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367 molecular biology > pp 16 > Flashcards

pp 16 Flashcards

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1
Q

Why is eukaryotic transcription regulated by chromatin structure?

A

Chromatin has to be open for promoter to be accessible to the transcription machinery. General transcription factors are needed to help recruit RNA polymerase to the promoter

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2
Q

What is the function of activators?

A
  1. Facilitate opening of chromatin
  2. Recruit general transcription factors and RNA polymerase as a preinitiation complex promoter. (can act with coactivators)
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3
Q

How do activators bind?

A

They recognize and bind regulatory DNA sequences called enhancers, and they can then stimulate transcription

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4
Q

What domains do activators have?

A

They have at least two functional domains, DNA-binding domain, transcription-activating domain and then some may have dimerization domain

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5
Q

What does the activation domain do?

A

Binds general transcription factors or mediator and helps initiation complex form

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6
Q

What are the different structures of DNA-binding domains?

A

Helix-turn-helix motif, homeodomain, zinc finger motif, leucine zipper, basic helix-loop helix

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7
Q

What are the characteristics of helix-turn-helix?

A

Found as part of larger DNA-binding domain.
Commonly found in DNA-binding domains of bacterial regulatory proteins (lac repressor, Y repressor)

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8
Q

What are the characteristics of homeodomains?

A

DNA-binding domains that include helix-turn-helix motifs found in large family of activators, consisting of three helices, second and third form helix-turn-helix.

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9
Q

What are the characteristics of Zinc finger?

A

30 residues forming an elongated loop held together at the base by a coordinated zinc ion (two cys and two His), Zinc stabilizes the motif, presents a recognition helix to the DNA

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10
Q

What are nuclear receptors?

A

Hormone-responsive activators that include zinc fingers in a DNA binding domain

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11
Q

What is the role of Zinc fingers in nuclear receptors?

A
  1. Type I nuclear receptors respond to steroid hormones, in absence of these hormones, sequestered in cytoplasm bound by Hsp70 but upon binding, Hsp70 dissociates, hormone receptor complex enters nucleus, binds hormone-response element, activating transcription
  2. Type II nuclear receptors bind DNA, but only interact with coactivator when it binds steroid hormone ligand
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12
Q

What are the characteristics of basic leucine zippers? (bZIP)

A

Dimerization. Transcription-activating domain is not shown.

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13
Q

What are the characteristics of Basic Helix-loop-helix (BHLH and bZIP)

A

both have basic (positively charged) DNA binding portion
ZIP refers to leucine zipper consisting of leucine residues on facing sides of the a-helices on each monomer facilitating dimerization by hydrophobic interactions.
HLH refers to the dimerization portion that consists of a helix-loop-helix motif

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14
Q

How and why do transcription factors bind as homodimers or heterodimers?

A
  • two members of the same family of similarity structured proteins can dimerize to make a heterodimer
  • combinations of heterodimers can recognize a range of sequences resulting from the combination of half-sites
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15
Q

What is the GAL4 protein?

A

Yeast transcription activator that acts as a dimer held together by a leucine zipper, while each DNA binding domain contains two Zinc ions

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16
Q

What is the function of GAL 4?

A

recognizes upstream activating sequences (UAS) of genes regulated by GAL4

17
Q

What is the GAL pathway?

A

galactose needs to be modified by GAL1,2,7,10 and 7 again to be converted to glucose 1-phosphate and so those genes are transcribed when galactose is present. Gal4 activator that binds to the enhancers called UAS

18
Q

How is the activity of Gal4 regulated?

A
  • galactose absent: Gal80 binds to transcription-activating domain of Gal4 and inhibits its activating function.
  • galactose present: binds Gal3 causing allosteric effect, increasing affinity for Gal80, then binds it and changes conformation preventing inhibition of Gal 4
19
Q

How is the combinatorial control of yeast regulated?

A

-glucose present: Gal genes repressed. Mig1 and Tup1 act as repressors (bind DNA directly) and corepressors (doesnt bind DNA). Mig1 only enters nucleis when glucose is present. if it isnt, Mig1 phosphorylated and cant enter nucleus.

20
Q

What are the other types of transcritpion-activating domains?

A
  1. acidic domains
  2. glutamine-rich domains
  3. proline-rich domains
21
Q

What are hybrid proteins?

A

Made with DNA-binding domain of one protein and the transcription-activating domain of another (Gal4-LexA hybird: activating domain of Gal4 can be fused to LexA)

22
Q

How is eukaryotic transcription regulated by chromatin structure?

A

activators recruit complexes to remodel/modify chromatin resulting in more open form. They then recruit general transcription machinery to promoter. Activation is reversible, resulting in more closed chromatin

23
Q

How is activation reversible?

A

repressors displace activators, blocking activation along with co-repressors, then recruit histone deacetylases, then deacetylation leads to condensed chromatin

24
Q

How do activators bind to enhancer sequences on DNA?

A

enchancers are DNA sequences that act as activator-binding sites, activators can bind the enhancers directly, co-activators bind activators instead.
together, they function to facilitates opening of chromatin, recruiting GTM or trigger RNA polymerase to release or clear the promoter and proceed along the DNA to elongate transcript

25
Q

What is the mechanism of activation?

A

activators + co-activators bind to enhancers at sites distant from promoter, recruiting components of GTM to a pre-initation complex to initiation complex.

26
Q

What is the mechanism of repression?

A

mediated by proteins that precent or disrupt essential contacts between transcription machinery and activators or co-activators

27
Q

What is DNA looping and why is it done?

A

activators that bind to a distant enhancer on DNA form a loop, allowing interaction with RNA polymerase at promoter. Transcription factors or architectural regulators can bend DNA significantly where they bind, facilitating looping.

28
Q

What is the role of co-activators and co-repressors during looping?

A
  1. co-activator can mediate interactions between activator and RNA polymerase
  2. co-repressor can bind and block interactions with RNA polymerase, preventing recruitment to the promoter
29
Q

What is the difference between regulators in eukaryotic genes and bacterial genes?

A
  • bacterial: have one or two transcription factors with binding sites.
  • eukaryotic: numerous regulator binding site spanning large region upstream or downstream of promoter
    (few transcription factors can regulate large number of genes)
30
Q

How can you tell if expression of gene is off or on?

A

specific combinations of transcription factors.
eve gene in fruit fly developement shows how

31
Q

What is the eve gene experiment?

A
  1. oocyte is surronded by nurse cells that secrete mRNA encoding various transcription factors
  2. diff. concentration gradients for mRNAs are established, when cell divides, it will have diff levels.
  3. mRNA translated, different ratios are observed. Fluorescently tagged antibodies if seen gene is activated, if not gene is repressed.
32
Q

What are enhancosomes?

A

Nucleoprotein complex containing a collection of activators bound to an enhancer in such a way that stimulates transcription in an all or nothing response.

33
Q

What are insulators?

A

DNA sequences that form boundaries between genes or groups of genes to prevent misregulation occuring from an enhancer of one gene interacting with the promoter of another gene. CTC-binding factor binds the insulator

34
Q

What is the signal transduction pathway?

A
  1. signaling molecule interacts with a receptor leading to a response in the nucleus
  2. Sends signal through the cell and frequently leads activator binding in nucleus
  3. Rely on protein phosphorylation to pass the signal from one protein to another

367 molecular biology (10 decks)

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