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7
Transcriptional Control
of Gene Expression

PART A: Linking Concepts and Facts

7.1 Control of Gene Expression in Bacteria
1. Operator constitutive mutants of the lac operon would
a. express the lac repressor constitutively.
b. block the binding of RNA polymerase to the promoter.
c. express -galactosidase constitutively.
d. prevent the inducer from binding to the repressor.

Ans: c

2. Which of the following statements regarding the E. coli two-component PhoR/PhoB regulatory system is (are)
true?
a. PhoB has kinase activity.
b. PhoB is a cytosolic protein.
c. Phosphorylated PhoB is an inactive transcriptional activator.
d. none of the above

Ans: b

3. How does binding of the lac repressor to the lac operator block transcription initiation?
a. lac repressor binding blocks RNA polymerase from interacting with DNA at the start site
b. lac repressor binding induces a DNase that cleaves the DNA at the transcription start site
c. lac repressor binding causes a conformational change in RNA polymerase
d. lac repressor binding induces a protease that degrades the sigma subunit of RNA polymerase

Ans: a

7.2 Overview of Eukaryotic Gene Control and RNA Polymerases

4. Most eukaryotic genes are controlled at the level of
a. transcription initiation.
b. transcription elongation.
c. transcription termination.
d. translation initiation.

Ans: d
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5. Which of the following is not a step in the run on transcription assay?

a. isolation of nuclei
b. incubation with 32P-labeled ribonucleoside triphosphate
c. exposure of cells to a labeled RNA precursor
d. hybridization of labeled RNA to cloned cDNAs

Ans: c

6. All of the following statements about the essential carboxy terminal domain (CTD) of RNA polymerase are true
except
a. The CTD is present in RNA polymerase I, II, and III.
b. The CTD can become phosphorylated.
c. The CTD is critical for viability.
d. The CTD of mammals contains more than 50 repeats of a heptapeptide.

Ans: a

7.3 Regulatory Sequences in Protein-Coding Genes

7. An enhancer
a. is a DNA element that stimulates transcription of eukaryotic promoters.
b. binds to RNA polymerase and stimulates transcription.
c. acts as a binding site for RNA polymerase.
d. interacts with repressor proteins to enhance transcriptional repression.

Ans: a

8. The TATA box

a. serves as a promoter sequence for genes transcribed by RNA polymerase III.
b. is located approximately 100 base pairs upstream of the start site for mRNAs.
c. is present in all eukaryotic genes.
d. acts to position RNA polymerase II for transcription initiation.

Ans: d

9. All the following elements can function as eukaryotic promoters except

a. a TATA box.
b. an initiator element.
c. CpG islands.
d. an enhancer.

Ans: d

7.4 Activators and Repressors of Transcription

10. Which of the following is not used in the electrophoretic mobility shift assay (EMSA)?
a. a radiolabeled DNA fragment
b. a polyacrylamide gel
c. a DNA binding protein
d. DNase I
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Ans: d

11. Which of the following proteins does not “footprint” the lac operon control region?
a. lac repressor
b. -galactosidase
c. RNA polymerase
d. cAMP-CAP

Ans: b

12. All the following statements about the Wilms’s tumor (WT1) gene or WT1 protein are true except
a. Loss-of-function WT1 protein leads to the development of kidney tumors.
b. The WT1 protein has a zinc finger binding domain.
c. WT1 protein is a transcription activator.
d. WT1 binds to the control region of the EGR1 gene.

Ans: c

13. A leucine zipper motif contains

a. a stretch of five leucine residues in a row.
b. a leucine residue at every seventh position.
c. a leucine residue complexed with a zinc ion.
d. an alternating leucine-alanine-proline structure.

Ans: b

14. Which of the following is not a structural motif found in a DNA-binding domain?
a. homeodomain
b. zinc-finger
c. helix-loop-helix
d. random-coil acidic domain

Ans: d

7.5 Transcription Initiation by RNA Polymerase II

15. Which of the following is the correct order of binding of general transcription factors to initiate transcription at
RNA polymerase II promoters?
a. TFIID, TFIIB, Pol II, TFIIH
b. PolII, TFIID, TFIIB, TFIIH
c. TFIIB, PolII, TFIIH, TFIID
d. TFIID, TFIIH, TFIIB, PolII

Ans: a

16. What is the function of TFIIH in the transcription initiation complex?

a. binding to the TATA box
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b. unwinding the DNA duplex

c. catalyzing the synthesis of RNA
d. all of the above

Ans: b

7.6 Molecular Mechanisms of Transcription Repression and Activation

17. All the following statements about heterochromatin are true except
a. Heterochromatin stains more darkly with DNA dyes than does euchromatin.
b. Heterochromatin contains more highly condensed DNA than does euchromatin.
c. Heterochromatin is associated with inactive genes.
d. Heterochromatin is more susceptible to DNaseI than is euchromatin.

Ans: d

18. All of the following events play a role in yeast mating type switching except
a. methylation of the silent-mating-type locus.
b. transcription of the gene at the MAT locus.
c. chromatin condensation at the silent mating type locus.
d. a recombination event known as gene conversion.

Ans: a

19. The mediator complex

a. can form a molecular bridge between activators of transcription and DNA replication machinery.
b. can function to maintain a promoter in a hypoacetylated state.
c. has histone acetylase activity.
d. none of the above

Ans: c

20. Transcriptionally inactive genes

a. are always located within euchromatin.
b. are not located within nucleosomes.
c. often are methylated.
d. are not resistant to DNase I.

Ans: c

7.7 Regulation of Transcription-Factor Activity

21. Lipid soluble hormones activate transcription by
a. binding to specific cell-surface receptors.
b. phosphorylating a protein kinase.
c. binding to a nuclear receptor.
d. inhibiting a histone deacetylase.

Ans: c
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22. Which protein domains are found in nuclear-receptor family members?

a. variable region, DNA-binding domain, ligand-binding domain
b. acetylase domain, DNA-binding domain, ligand-binding domain
c. variable region, acetylase domain, ligand-binding domain
d. variable region, DNA-binding domain, acetylase domain

Ans: a

23. Regulation of transcription by steroid hormones

a. involves hormone receptors normally found in the nucleus.
b. involves cytoplasmic hormone receptors that can move to the nucleus.
c. involves two ligase domains.
d. always activates transcription.

Ans: b

7.9 Other Eukaryotic Transcription Systems

24. Which of the following statement(s) regarding the transcription initiation and RNA Pol III is (are) true?
a. ATP hydrolysis is not required for initiation.
b. Pol III is responsible for synthesizing tRNAs and 5S-rRNA.
c. The promoter elements of tRNA genes lie entirely within the transcribed sequence.
d. all of the above

Ans. d

PART B: Testing on the Concepts

7.2 Overview of Eukaryotic Gene Control and RNA Polymerases

25. Define the terms cis-acting DNA sequences and trans-acting proteins.

Ans: Cis-acting DNA elements affect only the expression of genes on the same DNA molecule that are linked to the
DNA element. In contrast, trans-acting proteins are free to diffuse through the cell and can bind to any target
DNA sequence.

26. Describe the similarities and differences between prokaryotic and eukaryotic RNA polymerases.

Ans: In prokaryotes there is only one RNA polymerase, which consists of five subunits. In eukaryotes there are
three RNA polymerases. RNA polymerase I synthesizes ribosomal RNA; RNA polymerase II synthesizes
messenger RNA; and RNA polymerase III synthesizes tRNA and other small RNAs. The eukaryotic RNA
polymerases are more complex than the bacterial RNA polymerase. All three contain two large subunits and
12–15 smaller subunits. All three eukaryotic RNA polymerases contain subunits with some sequence
homology to the E. coli RNA polymerase subunits (, , and ´).

27. Describe the structure and function of the carboxy terminal domain (CTD) of RNA polymerase II.

Ans: The carboxy terminal domain (CTD) of RNA polymerase II consists of a heptapeptide repeat, with a consensus
sequence of Tyr-Ser-Pro-Thr-Ser-Pro-Ser. Yeast RNA polymerase II contains 26 or more repeats, while the
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mammalian RNA polymerase II contains 52 repeats. The CTD is critical for viability, and at least 10 copies of
the repeat must be present for survival. During formation of the transcription initiation complex, the CTD is
unphosphorylated. When the RNA polymerase transcribes away from the promoter, the CTD is phosphorylated
at serine and threonine residues. One hypothesis is that phosphorylation of the CTD causes the release of RNA
polymerase from the transcription initiation complex.

7.3 Regulatory Sequences in Protein-Coding Genes

28. What is the functional difference between enhancers and promoter proximal elements?

Ans: Enhancers can stimulate transcription from a promoter tens of thousands of base pairs away. In contrast,
promoter-proximal elements are located 100 to 200 base pairs upstream of the start site and usually lose the
ability to stimulate transcription from a promoter when moved only several tens of base pairs away.

29. Describe how the electrophoretic mobility shift assay (EMSA) and the DNase I footprinting techniques are
used to identify DNA-protein interactions.

Ans: In the electrophoretic mobility shift assay (EMSA), DNA-protein interactions are detected by changes in the
mobility of a DNA fragment bound to a protein. A DNA fragment containing a putative protein binding site is
first radiolabeled and then incubated in the presence of sequence-specific DNA binding proteins. The DNA
fragment containing a bound protein migrates slower in a gel, causing a shift in the location of the radiolabeled
DNA detected by autoradiography. In the DNase I footprinting technique, a DNA fragment is first labeled at
only one end with 32P. The radiolabeled DNA fragment is incubated with a DNA binding protein and then
digested with a limiting concentration of DNase I. The DNase I concentration is set such that on average each
DNA molecule is cut only once. The resulting DNA fragments are separated by denaturing gel electrophoresis
and visualized by autoradiography. In the absence of a DNA binding protein, a ladder of DNA bands is
detected on the autoradiogram. Binding of a protein to the DNA prevents DNase I from digesting the
radiolabeled DNA at the site of the DNA-protein interaction, resulting in a blank area (or “footprint”) in the
DNA ladder.

30. What is an enhancesome?

Ans: An enhancesome is a large nucleoprotein complex bound to an enhancer element. This complex is formed by
the cooperative assembly of transcription factors to their multiple binding sites in an enhancer.

7.4 Activators and Repressors of Transcription

31. How can transcription factors be purified using sequence-specific DNA-affinity chromatography?

Ans: Sequence-specific DNA-affinity chromatography is a technique that takes advantage of the binding specificity
of a protein to a specific DNA sequence. Once the DNA sequence to which a transcription factor binds is
identified, this DNA sequence can be coupled to a bead in a column. A protein mix containing the
transcription factor is applied to this column. Proteins that do not bind to the DNA fragment are washed out of
the column. The bound transcription factor can then be eluted from the column in the presence of a high
concentration of a salt.

32. Describe the structure and function of a zinc-finger motif.

Ans: A zinc finger is a structural motif found in DNA binding domains; it consists of a short length of the
polypeptide chain folded around a Zn2+ ion. The two basic classes of zinc finger domains are the C 2H2 and C4
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structures. The C2H2 zinc finger domain consists of two cysteine (C) and two histidine (H) residues bound to
one Zn2+ ion. The C4 zinc finger contains four cysteines bound to one Zn 2+ ion. The three-dimensional
structure of the zinc finger forms a compact domain, which can insert its  helix into the major groove of
DNA.

7.5 Transcription Initiation by RNA Polymerase II

33. Describe the structure of the RNA polymerase II transcription initiation complex.

Ans: The RNA polymerase II transcription initiation complex is a multiprotein complex. This complex consists of a
DNA promoter element to which general transcription factors (i.e., TFIIA, TFIIB, TFIID, TFIIE, TFIIH) bind
along with RNA polymerase II. This multisubunit nucleoprotein complex consists of 6070 polypeptides with
a mass of approximately 3 Mda and is nearly as large as a eukaryotic ribosome.

34. Describe the functional properties of TFIID and TFIIH.

Ans: TFIID is a large, multisubunit complex of approximately 750 kDa. TFIID consists of a 38 kDa TATA box–
binding protein (TBP) and 11 TBP-associated factors (TAFs). TBP is the first protein to bind to a TATA box–
containing promoter. TFIIH is the last protein to bind to the initiation complex. TFIIH contains helicase
activity, which unwinds the DNA duplex at the start site. As the polymerase transcribes away from the
promoter, a subunit of TFIIH phosphorylates the carboxy terminal domain (CTD) of RNA polymerase II.

7.6 Molecular Mechanisms of Transcription Repression and Activation

35. Describe the role of histone deacetylation and hyperacetylation in yeast transcriptional control.

Ans: Histone deacetylation/hyperacetylation is one mechanism for regulating transcriptional control in yeast.
Repressor proteins can cause deacetylation of histone N-termini in nucleosomes. Unacetylated histones
contain positive charges due to N-terminal lysines and interact strongly with DNA phosphates. These strong
interactions may restrict access of general transcription factors, thus leading to transcriptional repression. In
contrast, histones that have their N-termini hyperacetylated are neutralized, eliminating the strong electrostatic
interactions with the DNA phosphates. This more open chromatin configuration facilitates access of general
transcription factors and induces transcriptional activation.

7.7 Regulation of Transcription-Factor Activity

36. Describe how lipid soluble hormones, glucocorticoid for example, regulate gene transcription acting through
nuclear hormone receptors.

Ans: Glucocorticoid is a lipid soluble hormone that binds to a member of the nuclear hormone receptor family, the
glucocorticoid receptor (GR), and regulates gene transcription. In the absence of glucocorticoid, the GR in the
cytoplasm is bound to the protein HSP90. When glucocorticoid diffuses through the cell membrane, it binds to
the GR ligand-binding domain and causes a conformational change in the GR, releasing HSP90. The GR
bound to glucocorticoid is then translocated into the nucleus, where it interacts with glucocorticoid response
elements (GRE) and regulates transcription of responsive genes.

37. Describe the mechanism of transcriptional control for the heat shock genes. What advantage does this type of
control impart to the cell?
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Ans. During transcription of the heat-shock genes, RNA Pol II pauses after transcribing ≈25 nucleotides. Under
stress conditions, where intracellular proteins are denatured or may become denatured, heat shock transcription
factor (HSTF) is activated. In this state, HSTF binds to specific regions in the promoter of the heat shock
genes, stimulating RNA Pol II to continue chain elongation. Binding also facilitates the rapid re-initiation by
other RNA Pol II molecules, leading to a significant up-regulation in heat-shock-gene-expression. Thus the
mechanism of stalling the RNA Pol II and having partially completed transcripts ready to complete elongation
and readied for translation, when the need arises is a safeguard, protecting cells against unexpected, stressful
conditions.