Zhang Problem Set 1

1.
a. During development of multicellular organisms the fate of
millions of cells needs to be determined. Transcription factors can
determine cell fate by activating a specific subset of genes available in
the genome. Since transcription factors work either alone as well as in
combination to make such developmental decisions, a large number of
determination events can be specified by a much smaller number of
transcription factors. In theory, how many cell fates could be specified
by four transcription factors?
b. Transcription factors typically recognize a specific binding
sequence (within enhancer elements) to activate or repress a
set of target genes needed to specify cell fate. Describe
different ways that a transcription factor could be involved in
specifying more than one cell fate?
2.
A house fly mutant (ang1-1) lacking antenna was found to have a
mutation in the coding region of a gene encoding a homeodomain
transcription factor. The gene was named ANTENNA GONE1 (ANG1).
a. When and where would you predict the ANG1 gene is
transcribed. What major assumption are you making in your
hypothesis?
b. List at least three methods you could use to test your
hypothesis assuming that ANG1 gene has been identified and
isolated. Which test would you choose to do if you did only
one and explain why?
3.
Three new house fly mutants lacking antennae were discovered
subsequent to the discovery of the ang1-1 mutant. All three new
mutants were mapped to different positions (loci) of the house fly
genome (ANG2, ANG3, ANG4). Two of the genes (ANG2, ANG4) were
identified as encoding new homeodomain transcription factors. It was
found that ANG1 transcript was present in the wild type and ang1-1
mutant but not in ang2 or ang4 mutants. Conversely, ANG2 and ANG4
transcripts were found in each of wild type, ang1-1, ang2 and ang4
mutants.
a. Hypothesize a molecular model that is consistent with the
results described above for the function of ANG1, ANG2 and
ANG4 proteins (include a diagram).
b. A DNA fragment that included the ANG1 promoter and 3000bp
of DNA upstream (away from the coding region) was fused to
the lacZ reporter gene and transformed into flies. Forty

transformants carrying the chimeric gene were recovered but

none showed the expression pattern expected for ANG1.
Assuming that the sequence used was free of mutations, what
could have gone wrong?

c. The ang3 mutation mapped to the ang1 locus and failed to

complement ang1 (crossing the ang3 mutant to the ang1
mutant could not rescue the phenotype) suggesting that ang3
is a new allele of ANG1 (which we will call ang1-2). Unlike in
the original mutant (ang1-1), ANG1 transcript was not found.
Using these data and assuming that the mutation is a point
mutation (not a deletion) provide two hypotheses concerning
what part of the ANG1 gene the ang1-2 mutation disrupts.
4
.

Gene X
A
motif 1

motif 2

motif 3

motif 4

enhancer

promoter exon 1

enhancer
intron

exon 2

Transcription factor A binds Motif 1; Transcription factor B binds Motif 2;

Transcription factor C binds Motif 3; Transcription factor D binds Motif 4;

Transcription factors A, C and D are positive regulators of gene X. Any one of

the three alone bound to their respective enhancer can induce transcription.
However A and C cannot induce transcription when transcription factor B is present.
Transcription factor D is unaffected by transcription factor B.
---Using these data answer the following questions.

5.
The CERX gene of Arabidopsis thaliana encodes a condensing
enzyme required for epidermal wax biosynthesis. The gene has been
cloned (3 exons and 2 introns) and a polyclonal antibody made against
the CERX protein. The antibody was used to show that the CERX
protein is present in epidermal cells but not in other cell types.
a. List three general ways Arabidopsis could achieve such
epidermal specific localization of the CERX protein.
b. Briefly describe how you could use molecular techniques to
differentiate between the three possibilities you listed in part
a.
6.
If cells in a developing C. elegans embryo are ablated with a
laser, typically the adult will lack specific tissues or organs. In
contrast, the same experiment done on an A. thaliana embryo will
normally have little effect on the adult plant. Formulate a hypothesis
that can explain this difference.
7.
You generate a population of enhancer trap lines in the plant
Arabidopsis thaliana by transforming cells with an enhancer trap
construct and regenerating whole plants from single transformed cells.
Each of the transformed plants is heterozygous for a single enhancer
trap inserted in one place in the genome but a different place in each

independent transformant. You screen all the transformants (T1) for

expression of the reporter gene in flowers and find one plant that has
expression only in developing petals. This transformant was allowed to
self fertilize to produce T2 progeny. Sixtyfive T2 progeny were
examined for floral defects and 16 were found to lack petals. All 16
plants were homozygous for the enhancer trap insertion. If we assume
that the petal expression pattern is due to the enhancers of a gene
needed for petal development that you name PETALATA, provide
answers to the following questions:
a. Formulate a hypothesis to explain the origin of the petal-less
mutants observed in the population, carefully explaining your
logic.
b. Diagram the petalata gene in the enhancer trap line showing
the location of the enhancer trap such that your answer is
consistent with your hypothesis in part a.
c. Describe an experiment that could test some aspect of your
hypothesis.
d. If all 65 T2 plants were examined for reporter expression in
the flower petals how many would have such expression?
8.
Insulin is a protein that is secreted by the pancreas when the
level of blood sugar is high. It acts on many different tissues by
binding and activating a receptor tyrosine kinase that results in a
reduction in blood sugar. Diabetes is a disease in which the level of the
blood sugar remains high because some aspect of the insulin signaling
pathway is defective. One type of diabetes (A) can be treated by
giving the patient insulin while another type (B) does not respond to
such treatment.
a.

Which type of diabetes is likely to be due to a deficiency in

the pancreas and which type is likely due to a deficiency in
the target cells? Explain.

b.

Type B diabetes can be due to a mutation in one of several

genes. List several types of proteins that might be
encoded by such genes?