved from flies to man
- are selector genes that determine the fate of cells along the anterior-posterior axis.
- control the development of structures found in different locations throughout the body.
7
Q
- Understand that a pathway consisting of maternal effect genes, gap genes, pair-rule genes and segment polarity genes are needed to regulate the expression of the homeotic genes.
A
- Each set of genes further divides the embryo into smaller and smaller regions and ultimately you get expression of homeotic genes that determine the fate of cells.
- Maternal effect genes regulate gap genes
- Gap genes regulate pair-rule genes (7 stripes)
- Pair rule genes regulate segment polarity genes (14 stripes)
- Segment polarity genes regulate homeotic selector genes
8
Q
- Understand the importance of mutations in pair-rule genes
A
- mutations in pair-rule genes causes the loss of every other segment in the fly embryo.
- Fushi taratzu = pair-rule gene
9
Q
- Understand the importance of different concentration of gap genes.
A
- different concentrations of gap genes bind enhancers within pair-rule genes to regulate their expression
- these enhancers are modular (just like all enhancers).
10
Q
- Understand the regulation of pair-rule expression.
A
- pair-rule gene expression is a result of the interaction of
- modular cis-regulatory enhancer elements of pair-rule genes
- trans-regulatory proteins (gap genes) that bind enhancers.
- all gene expression is a result of this
11
Q
- Understand how the importance of enhancers can be demonstrated experimentally:
A
- i) mutations in enhancers disrupt expression in certain stripes
- ii) fusing the enhancer to lacZ causes lacZ to be expressed in that stripe
- iii) deleting a gap gene that regulates a stripe changes the location of the stripe.
12
Q
- Understand the importance of different combination of activators
A
- Different combinations of activators produce different levels of expression in different cells.
- Provide fine control over gene expression
13
Q
- Understand what kind of switches enhancers can be considered
A
- enhancers can be considered binary on/off switches.
- Provide fine control over gene expression
14
Q
- Understand how multiple enhancers act.
A
- multiple enhancers act cooperatively to regulate the expression of a gene.
- Provide fine control over gene expresison
15
Q
- Understand what happens when enhancers are located close to promoters.
A
- enhancers located close to promoters makes looping more difficult
16
Q
understand what architectural transcription factors do.
A
- architectural transcription factors cause the DNA to bend to stimulate transcription.
- They change the shape of the DNA control region so that other factors can interact successfully to stimulate transcription
17
Q
- Understand the sequence specificity of architectural transcription factors.
A
- architectural transcription factors exhibit little sequence specificity but bind mainly to AT-rich regions.
18
Q
- Understand the importance of binding between certain nuclear receptors and their ligands.
A
- binding between certain nuclear receptors and their ligands
- changes the receptors from repressors to activators.
- causes the receptor to dissociate from inhibitory protein in the cytoplasm, translocate to the nucleus, and activate transcription.
19
Q
- Understand the role of phosphorylation in the regulation of transcription factors.
A
- It causes certain activators to interact with mediators that stimulate transcription
- marks certain inhibitor proteins for destruction by proteolysis
- marks certain activators for destruction by proteolysis.
20
Q
- Understand the difference between the two types of signaling:
A
- Signaling molecule enters the cell
- Signaling molecule does not enter the cell
21
Q
signaling molecule enters the cell
A
- signaling molecule enters the cell
- binds to transcription factor,
- an allosteric change occurs allowing transcription factor to bind to specific DNA sequences in enhancers to stimulate transcription
22
Q
signaling molecule does not enter the cell
A
- the signaling molecule does not enter the cell or the cell nucleus
- rather it binds to a transmembrane receptor protein and
sets off a cascade of phosphorylation cascades
resulting in phosphorylation of a transcription factor, causing - an allosteric change, allowing the transcription factor to bind to specific DNA sites in enhancers to stimulate transcription
23
Q
- Understand what mediator is needed for
A
- mediator is needed for the transcription of most eukaryotic genes
- Called a mediator because it mediates effect of an activator
24
Q
Understand how mediator functions
A
- it functions by binding to both general transcription factors and gene specific transcription factors.
25
2. Understand the effect of mediator on basal transcription
- mediator has no effect of basal transcription
| - rather it elevates the level of expression of activated genes
26
Understand the cAMP signal transduction pathway
- cAMP levels increase
- Stimulates protein kinase A to move into nucleus
- Phosphorylates CREB
- Stimulates transcription
27
function of CREB
- CREB: trans-acting transcription factor
- Binds CRE, gets phosphorylated
- Interacts with mediator (CBP) to stabilize interactions with basal complex to get high levels of transcription.
28
function of CBP
- CBP (CREB binding protein) is a mediator
- brings histone acetyl transferase to the enhancer
histones are acetylated,
- Binds to other proteins called SCRs
- CBP contacts basal transcription apparatus
- SRCs interact with nuclear receptors
- gene is activated
29
4. Understand the composition of the mediator in different cell types.
- the composition of mediator varies in different cell types, but contains some subunits that are found in most cell types.
30
5. Understand that CBP interacts with at least 3 pathways:
- growth factors
- nuclear receptors
- cAMP signaling.
31
What is epigenetics?
- the study of heritable changes in gene function that occur without a change in the sequence of DNA.
32
7. Define epigenome
- the collection of biochemical modifications to chromatin that indexes genetic information.
- Determine if the information can be expressed or not.
33
8. Understand the roles of histone acetylation and deacetylation.
- Acetylation leads to activation - Most common histone modification
- deacetylation leads to silencing
34
Hallmarks of transcriptionally active chromatin
- Histones acetylated (particularly H3 and H4)
- Histone H3 K4 (lysine) methylated
- DNase I sensitive
35
Hallmarks of transcriptionally inactive chromatin
- DNA methylated
- Histone H3 K9 (lysine) methylated
- DNase I resistant
36
Understand the role of histone H1 in packaging DNA
- Chromatin containing H1 leads to nucleosomes with normal zigzag orientation rather than the “beads on a string” orientation.
37
H1 effect on trancription
- H1 represses transcription; binds to linker DNA between nucleosomes with a transcription start site
38
11. Understand the competition between transcription factors and nucleosome cores in DNA sequences.
- in most cases DNA sequences cannot be simultaneously occupied by transcription factors and nucleosome cores.
- If factors win, gene is active
- If histones win, gene is repressed when its control region is tied up with cross-linked nucleotides
39
12. Understand nucleosome "phasing"
- Nucleosomes are “phased” at least over short distances
- Nucleosome phasing and positioning: mechanisms to ensure that specific regions are in linker DNA and thus available for binding
- Not uniform throughout genome.
40
13. Understand the importance of the positioning of a nucleosome
- the positioning of a nucleosome at a one position can result in an open DNA sequence (in linker DNA) at another position. This is the phenomenon of "phasing."
41
14. Understand more of the competition between transcription factors and histones
- some transcription factors can out-compete histones for control regions of regions after replication
- other transcription factors can bind to a site already occupied by a nucleosome (by binding to a chromatin remodeling factor).
42
15. Understand how i) nucleosome-positioning sequences, ii) competition sequence-specific DNA binding proteins and iii) chromatin remodeling complexes affect transcription.
- They are all ways that a nucleosome can be positioned so that phasing will result in accessible DNA at a specific site for transcription factor binding
43
16. Understand the roles of H3K4 and H3K9 methylation and how they interact.
- Methylation of H3 K9 is a hallmark of transcriptionally inactive chromatin
- methylation of H3 K4 is a hallmark of transcriptionally active chromatin.
44
Methylation of H3 K4
- blocks methyl transferase from methylating H3 at K9
| - blocks the chromatin remodeling complex, NuRD, which has histone deacetylase activity (HDAC)
45
1. Understand the importance of acetylation and deacetylation of the amino groups of lysines within histone proteins.
- acetylation of the amino groups of lysines within histone proteins, particularly H3 and H4, helps activate genes
- deacetylation promotes the inactivation of genes.
46
2. Understand which lysines are acetylated in active chromatin and deacetylated in inactive chromatin.
- lysines 9 and 14 of histone H3
- 5, 8 and 16 of H4
- acetylated in active chromatin and deacetylated in inactive chromatin.
47
3. Understand what happens when transcriptional repressors without their ligands interact with co-repressors.
- transcriptional repressors, such as nuclear receptors without their ligands, interact with co-repressors, such as NCoR and SMRT, which interact with histone deacetylases.
- These remove acyl groups from basic tails of core histones in nearby nucleotides to repress transcription
48
4. Understand what happens when nuclear receptors bind to their ligands
- nuclear receptors, after binding to their ligands, interact with with mediator and other factors that acetylate histones, which helps to activate transcription.
49
5. Understand what chromatin remodeling complexes do.
- chromatin remodeling complexes remove or move nucleosomes within regulatory regions of genes to help activate transcription.
50
6. Understand what the Swi/Snf family and Iswi family do
- the Swi/Snf family and the Iswi family are chromatin remodeling complexes and need ATP to remove nucleosomes from DNA.
- They move nucleosomes on DNA and yield nucleosome-free regions around enhancers and promoters in inactive genes
51
1. Understand the difference between euchromatin and heterochromatin
- euchromatin is relatively extended and open and potentially active
- heterochromatin is condensed and inaccessible.
52
2. Understand the difference between constitutive and facultative heterochromatin and where they are found in the genome.
- Constitutive heterochromatin:
- regions that remain condensed throughout the cell cycle
- Found in centromeres and telomeres
- Facultative heterochromatin:
- regions that can unfold and become euchromatin
53
3. Understand what Barr bodies are
- Barr bodies are X chromosomes in females that are highly heterochromatic and silent.
- Barr bodies are condensed, inactive members of a pair of X chromosomes in the cell.
- The other X of the pair is not condensed and is active in transcription
54
4. Understand that methylation is used in some organisms more than others to regulate chromatin structure and what type of methylation that genes, repetitive, and heterochromatic DNA have.
- Genes are hypomethylated
| - Retrotransposons, other highly repetitive DNA, and heterochromatin are hypermethylated
55
5. Understand how heterochromatin can spread within a genome
heterochromatin can spread within a genome by the binding of HP1, which
i) brings in SUV39H1 methylase to methylate histones
ii) DNA methyl transferases that methylate DNA.
56
6. Understand what MeCRP2 does.
- MeCP2 also binds methylated DNA and recruits a deacetylase which deacetylates H3K9 unless H3K4 is methylated.
57
7. Understand what happens if H3K9 is methylated.
- if H3K9 is methylated, it can recruit HP1 and heterochromatin spreading continues.
58
8. Understand the relationship between histone methylation, DNA methylation, and histone methylation.
- histone methylation (H3K9) leads to DNA methylation and DNA methylation leads to histone methylation.
59
9. Understand what maintenance methylases do.
- maintenance methylases ensure that both DNA strands are methylated after replication, but they are not 100% efficient.
60
10. Understand the concept of Harold Weintraub
- If DNA in chromatin is inaccessible to transcription factors and RNA polymerase in vivo, then it should also be inaccessible to endonucleases in vitro.
61
11. Understand the concept of genes being transcriptionally "poised" but not transcriptionally active.
- DNA of transcriptionally-repressed genes is DNase I resistant
- DNA of transcriptionally-poised genes is DNase I sensitive
62
12. Understand Weintraub’s experiments with DNase I sensitivity of different genes in different tissues in chicken chromatin.
It is the active genes that are DNase sensitive in each cell
- Isolate nuclei from tissue (developing erythrocytes or oviduct)
- Treat nuclei with increasing concentration of DNase I
- Isolate DNA from DNase treated nuclei
- Digest the DNA with a restriction enzyme
- Perform a southern blot.
63
14. Understand the distinction between DNase I sensitive and DNase I resistance.
- the DNA that is not "DNase I sensitive" is DNase I "resistant," although given enough time or high enough concentration of DNase I, essentially all DNA in chromatin can be cleaved by DNase I.
64
15. Understand the relationship between DNase I sensitivity and the 30-nm fiber
- DNase I sensitivity is generally thought to indicate unfolding of the 30-nm fiber.
65
16. What does our best evidence to date indicate about transcriptionally active genes and DNase sensitivity
- Our best evidence to date indicates that all transcriptionally active genes are DNase I sensitive but that not all DNase I sensitive genes are transcriptionally active.
- These are the transcriptionally poised genes.
66
17. Understand when DNase I sensitivity occurs and what this means.
- DNase I sensitivity can be shown to occur BEFORE transcription
- This means that a DNase-I sensitive structure is a PREREQUISITE for transcription but NOT the result of transcription.
67
18. Understand the confines of DNase I-sensitive domains.
- DNase I-sensitive domains can include large regions of chromosomes and include several genes.
68
19. Understand the three general levels of DNase I sensitivity
- a. DNase I resistant (even this DNA can be digested if extensively treated)
- 30-nm fibers and higher orders of folding
- b. DNase I sensitive
- involves large regions and generally thought to be due to unfolding of the 30-nm fiber
- C. DNase I hypersensitivity
- involves small regions and is generally thought to be due to a missing nucleosome
69
20. Understand the procedures used to identify and map DNase I hypersensitive sites. If given data from DNase I hypersensitive site mapping experiment, be able to map the site.
How sensitive something is to DNase-I shows how open and active it is.
70
21. Understand the role of insulators or boundary elements in chromatin.
- insulators or boundary elements separate chromatin in topologically separate domains
- separate euchromatin from heterochromatin.
71
understand the precise definition of exon
- the portions of the gene that are transcribed and appear in the mature mRNA
72
understand the precise definition of intron
- the portions of the gene that are transcribed but do not appear in the mature mRNA
- introns are flanked on both sides by exons and are removed during processing.
73
2. Understand how sequence information is presented at genome web sites.
- Black - non-transcribed DNA
- Yellow- translated DNA
- Red - transcribed but not translated DNA
- Purple - introns
