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EpigeneticsHeritable alterations in chromatin structure can govern gene
expression without altering the DNA sequence.
ViterboUniversità degli Studi della Tuscia
Epigenetics denotes all those hereditary
phenomena in which the phenotype is not only
determined by the genotype (the DNA
sequence itself) but also by the
establishment over the genotype
(in greek “epi” means “over”) of an
imprint that modulates its
functional behavior
Epigenetic phenomena
Genic, chromosome and genomic imprinting
Heterochromatin formation
Centromere function
RNA interference (PTGS)
Paramutation
RIP e MIP (Quelling)
Polycomb group proteins
Transvection
Plants
Vertebrates,
Invertebrates and PlantsEukaryotes
Mammals
Drosophila
Drosophila
Fungi
Eukaryotes
Genic, chromosome or genomic
IMPRINTING
Differentialbehavior of homologous chromosome
s
The chromosome which passes through the male germ line aquires an imprint that results in behaviour exactly opposite to the imprint conferred on the same chromosome by the
female germ line(H. Crouse, 1960)
embryo
x AAx
x xx
maternal genomepaternal genome
zygote
xxAA
xSciara coprofila
embryo
x AA
androgenetic embryos(two male pronuclei)
Poor development of the embryo proper
MM
PP
M P
zygote
gynogenetic embryos(two female pronuclei)
Poor development of extraembryoniccomponents
PM
M
P
Nuclear transplantation in
mammals
Angelman, Prader-Willi syndromes
• Usually caused by large (megabase+) deletions of 15q11-q13
• Delete maternal chromosome = AS
• Delete paternal chromosome = PWS
–Prader-Willi Syndrome - obesity, mental retardation, short stature.
–Angelman Syndrome - uncontrollable laughter, jerky movements, and other motor and mental symptoms.
PWS
AS
PWSMousemodel
ASMousemodel
Imprinting cycle
establishment, maintenance and erasure
What Mendel (fortunately) didn’t find in his experiments with peas
1:1
Does the genomic imprinting falsifies the Mendel’s rules?
Neither the segregation of single gene alleles, nor the indipendent behavior of different genes are affected by the existence of imprinting
What the imprinting may mask are the dominance relations between alleles, and hence only the phenotypic output of a cross
NO
HETEROCHROMATIN
NUCLEATION AND MAINTENANCE
In 1928, Heitz defined the heterochromatin as regions of chromosomes that do not undergo cyclical changes in condensation during cell cycle as the other chromosome regions (euchromatin) do.
Heterochromatin is not only allocyclic but also very poor of active genes, leading to define it as genetically inert (junk DNA).
Heterochromatin can be subdivided into two classes: constitutive heterochromatin and facultative heterochromatin.
Constitutive heterochromatin indicates those chromatin regions that are permanently heterochromatic. These regions occupy fixed sites on the chromosomes of a given species, are present in both homologous chromosomes, throughout the life cycle of the individual.
Facultative heterochromatization is a phenomenon leading to the developmentally or tissue-specific co-ordinate
reversible inactivation of discrete chromosome regions,
entire chromosomes or whole haploid chromosome sets.
Position Effect Variegation (PEV)
inversion
White+ Wm4
Wm4
W-
Wm4
Y
White+
pericentricheterochromatin
Drosophila melanogaster X chromosome W+
W-
W+
Y
In all cases an inversion or translocation changed the position of
the gene from a euchromatic to heterochromatic position
this results in variegation
Some rearrangements gave large patches of red facets adjacent to
large patches of white
Conclusion: Decision on expression of white is made early during
tissue development and maintained through multiple cell divisions
Gene is not mutated – movement of the rearranged allele away
from heterochromatin can restore expression
PEV is not limited to Drosophila: see telomeric silencing in yeast
QuickTime™ e undecompressore TIFF (LZW)
sono necessari per visualizzare quest'immagine.
XY XX XXX
XXXXY XXXXX
The Barr body
X chromosome inactivation
Genotype is Xyellow/Xblack
Yellow patches: black allele is inactive Black patches: yellow allele is inactive
Xyellow/Xblack
Xyellow/Xblack
In mammals the dosage compensation of the X chromosome products, between XX females and XY males is achieved by inactivating one of the two Xs in each cell of a female (Mary Lyon, 1961)
imprinted facultative heterochromatization
Coccid chromosome system
embryo
zygote
maternal chromosomes paternal chromosomes
embryo
Planococcus citri (2n=10)
Female and male cells from P.citri
B-IB’
B’/B-I* B-I
x
x
B’/B-I B-I*/B-I
PARAMUTATIONAlexander Brink
MOLECULAR MECHANISMS
OF EPIGENETICS
The chromatin
DNA
histones
nucleosomes
DNA modifications
Histone protein modifications
HISTONE PROTEIN
MODIFICATIONS
Acetylation Phosforylation Methylation Ubiquitination
H3
H4
H2A
H2B
euchromatin heterochromatin
chromatin
…20KMe
20KMe
20KMe
20KMe
4KMe
4KMe
4KMe
…4KMe…9K
Me
9KMe
9KMe
9KMe
9KMe
…16KAc
16KAc
16KAc16KAc
16KAc
chromatin
HP1 and modified histone tails interactions during heterochromatin formation
euchromatin heterochromatin
9KMe
9KMe
9KMe
non histone chromatin proteins: HP1
Epigenetic modifications leading to gene silencing.
(A) Gene repression through histone methylation. Histone deacetylase deacetylates lysine 9 in H3, which can then be methylated by HMTs. Methylated lysine 9 in H3 is recognised by HP1, resulting in maintenance of gene silencing.
B) Gene repression involving DNA methylation. DNA methyltransferases methylate DNA by converting SAM to SAH, a mechanism that can be inhibited by DNMT inhibitors (DNMTi). MBPs recognise methylated DNA and recruit HDACs, which deacetylate lysines in the histone tails, leading to a repressive state.
(C) Interplay between DNMTs and HMTs results in methylation of DNA and lysine 9 in H3, and consequent local heterochromatin formation. The exact mechanism of this cooperation is still poorly understood.
Histone Code and Transcriptional Silencing
Epigenetic modifications leading to gene activation.
(A) Setting 'ON' marks in histone H3 to activate gene transcription. Lysine 4 in H3 is methylated by HMT (for example MLL) and lysine 9 is acetylated by HAT, allowing genes to be transcribed. It is not known, if HMTs and HATs have a direct connection to each other.
(B) In the postulated 'switch' hypothesis, phosphorylation of serines or threonines adjacent to lysines displaces histone methyl-binding proteins, accomplishing a binding platform for other proteins with different enzymatic activities. For example, phosphorylation of serine 10 in H3 may prevent HP1 from binding to the methyl mark on lysine 9. Other lysines in H3 may be acetylated by HATs, therefore overwriting the repressive lysine 9 methyl mark and allowing activation.
(C) Although there is no HDM identified to date, one can speculate that, if this enzyme exists, serine 10 phosphorylation in H3, for example, by Aurora kinases, can lead to recruitment of HDMs that in turn demethylate lysine 9 in H3. Histone acetyltransferases might then acetylate lysine 9 and HMTs methylate lysine 4, resulting in the loosening of the chromatin structure and allowing gene transcription.
Histone Code and Transcriptional Activation
Histone Modification Cassettes
Methylation of Lys-9 by DIM-5 (SUVAR39H1) recruits HP1 via its chromodomain.
In turn, HP1 can recruit additional SUVAR39H1 and other silencing proteins toestablish heterochromatin.
Phosphorylation of Ser-10 abolishes methylation of Lys9 by DIM-5 (SUVAR39H1) andbinding of the HP1, thereby blocking heterochromatin formation.
Phosphorylation of Ser-10 can modestly stimulate acetylation of Lys14 by GCN5,thus promoting transcription.
Lys-9 and Ser-10 have been referred to as a methyl/phos switch:
Fischle W, Wang Y, Allis CD. Nature. 2003;425:475-9.
DNA MODIFICATIONS
Imprinting cycle/DNA metylation cycleestablishment, maintenance and erasure
somatic cells
maintenance embryonic divisions
Maternal genome Paternal genome
zygote
gametes
gametogenesisreversion
de novo establishment
mm
maintenance methylase
mm
m mmm
m m
demethylasede novo methylase
m m
dapi
m9KH3
HP1
merge
Heterochromatin, HP1 and histone tail modifications
Histone H3 lysine 9 methylation
dapi
m9KH3
HP1
merge
Histone H4 lysine 20 methylation