Double Strand Breaks Can Initiate Gene Silencing and SIRT1-Dependent Onset of DNA Methylation in an Exogenous Promoter CpG Island

1
Double Strand Breaks Can Initiate Gene Silencing
and SIRT1-Dependent Onset of DNA Methylation in
an Exogenous Promoter CpG Island

• Heather M. OHagan, Helai P. Mohammad, Stephen B.
  Baylin

2
ARR Model for DNA Repair
Green et al. EMBO rep. 2002
3
Can the DNA Repair Process Lead to Aberrant Gene
Silencing?

• Tumor suppressor genes are often silenced in
  cancer cells.
• This silencing often occurs through epigenetic
  means such and chromatin modification and DNA
  methylation

Possible candidates for repair-induced silencing
1) SIRT1 - protein/histone deacetylase that can
be part of a PcG complex 2) EZH2 - HMT
responsible for repressive histone marks, also in
PcG complex 3) DNMT1 - involved in maintaining
DNA methylation 4) DNMT3B - involved in de novo
DNA methylation
4
Treatment with tetracycline induces a double
strand break in the inserted E-cad promoter
5
Treatment with tetracycline induces a double
strand break in the inserted E-cad promoter
6
DSB damage and/or repair induces the transient
recruitment of SIRT1, DNMT1, and DNMT3B
7
DSB damage and/or repair induces the transient
recruitment of SIRT1, DNMT1, and DNMT3B
8
Effects of knockdown of SIRT1 by siRNA
9
Effects of knockdown of SIRT1 by siRNA
10
Changes in enrichment of silencing proteins and
chromatin marks with knockdown of SIRT1
11
Changes in enrichment of silencing proteins and
chromatin marks with knockdown of SIRT1
12
Inducing a DSB in a promoter can lead to
silencing and the seeding of methylation
13
Inducing a DSB in a promoter can lead to
silencing and the seeding of methylation
14
Reduction of SIRT1 during DNA damage decreases
the number of silent clones that have methylation
15
Reduction of SIRT1 during DNA damage decreases
the number of silent clones that have methylation
16
Conclusions

• Silencing proteins can be recruited to the site
  near a double strand break
• Prolonged recruitment may lead to seeding and
  spreading of DNA methylation
• Supports a role for DNA damage in the epigenetic
  silencing of genes in tumors.

17
Final questions or thoughts

• Are silencing proteins recruited during gene
  expression in order to rapidly turn genes off?
• Could the same theory apply for genes being
  expressed and not repaired?