CMB2001 In-course exercise 2023

Instructions:
This is a formative exercise to be completed in your own time. It is envisaged that
this should take approx 45 mins.
Feedback and answers will be provided in a seminar/support session on Nov 17th 10-
11am

The exercise comprises a mixture of single answer MCQs and multiple

response questions
30 questions in total
Answer all questions in the space below the question
All questions are worth 1 mark
1. A transcription factor is known to regulate the expression of eight different genes.
Inspection of the DNA sequences of the promoter regions of these genes identified a
number of binding sites for this transcription factor. An alignment of the sequences is shown
in the diagram.

G C A N N C G
T
Based on this information, what is the consensus binding site for this transcription
factor?

Select one alternative:

List
a. GCTTNCG
b. G C A/T N T C G
c. G/A C A/T N N C G
d. G C A/T N N C G
e. G C/A A/T N N C G

Answer
2. The graph shows the mRNA levels of five different genes after fibroblast cells were
treated with either trichostatin (an inhibitor of zinc-dependent histone deacetylases) or
splitomycin (an inhibitor of NAD-dependent histone deacetylases).

Based on this information, which of the genes is most likely to have its transcription
repressed by a Sirtuin?

Select one alternative:

List
a. TPT1
b. ERG12
c. FOZ5
d. CYC8
e. SLP4

Answer
3. The positions of important regulatory elements in a promoter-lacZ reporter are shown in
(A). In (B) the results of β-galactosidase assays from cells harbouring plasmids containing
this reporter are shown; cells were either grown at a constant 37oC (black bars) or were
grown at 37oC and then incubated at 43oC for 1hr immediately prior to the assay (grey bars).
A series of mutant reporters were then constructed and assayed as described above. The
results are shown in (C).

(A) -295 -280 -260 -245 -54 -48 -40 -30 -22
+1

HSE HSE CAAT GC TATA lacZ

HSE = Heat Shock Element

CAAT = CAAT Box 300
(B)
-galactosidase Units
GC = GC Box
250
Wild Type
Reporter
200
150
100
50
0
(C) 37 oC 43 oC
for 1hr
300 300
Mutant 1 Mutant 2
-galactosidase Units
-galactosidase Units

250 250
200 200

150 150
100 100

50 50
0 0
37 oC 43 oC 37 oC 43 oC
for 1hr for 1hr

300 Mutant 3 300 Mutant 4

-galactosidase Units

-galactosidase Units

250 250
200 200

150 150
100 100

50 50
0 0
37 oC 43 oC 37 oC 43 oC
for 1hr for 1hr

What is the most likely identity of Mutant 1

Select one alternative:

List
a. A promoter with sequences between -48 and -54 deleted.
b. A promoter with sequences between -40 and -54 deleted.
c. A promoter with sequences between -40 and -295 deleted.
d. A promoter with sequences between -245 and -260 deleted.
e. A promoter with sequences between -245 and -295 deleted.

Answer
4. The positions of important regulatory elements in a promoter-lacZ reporter are shown in
(A). In (B) the results of β-galactosidase assays from cells harbouring plasmids containing
this reporter are shown; cells were either grown at a constant 37oC (black bars) or were
grown at 37oC and then incubated at 43oC for 1hr immediately prior to the assay (grey bars).
A series of mutant reporters were then constructed and assayed as described above. The
results are shown in (C).

(A) -295 -280 -260 -245 -54 -48 -40 -30 -22
+1

HSE HSE CAAT GC TATA lacZ

HSE = Heat Shock Element

CAAT = CAAT Box 300
(B)
-galactosidase Units
GC = GC Box
250
Wild Type
Reporter
200
150
100
50
0
(C) 37 oC 43 oC
for 1hr
300 300
Mutant 1 Mutant 2
-galactosidase Units
-galactosidase Units

250 250
200 200

150 150
100 100

50 50
0 0
37 oC 43 oC 37 oC 43 oC
for 1hr for 1hr

300 Mutant 3 300 Mutant 4

-galactosidase Units

-galactosidase Units

250 250
200 200

150 150
100 100

50 50
0 0
37 oC 43 oC 37 oC 43 oC
for 1hr for 1hr

What is the most likely identity of Mutant 2?

Select one alternative:

List
a. A promoter with sequences between -48 and -54 deleted.
b. A promoter with sequences between -48 and -260 deleted.
c. A promoter with sequences between -30 and -1 deleted.
d. A promoter with sequences between -54 and -245 inverted.
e. A promoter with sequences between -245 and -295 deleted.

Answer
5. The positions of important regulatory elements in a promoter-lacZ reporter are shown in
(A). In (B) the results of β-galactosidase assays from cells harbouring plasmids containing
this reporter are shown; cells were either grown at a constant 37oC (black bars) or were
grown at 37oC and then incubated at 43oC for 1hr immediately prior to the assay (grey bars).
A series of mutant reporters were then constructed and assayed as described above. The
results are shown in (C).

(A) -295 -280 -260 -245 -54 -48 -40 -30 -22
+1

HSE HSE CAAT GC TATA lacZ

HSE = Heat Shock Element

CAAT = CAAT Box 300
(B)
-galactosidase Units
GC = GC Box
250
Wild Type
Reporter
200
150
100
50
0
(C) 37 oC 43 oC
for 1hr
300 300
Mutant 1 Mutant 2
-galactosidase Units
-galactosidase Units

250 250
200 200

150 150
100 100

50 50
0 0
37 oC 43 oC 37 oC 43 oC
for 1hr for 1hr

300 Mutant 3 300 Mutant 4

-galactosidase Units

-galactosidase Units

250 250
200 200

150 150
100 100

50 50
0 0
37 oC 43 oC 37 oC 43 oC
for 1hr for 1hr

What is the most likely identity of Mutant 3?

Select one alternative:

List
a. A promoter with sequences between -40 and -54 deleted.
b. A promoter with sequences between -48 and -54 deleted.
c. A promoter with sequences between -22 and -295 deleted.
d. A promoter with sequences between -245 and -260 deleted.
e. A promoter with sequences between -245 and -295 deleted.

Answer
6. The positions of important regulatory elements in a promoter-lacZ reporter are shown in
(A). In (B) the results of β-galactosidase assays from cells harbouring plasmids containing
this reporter are shown; cells were either grown at a constant 37oC (black bars) or were
grown at 37oC and then incubated at 43oC for 1hr immediately prior to the assay (grey bars).
A series of mutant reporters were then constructed and assayed as described above. The
results are shown in (C).

(A) -295 -280 -260 -245 -54 -48 -40 -30 -22
+1

HSE HSE CAAT GC TATA lacZ

HSE = Heat Shock Element

CAAT = CAAT Box 300
GC = GC Box (B) -galactosidase Units
250
Wild Type
Reporter
200
150
100
50
0
(C) 37 oC 43 oC
for 1hr
300 300
Mutant 1 Mutant 2
-galactosidase Units
-galactosidase Units

250 250
200 200

150 150
100 100

50 50
0 0
37 oC 43 oC 37 oC 43 oC
for 1hr for 1hr

300 Mutant 3 300 Mutant 4

-galactosidase Units

-galactosidase Units

250 250
200 200

150 150
100 100

50 50
0 0
37 oC 43 oC 37 oC 43 oC
for 1hr for 1hr

What is the most likely identity of Mutant 4?

Select one alternative:

List
a. A promoter with sequences between -1 and -54 deleted.
b. A promoter with sequences between -48 and -54 deleted.
c. A promoter with sequences between -40 and -295 deleted.
d. A promoter with sequences between -245 and -260 deleted.
e. A promoter with sequences between -245 and -295 deleted.

Answer
7. A new compound (called KMTI223a) has been identified which acts as a specific inhibitor
of Suvar39. If cells are treated with KMTI223a which of the following scenarios is the most
likely?

Select one alternative:

List
a. Increased levels of histone H3 lysine 9 methylation
b. Increased levels of histone H3 lysine 36 methylation
c. Decreased levels of histone H3 lysine 9 methylation
d. Decreased levels of histone H3 lysine 36 methylation
e. No change in levels of histone H3 lysine 36 methylation or lysine 9 methylation

Answer
A8. The hormone β estradiol is suspected to influence the level of ZRRT2 mRNA. To
investigate this RNA is prepared from cells cultured in the presence and absence of β
estradiol. The RNA is subjected to northern blotting using probes for ZRRT2 and the
constitutively expressed gene, GAPDH. The results are shown in the Figure and the relative
levels of RNA are indicated.

Given these data, how much does exposure to β estradiol increase ZRRT2 mRNA
levels?

Select one alternative:

List
a. No increase
b. 1.3 fold
c. 1.9 fold
d. 2.7 fold
e. 5.0 fold

Answer
A9. What removes the structure that is ringed in the diagram?

Select one alternative:

List
a. DCP1/DCP2
b. DEAD box protein
c. Dicer
d. eIFG
e. exosome

Answer
A10. An osteosarcoma cell line was exposed to 4-Nitroquinoline 1-oxide (4-NQ) which
mimics UV irradiation. What is the most likely impact of 4-NQ on the level of p53 and its
phosphorylation at serine 15?

Select one alternative:

List
a. p53 levels increase with no change in phosphorylation of serine 15
b. p53 levels decrease with no change in phosphorylation of serine 15
c. p53 levels increase and phosphorylation of serine 15 increases
d. p53 levels decrease but phosphorylation of serine 15 increases
e. p53 levels do not change but phosphorylation of serine 15 increases

Answer
11. The lacZ gene from E. coli, which encodes β-galactosidase, is a commonly used reporter
gene. β-galactosidase activity can be determined using ortho-Nitrophenyl-β-galactoside
(ONPG) as a substrate. β-galactosidase hydrolyzes ONPG into galactose and ortho-
nitrophenol. The latter compound has a yellow colour that can be used to quantify enzyme
activity by measuring absorbance at 420 nm (OD420) in a spectrophotometer.
In order to measure expression from the yeast HQS1 promoter, cells containing a HQS1
promoter-lacZ reporter were collected for β-galactosidase assays. The following reaction mix
was assembled in an eppendorf: a 500 µl aliquot of buffer, a 200 µl aliquot of cells, and a
100 µl aliquot of 4 mg/ml ONPG.
Given that the molar mass of ONPG is 301.25 g/mol, what is the final mM
concentration of ONPG in the reaction mix?

Select one alternative:

List

a. 1.66 mM
b. 6.66 mM
c. 13.28 mM
d. 16.66 mM
e. 166.66 mM

Answer__________________________________
12. In order to measure expression from the yeast HQS1 promoter, cells containing a HQS1
promoter-lacZ reporter were collected for β-galactosidase assays. The following reaction mix
was assembled in an eppendorf tube: a 500 µl aliquot of buffer, a 200 µl aliquot of the cell
sample, and a 100 µl aliquot of 4 mg/ml ONPG.
The number of cells used in each assay can be estimated by measuring the absorbance of
the cell sample at 595 nm (OD595). An OD595 = 0.5 equates to 1 × 107 cells/ml.
A 40 µl aliquot of the cell sample was diluted with 960 µl H2O and the OD595 was found to be
= 0.35.
Given that 200 µl of an undiluted cell sample was added to the β-galactosidase
reaction, how many cells were used in the reaction mix?

Select one alternative:

List

a. 1.5 × 107 cells

b. 2.5 × 107 cells
c. 3.5 × 107 cells
d. 4.5 × 107 cells
e. 5.5 × 107 cells

Answer__________________________________
13. A transcription factor of 420 amino acid residues has been cloned and sequenced. In
order to define the important regions of this protein a number of truncated versions of the
protein were expressed and purified (see Figure A for details. Numbers refer to amino
acids). The ability of the protein to bind to a DNA sequence containing a specific binding site
was examined using an electrophoretic mobility shift assay (EMSA). The result of the EMSA
is shown in Figure B.

Given these data, in what region is the DNA binding domain located?

Select one alternative:

List

a. 220-370
b. 315-400
c. 315-420
d. 380-400
e. 380-420

Answer_________________________________
14. In order to identify genes that are regulated in response to Growth Hormone (GH) a
microarray experiment was performed on a cell line. A cDNA sample labelled with green
fluorescent dye was prepared from total mRNA derived from cells grown under control
conditions (no hormone present). Then a cDNA sample labelled with red fluorescent dye
was prepared from cells that had been exposed to GH for 4 hours. Equal amounts of the two
cDNA samples were mixed and hybridized to a microarray slide where each spot on the
microarray has a probe for a particular gene. A schematic diagram depicting the results from
of a subsection of the microarray slide is shown.

Key ns ns G ns
Y R G R
ns = no signal
R = red
R G ns Y ns Y ns Y
G = green
Y = yellow
G R ns Y Y ns R ns

R ns G G Y ns R Y

G R G ns ns Y G R

G G Y Y Y ns G G

Using this diagram, how many genes exhibit a change in expression in response to
Growth Hormone?

Select one alternative:

List

a. 9
b. 11
c. 13
d. 14
e. 22

Answer
15. To identify regions in a fission yeast chromosome that are subjected to heterochromatic
silencing a series of reporter strains was constructed using the ura4 reporter gene. Each
strain has the ura4 reporter gene inserted into a different locus: either the telomere
([Link] the meiosis genes (mei::ura4), the centromere (cen::ura4) or the mating type
genes (mat::ura4). (See Figure A). Total RNA was prepared from the reporter strains plus a
wild type strain which contains the ura4 gene at it natural euchromatic locus (ura4 wt). The
RNA was subjected to northern blotting using radiolabelled probes for ura4 and the
constitutively expressed act1 gene. The results are shown in Figure B.

Based on this assay, which region(s) of the fission yeast chromosome is/are
subjected to silencing?
Select one or more alternatives:

List

a. tel
b. cen
c. mat
d. mei
e. None of them

Answer____________________________________
16. Figure A shows sequences that are conserved at intron-exon boundaries in mRNA.
Figure B shows the sequence of a fission yeast gene around one intron. After splicing what
would be the sequence of the mRNA?

Select one alternative:

List

a. 5’-GUUAGGUGAGUCCU-3’
b. 5’-GTTAGGT-3’
c. 5’-GUUAGCUGGU
d. 5’-GUUAGGU-3’
e. 5’-GUUAGUGACGU-3’

Answer: _________________________________
17. Figure A shows sequences that are conserved at intron-exon boundaries in mRNA.
Figure B shows the sequence of a fission yeast gene around one intron. Five different
mutant fission yeast strains were constructed and the point mutations associated with each
is shown. Which mutation(s) would prevent splicing of the intron from the mRNA?

Select one or more alternatives:

List

a. 1
b. 2
c. 3
d. 4
e. 5

Answer: _________________________________
18. A compound has been identified that causes the activation of the TNF receptor. What
effect would you expect this compound to have on IB?

Select one alternative:

List

a. It would cause the translocation of IB to the nucleus

b. It would cause the export of IB from the nucleus
c. It would result in the stabilisation of IB
d. It would result in the proteolytic degradation IB
e. It would have no effect on the level or localisation of IB

Answer:
19. Which statement(s) about eukaryotic RNA polymerase II is/are CORRECT?

Select one or more alternatives:

List

a. It is responsible for the transcription of miRNA

b. It is responsible for the transcription of tRNA
c. Some of its subunits are homologous to those of bacterial RNA polymerase
d. It is located in the nucleolus
e. The largest subunit of has a series of repeats known as the carboxy terminal domain
(CTD)

Answer:
20. Which statement(s) about the spliceosome is/are CORRECT?

Select one or more alternatives:

List

a. It catalyses the removal of introns from pre-mRNA

b. Its function requires ATP
c. It is a small complex containing <20 proteins
d. It contains snRNAs that are responsible for the catalytic steps in splicing
e. It contains exonucleases required for the 3’ to 5’ degradation of mRNA

Answer:
21. What protein is shown in the figure below?

Select one alternative:

List

a. TAF
b. TBP
c. TFIIB
d. TFIID
e. TFIIH

Answer:
22. The diagram shows an RNA editing event that occurs in glutamate receptor mRNAs in
neurons. What enzyme catalyses this reaction?

Select one alternative:

List

Select one alternative:

a. APOBEC-1
b. FTP
c. IDAR1
d. ADAR2
e. AKLBH5

Answer:
23. PKR is a kinase that is activated by double-stranded RNA (viral infection). What
important regulator of translation does it phosphorylate?

Select one alternative:

List
a. eEF1
b. eEF2
c. eIF2α
d. eIF3
e. eIF4F

Answer:
24. The diagram shows the structure of an element in the 5’ UTR of a human mRNA. What
influence would you expect low iron levels to have on this mRNA?

Select one alternative:

List
a. increase the stability of the mRNA
b. decrease the stability of the mRNA
c. decrease splicing of the mRNA
d. increase translation of the mRNA
e. idecrease translation of the mRNA

Answer:
25. Which statement(s) about the m7GpppN 5' cap is/are CORRECT?

Select one or more alternatives:

List
a. it protects mRNA from degradation by 5’-3’ nucleases
b. it protects mRNA from degradation by 3’-5’ nucleases
c. it facilitates splicing
d. it facilitates export from the nucleus
e. it is critical for translation of most mRNAs

Answer:
26. Which of the following statements about pre-mRNA splicing is CORRECT?

Select one or more alternatives:

List
a. it causes a major change in the composition of the mRNA relative to the genomic
sequence
b. Almost all yeast genes have multiple introns
c. Splicing is protein catalysed event
d. Alternative splicing is key to providing multiple protein isoforms from one gene
e. Defects in mRNA splicing are linked to multiple genetic diseases

Answer:
27. Which was the first nuclear histone acetyl transferase to be identified?

Select one alternative:

List
a. Esa1
b. Gcn5
c. PCAF
d. Sas2
e. Tip60

Answer:
28. Which of the following is the main 3’ to 5’ exonuclease activity in the cell that is involved
in RNA turnover and processing and has multiple nuclease activities?

Select one alternative:

List

a. Endosome
b. Exosome
c. Spliceosome
d. Proteasome
e. XRN1

Answer:
29. What is the function of p14ARF?

Select one alternative:

List
a. To bind to p53 and inhibit its DNA-binding
b. To bind to Mdm2 and activate p53
c. To stabilise the interaction of Mdm2 with p53
d. To phosphorylate p53
e. To proteolytically degrade p53

Answer:
30. Concerning translation in eukaryotes: What protein interacts with both the 3’ end of the
mRNA and with eIF4G to mediate mRNA circularisation?

Select one alternative:

List

a. eEF1
b. eEF2
c. eEF3
d. eIF5A
e. poly A binding protein (PABP)

Answer: