1. How Genes Are Regulated
Somatic Mutation and the Genetics of Cancer
Overview: Initiation of Division
Cancer: A Failure of Control over Cell Division
The Normal Control of Cell Division
Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Hartwell et al., 4th edition, Chapter 17

2. Two unifying themes about cancer genetics
Cancer is a disease of genes
Multiple cancer phenotypes arise from mutations in genes that regulate cell growth and division
Environmental chemicals increase mutation rates and increase chances of cancer
Cancer has a different inheritance pattern than other genetic disorders
Inherited mutations can predispose to cancer, but the mutations causing cancer occur in somatic cells
Mutations accumulate in clonal descendants of a single cell

3. The relative percentages of new cancers in the United States that occur at different sites
Fig. 17.1

4. Overview of the initiation of cell division
How do cells know when to divide?
Two basic types of signals that tell cells whether to divide, metabolize, or die
Extracellular signals – act over long or short distances
Collectively known as hormones
Steroids, peptides, or proteins
Cell-bound signals – require direct contact between cells
histocompatibility proteins
The macrophages, helper T cells, and antibody-producing B cells communicate in the presence of viral particles, bacteria, and toxins.

5. An example of an extracellular signal that acts over large distances
Thyroid-stimulating hormone (TSH) produced in pituitary gland
Moves through blood to thyroid gland, which expresses thyroxine
Fig. 17.2a

6. An example of an extracellular signal that is mediated by cell-to-cell contact
Fig. 17.2b

7. Each signaling system has four components
Growth factors
Extracellular hormones or cell-bound signals that stimulate or inhibit cell proliferation
Receptors
Comprised of a signal-binding site outside the cell, a transmembrane segment, and an intracellular domain
Signal transducers
Located in cytoplasm
Transcription factors
Activate expression of specific genes to either promote or inhibit cell proliferation
Signal transduction - activation or inhibition of intracellular targets after binding of growth factor to its receptor

8. Hormones transmit signals into cells through receptors that span the cellular membrane
Fig. 17.3a

9. Signaling systems can stimulate or inhibit growth
Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Hartwell et al., 4th edition, Chapter 17
Hormones bind to a specific cell surface receptor. The extracellular surface of the receptor transmits a signal to the intracellular domain of the receptor, which, in turn, interacts with other signaling molecules in the cell either to stimulate growth or to inhibit growth
Fig. 17.3b&c

10. RAS is an intracellular signaling molecule
Fig. 17.3d
Guanosine diphosphate(GDP)
Guanosine triphosphate(GDP
The RAS protein is an intracellular signaling molecule that is induced to exchange a bound GDP (inactive) for a bound GTP (active) when a growth factor binds to the cellular receptor with which RAS interacts

11. Cancer phenotypes result from the accumulation of mutations
Cancer: A Failure of Control over Cell Division
Cancer phenotypes result from the accumulation of mutations
Mutations are in genes controlling proliferation as well as other processes
Result in a clone of cells that overgrows normal cells
Cancer phenotypes include:
Uncontrolled cell growth
Genomic and karyotypic instability
Potential for immortality
Ability to invade and disrupt local and distant tissues
we use the term “tumor” to designate cancerous tissue
and the term “growth” to designate a benign mass.

12. a) Phenotypic changes that produce uncontrolled cell growth (Figure17
Most normal cells
Many cancer cells
Autocrine stimulation:
Cancer cells can make their own stimulatory signals
a.1
Most normal cells
Many cancer cells
Autocrine stimulation. Most cells “decide” whether or not to divide only after receiving signals from neighboring cells. Many tumor cells, by contrast, make their
own stimulatory signals, in a process known as autocrine stimulation, or they have become insensitive to negative signals.
Loss of contact inhibition. Normal cells stop dividing when they come in contact with one another, as evidenced by the fact that normal cell types that grow in
culture form sheets one cell thick. Tumor cells, in contrast, climb all over each other to produce piles that are many cells thick. This change contributes to the disordered
array of cells seen in tumors.
Loss of contact inhibition:
Growth of cancer cells doesn't stop when the cells contact each other
a.2
Fig. 17.4

13. Phenotypic changes that produce uncontrolled cell growth (cont)
Most normal cells
Many cancer cells
Loss of cell death:
Cancer cells are more resistant to programmed cell death (apoptosis)
a.3
Most normal cells
Many cancer cells
Loss of gap junctions:
Cancer cells lose channels for communicating with adjacent cells
Loss of cell death. Normal cells die when starved of growth factors or when exposed to agents that damage them. Programmed cell death (apoptosis) is activated
by the expression of certain genes in the cell; it is probably a safeguard against the early stages of cancer. Most cancer cells are much more resistant than normal cells to programmed cell death.
Loss of gap junctions. Normal cells connect to their neighbors by small pores, or gap junctions, in their membranes. The gap junctions permit the transfer of small molecules that may be important in controlling cell growth. Most tumor cells have lost these channels of communication.
a.4
Fig. 17.4

14. b) Phenotypic changes that produce genomic and karyotypic instability
Defects in DNA replication machinery:
Cancer cells have lost the ability to replicate their DNA accurately
Increased mutation rates can occur because of defects in DNA replication machinery
Defects in the DNA replication machinery. Cancer arises most often in cells that have lost the ability to reproduce their genomes faithfully. You saw in Chapters 6 and 7
that cells have elaborate systems for repairing DNA damage; these systems include the enzymatic machinery for mismatch repair and the repair of damage caused
by radiation or ultraviolet light. Work on yeast and bacteria has shown that mutant organisms defective in DNA repair have enormously increased rates of mutation.
These increased mutation rates often lead to cancer in multicellular organisms.
Fig. 17.4

15. Phenotypic changes that produce genomic and karyotypic instability (cont)
Increased rate of chromosomal aberrations:
Cancer cells often have chromosome rearrangements (translocations, deletions, aneuploidy, etc)
Some rearrangements appear regularly in specific tumor types
(the translocation between chromosomes 8 and 14 in patients with certain kinds of lymphoma )
Increased rate of chromosomal aberrations. Tumorcell karyotypes often carry gross rearrangements, including broken chromosomes, with some of the
pieces rejoined to other chromosomes; multiple copies of individual chromosomes, rather than the normal two; and deletions of large chromosomal segments
and of whole chromosomes. Studies have confi rmed that the fi delity of chromosome reproduction is greatly diminished in tumor cells. Normal fi broblast cells, for
example, have an undetectable rate of gene amplifi cation (an increase in the number of copies of a gene), whereas tumor cells have amplifi cation rates as high
as 1 in 100 cells. Probably only a small fraction of these chromosomal rearrangements lead to cancer; for example, tumors from solid tissues typically carry many chromosomal
rearrangements, but most of these aberrations do not recur in all tumors. A few rearrangements, however, regularly appear in specifi c tumor types.
Examples include the translocation between chromosomes 8 and 14 found in patients with certain kinds of lymphoma and the translocation between chromosomes
9 and 22 found in certain types of leukemias (see Fig on p. 492).
Fig. 17.4b.2
Fig. 17.4b.2

16. c) Phenotypic changes that produce a potential for immortality
Loss of limitations on the number of cell divisions:
Tumor cells can divide indefinitely in culture (below) and express telomerase (not shown)
Most normal cells
Many cancer cells
c.1
Immortality
Loss of limitations on the number of cell divisions. Most normal cells (except for the rare stem cells) die spontaneously after a specifi able number of cell divisions. Tumor
cells, by contrast, can divide indefi nitely.
2. Ability to grow in culture. Cells derived from tumor cells usually grow readily in culture, making cancerous cell lines available for study. Normal cells do not grow well in culture.
c.2
Growth in soft agar
Ability to grow in culture:
Cells derived from tumor cells usually grow readily in culture, making cancerous cell lines available for study.

17. d) Phenotypic changes that enable a tumor to disrupt local tissue and invade distant tissues
Ability to metastasize:
Tumor cells can invade the surrounding tissue and travel through the bloodstream
d.1
Angiogenesis:
Tumor cells can secrete substances that promote growth of blood vessels
d.2
The ability to metastasize. Normal cells stay within rigidly defi ned boundaries. Tumor cells, by comparison, often acquire the capacity to invade surrounding tissues and
eventually to travel through the bloodstream to colonize distant tissues. Metastasis—the invasion of other tissues— is a complicated behavior requiring many genetic
changes. wound. Tumor cells, however, secrete substances that cause blood vessels to grow toward them. The new vessels serve as supply lines through which the tumor can tap new
sources of nutrients and as escape routes through which tumor cells can metastasize.
3. Evasion of immune surveillance (not shown). The human immune system may recognize cancer cells as foreign and
attack them, thereby helping to eliminate tumors even before they become large enough for clinical detection. As
evidence, cancer patients often have antibodies and/or killer T cells directed against their cancer cells. Successful
tumor cells, however, somehow develop the ability to evade detection by the immune system.
Fig. 17.4

18. Multiple mutations leading to conversion
DNA sequencing of tumor cells has revealed thousands of mutations in each tumor, but how many actually contribute to the cancer phenotype is unclear because most of these mutations are different in different tumors of the same type.

19. Evidence from mouse models that cancer is caused by several mutations
b) Transgenic mice with dominant mutations in the myc gene and in the ras gene
The activated myc oncogene produces tumors more slowly than the ras oncogene. Mice containing both oncogenes develop tumors even faster than mice with ras.
b) Mice with recessive mutations in the p53 gene
the p53 gene is not essential for development or for normal cell function, but it does play a role in preventing tumor formation
Fig. 17.5a
Fig. 17.5

20. Evidence that cancer cells are clonal descendants of a single somatic cell
CLONAL PROLIFERATION
Analysis of polymorphic enzymes encoded by the X chromosome in females
Sample from normal tissues has mixture of both alleles
Clones of normal cells has only one allele
Sample from tumor has only one allele
Fig. 17.6

21. The role of environmental mutagens in cancer
The mutations that produce these cancers are not inherited through the germ line in a dominant or recessive pattern…
Concordance for the same type of cancer in first degree relatives (i.e. siblings) is low for most forms of cancer
The incidence of some cancers varies between countries (see Table 17.2)
When a population migrates to a new location, the cancer profile becomes like that of the indigenous population
Numerous environmental agents are mutagens and increase the likelihood of cancer
Some viruses, cigarette smoke

22. The incidence of some common cancers varies between countries
The role of environmental mutagens
The incidence of some common cancers varies between countries
Table 17.1
*Incidence indicates number of new cases per year per 100,000 population, adjusted for a standardized population age distribution (so as to eliminate effects due merely to differences of population age distribution). Figures for cancers of breast, cervix, and ovary are for women; other figures are for men.

23. Cancer development over time
Lung cancer death rates and incidence of cancer with age
(b) The incidence of most cancers shows a dramatic increase with age, a result thought to reflect the accumulation of mutations in somatic cells.
(a) Lung cancer death rates in the United States during the twentieth century began increasing rapidly for men in the 1940s and for women in the 1960s. This refl ects the fact that smoking became prevalent among men about 20 years before it did among women.
Fig. 17.7

24. Some families have a genetic predisposition to certain types of cancer
Cancers that run in families
Some families have a genetic predisposition to certain types of cancer
Example: retinoblastoma caused by mutations in RB gene
Individuals who inherit one copy of the RB− allele are prone to cancer of the retina
During proliferation of retinal cells, the RB+ allele is lost or mutated
Tumors develop as a clone of RB−/RB− cells
Fig. 17.8

25. Cancer is thought to arise by successive mutations in a clone of proliferating cells
Fig. 17.9

26. Cancer-producing mutations are of two general types:
Improperly activate genes (exp. the genes responsible for promoting cell proliferation)
Improperly inactivate genes (exp. the genes responsible for preventing excessive cell proliferation).
Fig

27. Oncogenes act dominantly and cause increased proliferation
Oncogenes are produced when mutations cause improper activation a gene
Two approaches to identifying oncogenes:
Tumor-causing viruses (Fig 17.11a)
Many tumor viruses in animals are retroviruses
Some DNA viruses carry oncogenes [e.g. Human papillomavirus (HPV)]
Tumor DNA (Fig b)
Transform normal mouse cells in culture with human tumor DNA

28. Cancer-causing retroviruses carry a mutant or overexpressed copy of a cellular gene
After infection, retroviral genome integrates into host genome
If the retrovirus integrates near a proto-oncogene, the proto-oncogene can be packaged with the viral genome
Fig a
Normal genes change to abnormally activated oncogenes through mutations that occur during viral propagation or through their placement near powerful promoters and enhancers in the viral genome

29. Retroviruses and their associated oncogenes
When a virus carrying one or more oncogenes infects a cell, the oncogenes cause abnormal proliferation that can lead to the accumulation of more mutations and eventually to cancer.
Retroviruses and their associated oncogenes
Table 17.2

30. DNA from human tumor cells is able to transform normal mouse cells into tumor cells
Human gene that is oncogenic can be identified and cloned from transformed mouse cells
the DNA responsible for the transformation of mouse cells by human tumor DNA can be identifed by reisolating the human DNA from the transformed mouse cells with probes for the short interspersed elements known as Alu sequences. These sequences appear only in the human genome
Fig b

31. The RAS oncogene is the mutant form of the RAS proto-oncogene
Normal RAS is inactive until it becomes activated by binding of growth factors to their receptors
Oncogenic forms of RAS are constitutively activated
They do this by encoding receptors, signal transmitters and transcription factors that are active with or without growth factor
Fig c

32. Oncogenes are members of signal transduction systems
This table and the previous one were labeled as Table 17.2 in the pdf, but the next one is 17.4
Table 17.3
Retroviruses identified as causative agents of tumors in animals contain oncogenes that were derived from a cellular gene. Adapted from Lewin, Genetics, 1e, Oxford University Press, Inc. by permission

33. Enhanced mutation potential in proliferating cells
many of the oncogenes so far identifed affect cell-signaling pathways
an increase in proliferation alone, without other changes, generates benign growths
can be removed by surgery
further mutations can occur and may eventually lead to malignancy.

34. Cancer can be caused by mutations that improperly inactivate tumor suppressor genes
Function of normal allele of tumor suppressor genes is to control cell proliferation
Mutant tumor suppressor alleles act recessively and cause increased cell proliferation
Tumor suppressor genes identified through genetic analysis of families with inherited predisposition to cancer
Inheritance of a mutant tumor suppressor allele
One normal allele sufficient for normal cell proliferation in heterozygotes
Wild-type allele in somatic cells of heterozygote can be lost or mutated  abnormal cell proliferation

35. The retinoblastoma tumor-suppressor gene
the deletions vary in size and position from patient to patient, they all remove band 13q14
Retinoblastoma provides an example of this identifcation process. A cancer of the color-perceiving cone cells in the retina, retinoblastoma is one of several cancers inherited in a dominant fashion in human families
heterozygous cells in a patient’s normal tissues carry one copy of the gene’s wild-type allele (RB), and this one copy prevents the cells from becoming cancerous. Tumor cells homozygous for the deletion, however, do not carry any copies of RB, and without it, they begin to divide out of control
Fig

36. How can the retinoblastoma trait be inherited in a dominant fashion if a deletion of the RB gene is recessive to the wild-type RB allele?
the strong likelihood that in at least one of the hundreds of thousands of retinal cells heterozygous for the deletion, a subsequent genetic event will disable the single remaining RB allele
both copies of the p16 gene on chromosome 9 are deleted in roughly 75% of all melanomas
in approximately 85% of all gliomas

37. Mutant alleles of these tumor-suppressor genes decrease the accuracy of cell reproduction
Table 17.4
Many tumor-suppressor mutations occur in genes that control the cell cycle and, with it, the accuracy of genomic replication.

38. Alterations in genes that control proliferation result in an enlarged clone of cells, but aside from their increase in number,
these cells—if they sustain no further mutations—are normal and thus form a benign growth
mutations in genes that control the cell cycle can alter the accuracy with which a cell reproduces its genome.
The resulting mutant cells can produce offspring with many more mutations than occur in normal cells
increase the mutational rate as errors accumulate with each round of division.
The resulting mutant cells can produce offspring with many more mutations than occur in normal cells, and this increase in the frequency of mutation vastly increases the probability that the cascade of mutations necessary to produce the phenotypic changes of tumor cells will occur

39. The normal control of cell division
Copyright © The McGraw-Hill Companies, Inc. Permission required to reproduce or display Hartwell et al., 4th edition, Chapter 17
Four phases of the cell cycle: G1, S, G2, and M
A variety of genes and proteins control the events of the cell cycle. These genes and proteins allow progression to the next stage of the cycle when all is well, but they cause the cellular machinery to slow down when damage to the genome or to the machinery itself requires repair.
During M, the cell must coordinate the activities of a variety of proteins: those that cause chromosome condensation,
tubulins that polymerize to form the mitotic spindle on which the chromosomes move, motor proteins in the kinetochores that power chromosome movement, proteins that dissolve and re-form the nuclear membrane at the beginning and end of mitosis, and others.
Fig

40. Experiments with yeast helped identify genes that control cell division
Two kinds of used: Saccharomyces cerevisiae (budding yeast) and Schizosaccharomyces pombe (fission yeast)
Usefulness of yeast for studies of the cell cycle
Both grow as haploids or diploids
Can identify recessive mutations in haploids
Can do complementation analysis in diploids
S. cerevisiae – size of buds serves as a marker of progress through the cell cycle
Daughter cells arise as small buds on mother cell at end of G1 and grow during mitosis
Stage of cell cycle can be determined by relative appearance of buds (see Fig 17.14)

41. The isolation of temperature-sensitive mutants of yeast
Mutants grow normally at permissive temperature (22°)
At restrictive temperature (36°), mutants lose gene function
After replica plating, colonies that grow at 22° but not at 36° have temperature-sensitive mutation
Fig

42. A temperature-sensitive cell-cycle mutant in S. cerevesiae
Cells grown at permissive temperature display buds of all sizes (asynchronous division)
Growth of the same cells at restrictive temperature – all have large buds
Fig a
Fig b

43. The discovery of cyclins and cyclin-dependent kinases through experiments with cell-cycle mutants in yeasts have provided insight into the control of cell division.

44. Some important cell-cycle and DNA repair genes
Table 17.5

45. CDKs interact with cyclins and control the cell cycle by phosphorylating other proteins
Cyclin-dependent kinases (CDKs) – family of kinases that regulate the transition from G1 to S and from G2 to M
Cyclin specifies the protein targets for CDK
Phosphorylation by CDKs can activate or inactive a protein
CDK
cyclin
A CDK combines with a cyclin and acquires the capacity to phosphorylate other proteins. Phosphorylation of a protein can either inactivate or activate it.
Fig a
A CDK combines with a cyclin and acquires the capacity to phosphorylate other proteins.

46. One CDK–cyclin complex activates target proteins required for DNA replication at the onset of the S phase
another CDK–cyclin activates proteins necessary for chromosome condensation and segregation at the beginning of the M phase
After they associate with the appropriate CDKs and point out the proper protein targets

47. CDKs control the dissolution of the nuclear membrane at mitosis
Lamins – provide structural support to the nucleus
Form an insoluble matrix during most of the cell cycle
At mitosis, lamins are phosphorylated by CDKs and become soluble
CDK on the nuclear lamins, a group of proteins that underlie the inner surface of the nuclear membrane (Fig b). The nuclear lamins probably provide structural support for the nucleus and possibly provide sites for the assembly of proteins that function in DNA replication, transcription, RNA transport, and chromosome structure. During most of the cell cycle, the lamins form an insoluble structural matrix. At mitosis, however, the lamins become soluble, and this solubility allows dissolution of the nuclear membrane into vesicles
Fig

48. Mutant yeast permit the cloning of a human CDK gene
Genetic studies of yeast provided much of the evidence that CDK–cyclin complexes are key controlling agents in all eukaryotic cell cycles
Human CDKs and cyclins can function in yeast and replace the corresponding yeast proteins
Fig

49. CDKs mediate the transition from the G1 to the S phase of the cell cycle
CDKs mediate the transition from the G1 to the S phase of the cell cycle. In human cells, CDK4 complexed to cyclinD, and CDK2 complexed to cyclinE, phosphorylate the Rb protein, causing it to dissociate from, and activate, the E2F transcription factor. E2F stimulates transcription of many genes needed for DNA replication. At the transition into S phase, cyclinD is destroyed, cyclinA is synthesized, and the CDK2–cyclinA complex activates DNA replication.
Fig

50. CDK activity in yeast is controlled by phosphorylation and dephosphorylation
Fig
The CDC2 proteincomplexed with cyclinB is inactivated prior to mitosis through phosphorylation by a specific kinase and then activated at the onset of mitosis through dephosphorylation by a specific phosphatase

51. Cell-cycle checkpoints ensure genomic stability
Checkpoints monitor the genome and cell-cycle machinery before allowing progression to the next stage of cell cycle
G1-to-S checkpoint
DNA synthesis can be delayed to allow time for repair of DNA that was damaged during G1
The G2-to-M checkpoint
Mitosis can be delayed to allow time for repair of DNA that was damaged during G2
Spindle checkpoint
Monitors formation of mitotic spindle and engagement of all pairs of sister chromatids

52. The G1-to-S checkpoint is activated by DNA damage
Fig a

53. Disruption of the G1-to-S checkpoint in p53-deficient cells can lead to amplified DNA
Tumor cells often have homogenously staining regions (HSRs) or small, extrachromosomal pieces of DNA (double minutes)
Fig b

54. Disruption of the G1-to-S checkpoint in p53-deficient cells can lead to many types of chromosome rearrangements
Fig c

55. Checkpoints acting at the G2-to-M cell-cycle transition or during M phase
Fig

56. The necessity of checkpoints
Checkpoints are not essential for cell division
Cells with defective checkpoints are viable and divide at normal rates
But, they are much more vulnerable to DNA damage than normal cells
Checkpoints help prevent transmission of three kinds of genomic instability (Fig 17.22)
Chromosome aberrations
Changes in ploidy
Aneuploidy

57. Three classes of error lead to aneuploidy in tumor cells
Fig a

58. Chromosome painting can be used to detect chromosome rearrangements
Chromosomes from normal cells
Chromosomes from tumor cells
Fig