Download 22 Cancer Genetics

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22
Cancer Genetics
CHAPTER SUMMARY QUESTIONS
2. Cancer is a group of diseases characterized by uncontrolled cell growth and the
spread of abnormal cells.
4. Hyperplastic cells accumulate additional mutations and become dysmorphic
(abnormal in addition to uncontrolled division); dysmorphic cells may acquire the
ability to separate from the mass of cells, migrate between cells, and enter the
bloodstream. A metastatic tumor is one where the cells have gained the ability to
travel throughout the body and establish secondary tumor locations.
6. They may occur in intragenic regions or introns, or they may affect wobble
positions; in these examples they will have no effect on protein structure. Mutations
that cause a change in amino acid sequence may not affect protein function if they are
conservative missense and/or affect nonessential amino acids. Mutations that destroy
protein function will not affect a cell's phenotype if that function is still performed—
for example, by proteins encoded by the other allele.
8. The role of these proteins is to regulate the cell cycle by causing the cell to arrest at a
cell cycle checkpoint. If Rb and p53 are inactivated, a cell may divide
inappropriately, resulting in hyperplasia.
10. Cyclin levels vary throughout the cell cycle, while cdk levels are fairly constant.
Expression of cdk’s at the level of functional protein requires an additional activation
step (association with a cyclin protein), which is reversible.
12. Precondensation. The period after the cell has received a signal to induce cell death
but before there are any visible signs.
Condensation. Loss of interactions between the dying cell and its neighbors;
degradation of the extracellular matrix (if any) and a decrease in the cytoplasmic
volume.
Nuclear condensation. DNA cleavage and redistribution to nuclear margins.
Fragmentation. Division of the apoptotic cell into apoptotic bodies.
Phagocytosis. Neighboring cells engulf the apoptotic bodies.
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14. APC, KRAS, SMAD4/DCC, p53. This is unusual because in most cases the order
of mutations is random rather than following a typical sequence.
EXERCISES AND PROBLEMS
16. a.
Hypomorphic or null alleles of tumor suppressor genes are recessive mutations
but will cause errors to accumulate in the cellular genome. Hypomorphic or null
mutations of protooncogenes are unlikely to cause cancer.
b. Hypermorphic alleles of tumor suppressor genes are unlikely to cause cancer.
Hypermorphic alleles of protooncogenes are likely to be oncogenic, because
they increase the amount of cell signaling that stimulated cell division.
18. We see one band that is common to both cell lines; this band must represent the
normal protooncogene. The fact that this band is present in both lines indicates that
the insertion of a virus has occurred in only one of the two copies of the gene present
in the clone 1 cell. If it had inserted within both genes, we should not have seen the
normal band. We see a larger fragment in clone 1, indicating that the DNA of the
virus does not contain a site for the restriction enzyme used and that the virus has
inserted within the restriction sites that define the band, lengthening the region
probed. Alternatively, the virus could contain a restriction site and has still inserted
in such a way as to lengthen the band probed by inserting between the sequence
probed and the original restriction site.
20. If the viral proteins completely prevent RB1 protein function, this would be very
similar to a homozygous loss of function mutation. If the viral proteins only reduce
the level of RB1 function, the situation would be more like a hypomorph. In either
case, the RB1 protein is still produced normally by the cell. A loss of function
mutation is different because it affects the production or function of the protein
directly.
22. Contact inhibition requires signals from nearby cells to repress cell division, or
influence the G1 restriction point. Signals that allow cells to ignore repressive signals
from neighbors will allow transition to S rather than G0. For anchorage dependence,
it seems the cells must normally require some sort of signal from a cell:cell or
cell:matrix connection in order to prevent apoptosis. Mutations in the pathway
through which this signal is mediated could prevent stimulation of apoptosis even if
not connected; such a cell could continue dividing and pass that trait on to daughter
cells.
24. The specific checkpoints affected will determine the nature of the cell cycle of each
type of cancerous cell. The key comparison is that the cell cycle functions without
normal controls—not necessarily that cell division is rapid. For instance, in many
tumors, a G1-S signal is excessive or constant and G1 phase will be very short,
leading to rapid division. In tissues such as the prostate, which normally have very
low rates of cell division, a mutation that interferes with G2-M arrest will allow
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cancerous cells to divide but will not affect the overall rate of cell division and thus
will result in a slow-growing tumor.
If the spindle assembly checkpoint in M has been disrupted, metaphase may be
shorter as the cell does not have to wait to ensure all chromosomes are attached to
spindle fibers. This type of mutation may not directly affect the cell cycle per se, but
will cause genomic instability and rapid development of abnormalities in the tumor
cells.
26. The extracellular signal binds to a receptor protein located in the cell membrane.
The intracytoplasmic portion of the receptor then interacts with other proteins such
as adapter proteins and G proteins to initiate a signal cascade composed of protein
kinases. Ultimately, phosphorylation of transcription factors leads to a specific
pattern of transcription factor binding at the upstream regulatory sites of cyclin D,
activating transcription.
28. a.
The initial Myc mutation is a hypomorph; along with a wild-type copy, this
allele produces an adequate amount of functional Myc for wild-type protein
activity. However, two hypomorphs produce so little functional Myc that Myc’s
normal role to maintain proliferation is compromised and cell division fails,
resulting in embryonic lethality. The suppressor mutation is likely to be another
gene that normally competes with Myc. If levels of Myc are normal, loss of
function at this gene results in relative overactivity of Myc and increased
proliferation. The reduced levels of Myc activity in the homozygous hypomorph
balance the levels of activity of the mutant allele at the new gene, returning the
cell cycle control to its usual balance.
b. Since these two proteins do not directly interact, there must be some other
protein that mediates their interaction. A straightforward method would be to
identify known targets of Myc regulation and study potential protein:protein
interactions between Myc, the target genes, and the unknown gene identified
earlier. If no known genes are identified, the yeast two-hybrid system could help
identify other proteins involved in this pathway.
30. a.
If free Bad is present, it will bind with Bcl-2, preventing Bcl-2 from binding
Bax. Free Bax will be cleaved and lead to apoptosis. Overexpression of Bcl-2
will allow binding to both Bax and Bad, preventing apoptosis unless Bax levels
are very high. This is potentially an oncogenic mutation, as cells that should
undergo apoptosis will be allowed to survive and continue through the cell
cycle.
b. Reduced levels of Bad will reduce the amount of Bcl-2 required to sequester
Bad protein and, conversely, increase the relative amount of Bcl-2 available to
bind Bax, preventing apoptosis unless Bax levels are very high. Again, this is
likely to have an oncogenic effect because cells that should undergo apoptosis
are allowed to survive and divide.
32. 20,000 lesions per day 3 days = 60,000 lesions. If 99% of these lesions are
repaired, 60,000 0.01 = 600 remain.
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34. Tissue-specific expression (for example, DCC—colon) of the wild-type gene will
often result in tissue-specific expression of the mutant allele, or tissue-specific
responses to loss of gene function. Some tissues are also more likely to be exposed to
mutagens, or more likely to accumulate a mutation due to rapid cell division or
inefficient DNA repair in that tissue. Normal levels of oxidative damage caused by
the presence of ROS will also vary between tissues, with tissues that have high
energy demands having higher levels of ROS.
CHAPTER INTEGRATION PROBLEM
a.
A first step would be sectioning and staining of the tissue. Various staining
techniques would allow visualization of cellular and extracellular structures. An
increased frequency of mitotic cells than would be expected for that tissue indicates
hyperplasia. In the epidermal reference sample provided, the only mitotic cells are
the basal layer. Abnormally shaped cells, presence of abnormal cytoskeletal
structures, and evidence of unusual cell behavior would be indicators of potential
malignancy. A further indication is proliferation of a single cell type, rather than the
mix of cell types typically present in the epithelial tissue. If a portion of the edge of
the growth is included, observation of the cells at the border would provide an
indication if the growth is fully contained or has potentially become migratory.
b. Preparation of a karyotype with FISH will provide detailed information regarding
chromosomal rearrangements or changes in the number of chromosomes present.
Alternative staining procedures such as G-banding may allow identification of
deletions, but are unlikely to add significant information to a FISH karyotype. PCR
amplification and analysis of genes that are known or suspected to play a role in
development of carcinomas from epithelial tissues is a more time intensive
procedure, and it may or may not lead to identification of specific mutations or
rearrangements. However, if specific mutations are suspected, PCR amplification
and individual gene analysis may provide additional useful information.
c. The best method for studying gene expression for a large number of genes is
microarray analysis. mRNA from healthy tissue can be compared to mRNA from the
biopsy to identify any changes. Subtractive cDNA hybridization and library
screening for differentially active genes is a second method that could be used. This
method is useful if cloning the genes of interest and further study is needed. For
diagnostic purposes, microarray analysis is faster and more accurate. A third
technique to consider is Northern blot analysis of mRNA. This procedure requires a
large amount of mRNA and is not technically feasible for a small amount of tissue
such as a typical biopsy. However, after removal of a large tumor, Northern blot
analysis of mRNA from the tumor could provide detailed information about the
expression of specific genes.
d. Genes 1, 4, and 8 are only active during tumor growth or late in the process of
carcinogenesis. As these genes seem to promote growth, they are potentially
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involved in growth factor response. Genes that are inactivated in the process of
tumorigenesis, such as 2, 5, 6, and 7, are more likely to be tumor suppressors. If they
are down-regulated, tumor formation is allowed.
e.
This will depend on the specific function of these genes. Sunburn 2 may be involved
in precancerous changes, but does not appear to play a role in late development of
tumors, as it is inactive both in healthy skin and in late or metastatic tumors. Sunburn
5 may play a role in preventing precancerous changes, as it is activated only in acute
exposure. One possible mechanism would be participation in DNA repair.
Preventing activation of these genes during the normal response to acute UV
exposure could potentially increase risk of future cancer development.
f.
Inhibition of genes that play a role in tumor development or that increase rates of cell
division could potentially reduce cancer development. Inhibition of genes that
protect against tumor development could have the opposite effect.
g.
In an adult patient without a history of skin cancer, new epidermal growths are more
likely to be caused by environmental exposure to mutagens such as UV radiation.
Such exposures are not heritable. However, a complete answer would require
information about the patient’s past history as well as a complete family history.
Heritable factors such as an underlying predisposition or genetic disease related to
DNA repair could be involved and would likely be identified in a thorough
discussion of personal and family history. No, the stage of development of the tumor
would not affect this reasoning. A relatively common type of skin tumor would
support it; if the growth were very rare and/or almost always seen only in cases of an
underlying predisposition, the possibility of a family history becomes more relevant.
h. No. The most common risk factors for breast cancer are sex (female) and age (most
breast cancers are diagnosed after the age of 70). Only about 15% of breast cancer
patients have a family history of breast cancer; of those only about 85% have an
identifiable genetic mutation.