Announcements

1. Announcements 1. Please let me know if you have a final exam scheduling conflict; we can reschedule your final now. 2. Reminder - I will calculate your current grade for you if you are interested. Final exam (20%) can help! Possible adjustment after final…. 3. No labs this week - happy turkey day. 4. Study guide for final (final in 2 weeks) available in class Wednesday 5. Reminder - look over Ch. 23: 1, 3, 6, 8, 10

2. Review of lecture 35 C. elegans chemotaxis behavior and genetics Statistical analysis of chemotaxis data - 7 t -tests total

3. So, how exactly will you discriminate among unknown possibilities? osm-1 encodes novel protein osm-3 encodes kinesin che-3 encodes a dynein che-3;dpy-13 is short and fat wt is not short and fat and should behave like wt control! osm-1, osm-3, che-3, che-3;dpy-13, wild-type If unknown is wt, will you expect a significant difference bet. index of unknown and wt control? Will you be able to distinguish among osm-1, osm-3, and che-3?

4. Overview of lectures 36/37 Genetics of cancer - Ch. 23 I. cell cycle regulation • II. mutant genes confer predisposition to cancer • III. Tumor suppressor genes - normally halt cell cycle • IV. Oncogenes - normally promote cell cycle • V. Translocations • VI. genomic instability

5. Mutations play a central role in cancer • background rate of spontaneous mutation - due to ? • therefore, always baseline rate of cancer • above baseline rate, environmental agents that promote mutation also contribute to cancer = carcinogens • Which mutant genes are most likely to result in cancer? • How many mutations are needed to cause cancer? • How do mutations convert normal cells into malignant tumors? What are the differences between these cells? 1) uncontrolled growth 2) metastasis

6. I. The “cell cycle” Many cells alternate between dividing and “resting” or not dividing Gap 1; metabolic activity and cell growth G0(resting phase) Mitosis DNA replication ex. 1 hour of 16 hour cell cycle Gap 2; metabolic activity and cell growth

7. Three main checkpoints in the cell cycle • 2001 Nobel Prize was awarded to 3 scientists who studied genes that regulate the cell cycle • G1/S Is cell the correct size? Is DNA damaged? • G2/M Is DNA fully replicated? Is DNA damage repaired? • M Have spindle fibers formed? Have they attached to chromosomes correctly? 1. 3. 2.

8. Why are cell cycle checkpoints important? What might result if DNA repair has not finished? Uncontrolled cell division could occur - cancerous cell Example: p53 protein normally targets cells with severe DNA damage to undergo programmed cell death. (this removes them from the population) If the p53 gene is mutated, damaged cells will not be removed and may continue dividing in an uncontrolled manner. Many different types of cancers involve mutations of p53.

9. Checkpoint Control of Cell Cycle G1/S G1 S G2/M G2 Cdk-G1 cyclin M Cdk- Mitotic (B) cyclin (MPF)

10. II. Which mutant genes most likely to cause cancer? - Retinoblastoma • Diagnosis: “Cat’s eye” reflection (leukocoria) in affected eye. • Most common cancer of infants and children (1/20,000 U.S. live births). • Survival > 90% with early diagnosis and treatment. • Individuals at greater risk of developing other cancers.

11. Retinoblastoma: Familial v. Sporadic +/+ RB/+ RB/+ RB/+ RB/+ RB/+ RB/RB “Loss of Heterozygosity” RB/RB Common Rare - Why?

12. III. Retinoblastoma Gene - first ex. of a tumor suppressor gene • A Tumor Suppressor, which normally suppresses unregulated cell growth: must be inactive for cell division; if permanently inactive, no control over division • Discovered as a regulator of growth of neuroblasts in developing retina of the eye. • Inactivation of both copies of the Rb Gene removes a “brake” on growth, leading to increased incidence of retinal cancer. • Since found to be active in all cells.

13. Rb Protein is Inactivated By CDK-Cyclin During G1  S

14. p53 Gene (tumor suppressor)Normal Functions - controls G1/S • The “Last Gatekeeper” gene since malignant state not attained despite the presence of other cancer-causing mutations until p53 is inactivated by mutation. • 2 possible responses to DNA damage: 1) Acts as a Transcription Factor to activate expression of p21, which inhibits CDK/G1 cyclin to halt the cell cycle; then activates DNA repair. 2) Triggers apoptosis (programmed cell death) if damage can’t be repaired.

15. Role of p53 in Cell Cycle Control

16. p53 Mutations • Most commonly mutated gene in cancers (50% of total). • When p53 is mutated, DNA-damaged cells are not arrested in G1 and DNA repair does not take place. This failure to arrest DNA-damaged cells will be repeated in subsequent cell cycles permitting other mutations to accumulate, culminating in neoplastic transformation... tumor formation and cancer.

17. Breast Cancer Tumor Suppressors • A small proportion of breast cancer is heritable. Two genes are associated with predisposition to breast cancer. • BRCA1 on chromosome 17 • BRCA2 on chromosome 13 • Normal function of both is in repair of ds DNA breaks.

18. IV. Oncogenes • Arise from mutation in normal gene called a proto-oncogene; these promote cell division • Dominant mutation: one copy is sufficient to cause cancer. (different than tumor-suppressors) - when switched on permanently, uncontrolled cell division. • First link between viruses and cancer proposed by Francis Peyton Rous in 1910 (Nobel Prize, 1966): cell-free extracts from chicken tumors injected into healthy chickens caused new tumors.

19. Rous Sarcoma Virus (RSV) • Discovered by Harold Varmus and Bishop, 1975-76 (Nobel Prize, 1989). • A transforming retrovirus (TRV): a cancer-causing single-stranded RNA virus that uses reverse transcriptase enzyme to make ssDNA, then ds DNA, which integrates into host DNA. • Note: not all retroviruses are TRV’s, not all oncogenes caused by viruses. • 100’s of oncogenes now known. • Human T-cell leukemia virus (HTLV) is only human TRV known; codes a TF.

20. Southern Blots Probed with viral src Gene Revealed Cellular Origin of Oncogenes Infected chicken #1 Infected chicken #2 Uninfected chicken (Negative Control) v-src c-src Proto-oncogene SURPRISE!

21. Origin of Transforming Retroviruses Capsid protein Reverse Transcriptase Envelope Protein Mutation creates oncogene

22. Ras Proto-oncogene • Mutated in 30% of all cancers. • A “molecular switch” in the signal transduction pathway leading from growth factors to gene expression controlling cell proliferation: GF  receptor   Ras    TF  target genes  growth. • A single amino acid change in Ras protein can cause constant stimulation of the pathway, even in the absence of growth factors.

23. Cancers Usually Result from a Series of Mutations in a Single Cell • Colon Cancer: oncogene Tumor suppressors oncogene

24. Tumor Progression: Evolution at the Cellular Level Benign tumor (polyp in epithelial cells) is confined by basal lamina; then additional mutation occurs. Malignant tumor (carcinoma in epithelial cells) grows very fast, becomes invasive, and metastasizes.

25. Cancer Cells Evade Two “Safety” Mechanisms Built into the Cell Cycle 1. Once p53 is inactivated, cells with DNA damage don’t arrest from G1 and don’t undergo apoptosis. 2. Telomerase enzyme is activated, avoiding the limit to cell divisions imposed by telomere shortening.