Position-effect variegation (PEV) - Large segments of
eukaryotic genomes are made of repetitive sequences that are
constitutively heterochromatin - Juxtaposition of a gene to the
heterochromatic regions derives PEV. - Spreading heterochromatic
features to a nearby gene in a clonal fashion. - The drosophila
white gene is the first known example, which has been an important
tool for identifying all the machineries involved in the
heterochromatin formation.
Slide 3
Screening of PEV modifiers - The white gene in heterochromatic
region was used as a reporter. - A large-scale mutagenesis
experiments to find either suppressing or enhancing the PEV. -
su(var) -> loss of silencing -> components for repression and
heterochromatin - e(var) -> enhancing of silencing ->
components for activation
Slide 4
P-element based screening - Transposon P element = inverted
element + transposase - co-injection of the P-based reporter with
active transposase into embryos - 1% of the recovered line with PEV
- Visualizing the heterochromatin structure by MNase assays.
Slide 5
Su(var) and E(var) - Nomenclature Su(var)3-9 17 : Suppressor of
Variegation, chromosome 3, 9 th gene, 17 th allele ---- H3K9
methylase 5 known K3K9 methylase in mammals, including Suv39h1,
Suv39h2. - Su(var)2-5 : Heterochromatic protein 1 (HP1) recognizes
H3K9me2 and spreads through interacting Su(var)3-9 - About 150
genes are involved in PEV; chromosomal proteins + histone modifiers
- PcG and TrxG are also identified as Su(var) or E(var)
Slide 6
Immuno-staining: demonstration of heterochromatin components
-HP1 = su(var)2-5 -HKMT for H3K9 = su(var)3-9
Slide 7
Histone Modification on H3K9 - su(var)3-9 dimethylation on H3K9
- not known for mono and tri-methylases - HP1 and SU(VAR)3-9 are
inter-dependent for the formation of heterochromatin
Slide 8
Many pre-steps for the transition between hetero and
euchromatins
Slide 9
Targeting HP1 and SU(VAR)3-9 repetitive sequences location
within pericentromeric, centromeric, and telomeric regions well
conserved through eukaryotes (S. pombe to mammals however, the
actual histone modifications are interpreted in a different manner
between Individual species!!
Slide 10
Paper to discuss Thursday (Sept.25 th ) Ooi, S.K., Qiu, C.,
Bernstein, E., Li, K., Jia, D., Yang, Z., Erdjument-Bromage, H.,
Tempst, P., Lin, S.P., Allis, C.D., Cheng, X., and Bestor, T.H.
(2007). DNMT3L connects unmethylated lysine 4 of histone H3 to de
novo methylation of DNA. Nature 448, 714-717. Ciccone, D.N., Su,
H., Hevi, S., Gay, F., Lei, H., Bakjo, J., Xu, G., Li, E., and
Chen, T. (2009). KDM1B is a histone H3K4 demethylase required to
establish maternal genomic imprints. Nature 461, 415-418.