Gene regulation in cancer

1. Gene regulation in cancer 11/14/07

2. Overview • The hallmark of cancer is uncontrolled cell proliferation. • Oncogenes code for proteins that help to regulate cell growth and differentiation. A mutation in an oncogene causes uncontrolled cell growth. • Tumor suppressors suppress cell division or promote apoptosis (regulated cell-death).

3. Different paths leading to transcription factor activation Signal • Steroid receptor extra-cellular cytoplasm TF nucleus

4. Different paths leading to transcription factor activation • Steroid receptor extra-cellular cytoplasm Signal TF nucleus

5. TF * Different paths leading to transcription factor activation • Steroid receptor extra-cellular cytoplasm Signal nucleus

6. Different paths leading to transcription factor activation Signal • Steroid receptor • Nuclear TF extra-cellular Kinase cytoplasm TF nucleus

7. Different paths leading to transcription factor activation • Steroid receptor • Nuclear TF extra-cellular Signal Kinase cytoplasm TF nucleus

8. Different paths leading to transcription factor activation • Steroid receptor • Nuclear TF extra-cellular Signal Kinase * cytoplasm TF nucleus

9. Kinase * Different paths leading to transcription factor activation • Steroid receptor • Nuclear TF extra-cellular Signal cytoplasm TF nucleus

10. TF * Different paths leading to transcription factor activation • Steroid receptor • Nuclear TF extra-cellular Signal cytoplasm Kinase nucleus

11. Different paths leading to transcription factor activation Signal • Steroid receptor • Nuclear TF • Latent cytoplasmic TF extra-cellular TF cytoplasm nucleus

12. Different paths leading to transcription factor activation • Steroid receptor • Nuclear TF • Latent cytoplasmic TF extra-cellular Signal TF cytoplasm nucleus

13. TF * Different paths leading to transcription factor activation • Steroid receptor • Nuclear TF • Latent cytoplasmic TF extra-cellular Signal cytoplasm nucleus

14. TF * Different paths leading to transcription factor activation • Steroid receptor • Nuclear TF • Latent cytoplasmic TF extra-cellular Signal cytoplasm nucleus

15. Transcription factors in cancer Oncogenes • Steroid receptors • Estrogen receptors (breast cancer) and androgen receptors (prostate cancer) • Nuclear proteins • JUN • Latent cytoplasmic factors • STAT Tumor suppressors • p53, RB, etc.

16. Example: STAT pathway Darnell 2000

17. P53: overview • Known as the “guardian of the genome” • The first discovered tumor suppressor • Inactivated in most types of tumors. • 10,000 tumor related mutations have been identified from human to clam.

18. P53: overview • Sequence-specific transcription factor (both an activator and a repressor)

19. Vousden and Lane 2007

20. (Vogelstein et al. 2000)

21. p53 pathway Oren 2003

22. Different biological outcomes Oren 2003

23. p53 activation • Nuclear protein • Activated by phosphorylation. • Contain multiple phosphorylation sites.

24. Different activation of different subsets of genes. Oren 2003

25. Life and death choices of p53 How do p53 choose which set of genes to activate?

26. Life and death choices of p53 • How do p53 choose which set of genes to activate? • Different modifications • Different partners • Others?

27. What genes are regulated by p53?

28. Gene expression profiling • Ovarian cancer cell line (p53 is inactivated) • Expression p53 by infection with adenovirus • Label the DNA from the two cell lines differently and hybridize using a 2-color microarray • Measure gene expression by microarray (60,000 cDNAs) at multiple time points. • Monitor whether genes are activated or repressed (fold change > 2.5). Mirza et al. 2003

29. Mirza et al. 2003

30. Mirza et al. 2003

32. Target genes • Differentially expressed genes can be due to direct or indirect regulation. How to identify direct targets? Mirza et al. 2003

33. Target genes • Differentially expressed genes can be due to direct or indirect regulation. How to identify direct targets? • Use known motif information, scan genome for motif sites. These sites are viewed as target genes. • 294 repressed genes contain p53 motif sites; 67 activated genes contain p53 motif sites Mirza et al. 2003

34. Identifying p53 targets by CHIP-chip • Affymetrix tiling array chr 21 and 22 • 35 bp resolution on average Cawley et al. 2004

35. Identifying p53 targets by CHIP-chip Data analysis • Apply Wilcoxon rank-sum test to probes in each sliding window • P-value cutoff at 10-5 • 1600 p53 sites identified. Cawley et al. 2004

36. Cawley et al. 2004

37. Distribution of TFBS Cawley et al. 2004

38. Novel transcript related to TFBS Cawley et al. 2004

39. Novel transcript related to TFBS Cawley et al. 2004

40. Co-expression between coding and non-coding RNA Cawley et al. 2004

41. Can tiling array data be used to obtain a better motif?

42. CHIP-PET: A new method for detecting TFBS (Wei et al. 2006)

43. counts Detected 122 novel target genes.7 (Wei et al. 2006)

44. Motif finding from CHIP-PET data (Wei et al. 2006)

45. Expression profile from multiple tumors • 193 tumors with p53 wild-type • 58 tumors with p53 mutant • Measure gene expression for each tumor tissue

46. Expression profile from multiple tumors • 193 tumors with p53 wild-type • 58 tumors with p53 mutant • Measure gene expression for each tumor tissue Idea: • For p53 target genes, differential expression should be observed.

47. Gene expression profile of p53 wild-type vs mutant tumors (Wei et al. 2006)

48. Gene expression profile of p53 wild-type vs mutant tumors (Wei et al. 2006)

49. Clinical implications

50. p53 network (Vogelstein et al. 2000)