Gene regulation in cancer 11/14/07. Overview The hallmark of cancer is uncontrolled cell proliferation. Oncogenes code for proteins that help to regulate.

Gene regulation in cancer

11/14/07

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).

Different paths leading to transcription factor activation

extra-cellular

cytoplasm

nucleus

Signal• Steroid receptor

TF

Different paths leading to transcription factor activation

extra-cellular

cytoplasm

nucleus

SignalTF

• Steroid receptor

Different paths leading to transcription factor activation

extra-cellular

cytoplasm

nucleus

Signal

*TF

• Steroid receptor

Different paths leading to transcription factor activation

extra-cellular

cytoplasm

nucleus

Kinase

Signal

TF

• Steroid receptor• Nuclear TF

Different paths leading to transcription factor activation

extra-cellular

cytoplasm

nucleus

Kinase

TF

Signal• Steroid receptor• Nuclear TF

Different paths leading to transcription factor activation

extra-cellular

cytoplasm

nucleus

Signal

Kinase

*

TF

• Steroid receptor• Nuclear TF

Different paths leading to transcription factor activation

extra-cellular

cytoplasm

nucleus

Signal

Kinase

*TF

• Steroid receptor• Nuclear TF

Different paths leading to transcription factor activation

extra-cellular

cytoplasm

nucleus

Signal

Kinase

*TF

• Steroid receptor• Nuclear TF

Different paths leading to transcription factor activation

extra-cellular

cytoplasm

nucleus

TF

Signal• Steroid receptor• Nuclear TF• Latent cytoplasmic TF

Different paths leading to transcription factor activation

extra-cellular

cytoplasm

nucleus

TF

Signal• Steroid receptor• Nuclear TF• Latent cytoplasmic TF

Different paths leading to transcription factor activation

extra-cellular

cytoplasm

nucleus

Signal

TF

*

• Steroid receptor• Nuclear TF• Latent cytoplasmic TF

Different paths leading to transcription factor activation

extra-cellular

cytoplasm

nucleus

Signal

TF

*

• Steroid receptor• Nuclear TF• Latent cytoplasmic TF

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.

Example: STAT pathway

Darnell 2000

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.

P53: overview

• Sequence-specific transcription factor (both an activator and a repressor)

Vousden and Lane 2007

(Vogelstein et al. 2000)

p53 pathway

Oren 2003

Oren 2003

Different biological outcomes

p53 activation

• Nuclear protein

• Activated by phosphorylation.

• Contain multiple phosphorylation sites.

Different activation of different subsets of genes.

Oren 2003

Life and death choices of p53

• How do p53 choose which set of genes to activate?

Life and death choices of p53

• How do p53 choose which set of genes to activate?– Different modifications– Different partners– Others?

What genes are regulated by p53?

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

Mirza et al. 2003

Mirza et al. 2003

Target genes

• Differentially expressed genes can be due to direct or indirect regulation. How to identify direct targets?

Mirza et al. 2003

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

Identifying p53 targets by CHIP-chip

Cawley et al. 2004

• Affymetrix tiling array chr 21 and 22

• 35 bp resolution on average

Identifying p53 targets by CHIP-chip

Cawley et al. 2004

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

Distribution of TFBS

Cawley et al. 2004

Novel transcript related to TFBS

Cawley et al. 2004

Novel transcript related to TFBS

Cawley et al. 2004

Co-expression between coding and non-coding RNA

Cawley et al. 2004

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

CHIP-PET: A new method for detecting TFBS

(Wei et al. 2006)

(Wei et al. 2006)

coun

ts

Detected 122 novel target genes.7

Motif finding from CHIP-PET data

(Wei et al. 2006)

Expression profile from multiple tumors

• 193 tumors with p53 wild-type

• 58 tumors with p53 mutant

• Measure gene expression for each tumor tissue

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.

Gene expression profile of p53 wild-type vs mutant tumors

(Wei et al. 2006)

Gene expression profile of p53 wild-type vs mutant tumors

(Wei et al. 2006)

Clinical implications

p53 network

(Vogelstein et al. 2000)

Oren 2003

Reading List

• Oren 2003– A review of p53 pathway

• Mirza et al. 2003– Gene expression profile of p53

• Cawley et al. 2004– Map p53 binding sites using high resolution tiling

array• Wei et al. 2006

– Use CHIP-PET to map p53 binding sites, rediscovered p53 motif; linked p53 targets with gene expression profile