DNA Methylation: Basic Mechanisms - Part 4 ppsx

94
B. F. Van
y
ushin
t
h
e maize en
d
osperm, genes
f
o
r
α
-zeins an
d
α
-tu
b
u
l
ins met
h
y
l
ate
d
in sporo
-
ph
y
tic diploid tissues b ecome undermeth

y
lated in the triploid endosperm,
and the demeth
y
lation correlatin
g
with
g
ene expression is often restricted t
o
t
h
e two c
h
romosomes o
f
materna
l
origin (Lun
d
et a
l
. 1995a,
b
). I
n
Ara
b
i
d

op
-
sis
the paternall
y
inherited
MEA
alleles are transcriptionall
y
silent in both
y
oung em
b
ryo an
d
en
d
osperm.
MEA
gene imprinte
d
in t
he
Ara
b
i
d
opsis en-
dosperm enc odes a SET-domain pr otein of the Pol
y

comb
g
roup that re
g
ulates
ce
ll
pro
l
i
f
eration
b
y exerting a gametop
h
ytic materna
l
contro
ld
uring see
d
development.
ddm
1 m
utat
i
o
n
sa
r

eab
l
eto
r
escue
mea
seeds b
y
functionall
y
r
eactivatin
g
paternall
y
inherited MEA alleles durin
g
seed development. Thus
,
t
h
e maintenance o
f
t
h
e genomic imprint at t
he
m
e
a

l
ocus requires zygotic
D
DM1 activit
y
(Vielle-Calzada et al. 1999). Imprintin
g
of th
e
MEA
Pol
y
com
b
gene is co ntro
ll
e
d
in t
h
e
f
ema
l
e gametop
h
yte
b
y an tagonism
b

etween t
he
two DNA-modif
y
in
g
enz
y
mes, MET1 meth
y
ltransferase and DME
g
l
y
cos
y
-
l
ase (Xiao et a
l
. 2003). DME DNA g
l
ycosy
l
ase activates materna
l
MEA
all
e
l

e
expression in the central cell of the female
g
ametoph
y
te, the pro
g
enitor of th
e
endos
p
erm. Maternal mutan
t
dme
or
mea
a
ll
e
l
es
r
esu
l
t
in
seed abo
r
t
i

o
n
.
Mutations that suppres
s
dme
seed abo
r
t
i
o
nh
a
v
ebee
nf
ou
n
dto
r
es
i
de
in
the MET1 meth
y
ltransferase
g
ene
.

MET1
functions u
p
stream of, or at
,
MEA
a
n
d
is require
df
or DNA met
h
y
l
ation o
f
t
h
ree regions in t
he
MEA
p
romoter in
s
eeds (Xiao et al. 2003). Parental imprintin
g
in A
. thaliana
i

nvolves the activit
y
o
f
t
h
eDNA
MET1
g
ene. P
l
ants trans
f
orme
d
wit
h
an antisens
e
MET1
cons
t
ruc
t
h
ave h
y
pometh
y
lated

g
enomes and show alterations in the behaviour of their
g
ametes in crosses with wild-t
y
pe plants. A h
y
bridization barrier between 2
x
A. thaliana
(when used as a seed parent) and 4x A
. arenosa
(
when used as
a
pollen parent) can be o vercome b
y
increasin
g
maternal ploid
y
butrestored b
y
h
ypomet
h
y
l
ation. T
h

us,
h
ypomet
h
y
l
ation restores t
h
e
h
y
b
ri
d
ization
b
arrier
throu
g
h paternalization of endosperm. Manipulation of DNA meth
y
lation ca
n
b
esu
ffi
cient to erect
h
y
b

ri
d
ization
b
arriers, o
ff
ering a potentia
l
mec
h
anis
m
for speciation and a means of controllin
gg
ene flow between species (Bushell
et al. 2003).
Th
e
A
rabid o
p
sis FWA
g
ene displa
y
s imprinted (maternal ori
g
in-specific)
expression associated with heritable hypomethylation of repeats around tran-
s

cription starting sites in en
d
osperm. T
he
F
WA
imprint
d
epen
d
sont
h
e main
-
tenance DNA methyltransferase MET1 and is not established by allele-specifi
c
d
e novo met
h
y
l
ation
b
ut
b
y materna
l
gametop
h
yte-speci

fi
c gene activation
,
which depends on a DNA
g
l
y
cos
y
lase
g
ene, DEMETE
R
(
Kinoshita et al. 2004
).
D
NA met
h
y
l
ation is essentia
lf
or genome management in p
l
ants: It contro
l
s
the activit
y

of transposable elements and introduced DNA se
g
ments and is
r
espo nsible for transgene silencing (Kooter et al. 1999; Kumpatla and Hal
l
1
999; Meyer 1999). Met
h
y
l
ation o
f
t
h
e
fi
rst untrans
l
ate
d
exon an
d
5

-en
d
o
f
DNA Meth

y
lation in Plants
95
th
eintronint
h
e maize u
b
i
q
uitin 1
p
romoter com
pl
ex an
d
con
d
ensation o
f
t
he chromatin in re
g
ions containin
g
trans
g
enes correlate with transcriptional
t
rans

g
ene silencin
g
in barle
y
(Men
g
et al. 2003)
.
T
h
e
h
omozygous
dd
m1 (
f
or
d
ecrease in DNA met
h
y
l
ation) mutation o
f
Arabido
p
sis
r
esults in

g
enomic DNA h
y
pometh
y
lation and the release of si-
l
encing in various genes. W
h
en t
he
dd
m1 mutation was intro
d
uce
d
into a
n
Arabido
p
sis cell line carr
y
in
g
inactivated tobacco retrotransposon Tto1, this
el
ement
b
ecame
h

ypomet
h
y
l
ate
d
an
d
transcriptiona
ll
yan
d
transpositiona
ll
y
active. Therefore, the inactivation of re trotransposons and the silencin
g
o
f
repeated
g
enes have mechanisms in common (Hirochika et al. 2000). A re-
m
ar
k
a
bl
e
f
eature o

f
t
h
e
ddm
1
mutation is t
h
at it in
d
uces
d
eve
l
o
p
menta
l
a
b
nor-
m
alities b
y
causin
g
heritable chan
g
es in other loci. One of th
e

ddm
1-in
duced
a
b
norma
l
ities is cause
db
y insertion o
f
CAC1, an en
d
ogenous CACTA
f
ami
l
y
t
ransposon. This class of Arabido
p
sis
e
lements transposes and increases i
n
copy num
b
er at
h
ig

hf
requencies speci
fi
ca
ll
yint
he
dd
m1
h
ypomet
h
y
l
atio
n
back
g
round. Thus, the
DDM
1
g
ene not onl
y
epi
g
eneticall
y
ensur es proper
g

ene expression, but also stabilizes transposon behaviour, possibl
y
throu
gh
chromatin remodellin
g
or DNA meth
y
lation (Miura et al. 2001). Robertson’
s
m
utator trans
p
osons in the A rabido
p
sis
g
enome are heavil
y
meth
y
lat ed an
d
inactive. T
h
ese e
l
ements
b
ecome

d
emet
h
y
l
ate
d
an
d
active in t
h
ec
h
romatin-
remodellin
g
mutant
ddm1
, which lost the heterochromatic DNA meth
y
latio
n
(
Singer et a
l
. 2001). T
h
us, DNA transposons in p
l
ants are regu

l
ate
db
yc
h
ro
-
m
atin remodellin
g
. Since
g
ene silencin
g
and paramu tation are also re
g
ulated
b
y
DDM1
,theepi
g
enetic silencin
g
is considered to be related to transposo
n
re
g
ulation (Sin
g

er et al. 2001). Plant S1 SINE retroposons mainl
y
inte
g
rate in
hy
pometh
y
lated DNA re
g
ionsandare tar
g
eted b
y
meth
y
lases; meth
y
lationca
n
th
en sprea
df
rom t
h
e SINE into
fl
an
k
ing genomic sequences, creating

d
ista
l
epi
g
enetic modifications. This meth
y
lation spreadin
g
is vectoriall
y
directe
d
upstream or
d
ownstream o
f
t
h
eS1e
l
ement, suggesting t
h
at it cou
ld b
e
f
aci
l
i-

t
ated when a potentiall
yg
ood meth
y
latable sequence is sin
g
le stranded durin
g
D
NA replication, particularly when located on the lagging strand. Replicatio
n
of a short meth
y
lated DNA re
g
ion could th us lead to the de novo meth
y
latio
n
of upstream or downstream adjacent sequences (Arnaud et al. 2000)
.
DNA met
h
y
l
ation in
fl
uences t
h

emo
b
i
l
ity o
f
transposons. T
h
ein
fl
uence
s
eems to be associated, particularly, with different affinity for Ac transposase
b
in
d
ing to
h
o
l
o-,
h
emi- an
d
unmet
h
y
l
ate
d

transposon en
d
s. In petunia ce
ll
s
,
a holometh
y
lated Ds is unable to excise from a nonreplicatin
g
vector, an
d
rep
l
ication restor es excision. A Ds e
l
ement
h
emimet
h
y
l
ate
d
on one DNA
s
trand transposes in the absence of replication, whereas hemimeth
y
lation o
f

t
he complementary strand causes an inhibition of Ds excision. In the activ
e
h
emimet
h
y
l
ate
d
state, t
h
eDsen
d
s
h
ave a
h
ig
hb
in
d
ing a
ffi
nity
f
or t
h
e trans-
9

6B.F.Van
y
ushin
posase, w
h
ereas
b
in
d
ing to inactive en
d
sisstrong
l
yre
d
uce
d
(Ros an
d
Kunze
2001). Hi
g
h-frequenc
y
transposition of endo
g
enous CACTA transposons in
A
rabid o
p

sis
C
ACTA elements was detected in
cmt3met
1 double mutants. Sin-
gl
emutantsineit
h
er
m
et
1
or
c
mt
3
w
ere muc
hl
ess e
ff
ective in mo
b
i
l
ization
,
despite si
g
nificant induction of CACTA transcript accumulation. Thus, CG

a
n
d
non-CG met
h
y
l
ation systems re
d
un
d
ant
l
y
f
unction
f
or immo
b
i
l
izatio
n
of transposons (Kato et al. 2003). DNA meth
y
lation in the Tam3 end re
g
ions
in
A

ntirr
h
inu
m
t
en
d
e
d
to suppress t
h
e excision activity, an
d
t
h
e
d
egree o
f
meth
y
lation was dependent on the chromosomal position (Kitamura et al
.
2001).
P
aramutation an
d
mutator
(
Mu

)
trans
p
oson inactivation in maize are
l
inked mechanisticall
y
(Lisch et al. 2002). A muta tion of a
g
ene, modifie
r
o
fp
aramutation 1 (
m
op
1
)
,w
h
ic
hp
revents
p
aramutation at t
h
ree
d
i
ff

erent
l
oci in maize, can reverse meth
y
lation of mutator elements. In mo
p1
muta
n
t
b
ac
k
groun
d
s, met
h
y
l
ation o
f
nonautonomou
s
Mu
e
l
ements can
b
e reverse
d
even in the absence of the re

g
ulator
y
MuDR
e
l
e
m
e
n
t.
MuDR
meth
y
lation is
s
e
p
arable fro
m
Mu
D
R
s
ilencin
g
because removal o f meth
y
lation does no
t

cause
imm
ed
i
ate
r
eact
iv
at
i
o
n
.
Th
e
m
o
p
1
m
utation does not alter the meth
y
la
-
tion of certain other transposable elements includin
g
those
j
ust upstream o
f

ap
aramuta
ble
b
1
g
ene. T
h
us, t
h
e mop
1
gene acts on a su
b
set o
f
epigenetica
ll
y
r
e
g
ulated seq uences in the maize
g
enome, and paramutation an
d
Mu
e
l
e

m
e
n
t
met
h
y
l
ation require a common
f
actor (Lisc
h
et a
l
. 2002)
.
Due to
kn
o
wn r
eact
i
o
n
o
f
t
h
eo
xi

dat
iv
e
m
5
Cd
eamination con
j
u
g
ate
d
wit
h
c
y
tosine meth
y
lation (Mazin et al. 1985), DNA meth
y
lation is an essential
muta
g
enic
f
actor t
h
at is responsi
bl
e

f
or a we
ll
-
k
nown p
h
enomeno n o
f
CG an
d
CNG suppressions that are common for man
y
plant
g
enes (Lund et al. 2003).
T
h
us, DNA met
h
y
l
ation is an important
f
actor o
f
p
l
ant ev o
l

ution.
DNA meth
y
lation ma
y
be essentiall
y
modulated b
y
various biolo
g
ical (vi
-
ra
l
,
b
acteria
lf
unga
l
, parasitic p
l
ant in
f
ections) or a
b
iotic
f
actors t

h
at may
in
fl
uence p
l
ant
g
rowt
h
an
dd
eve
l
opment. Interestin
gly
,t
h
eC
h
erno
byl
ra
d
i-
ation accident resulted in a
g
lobal DNA h
y
permeth

y
lation in some plant
s
investi
g
ated (Kovalchuk et al. 2003). Fun
g
al infections most stron
g
l
y
distort
meth
y
lation in repetitive butnot unique sequences in plant
g
enome (Guseinov
an
d
Vanyus
h
in 1975). By t
h
is met
h
o
d
,
f
ungi, viruses an

d
ot
h
er in
f
ective agents
ma
y
switch ov er the
g
ene transcription pro
g
ram in the host plant mostl
y
i
n
f
avour o
f
t
h
e respective in
f
ective agent. On t
h
eot
h
er
h
an

d
,p
l
ants are a
bl
et
o
modif
y
viral DNA that is not inte
g
rated into the plant
g
enome. A few da
ys
a
f
ter inocu
l
ation into turni
pl
eaves, t
h
e unenca
p
si
d
ate
d
cau

l
i
fl
ower mosaic
virus DNA was found to be in a meth
y
lated stat e at almost all HpaII/Msp
I
s
ites (Tan
g
and Leisner 1998). In fact, proper DNA meth
y
lation ma
y
stabi
-
l
ize
f
oreign DNA in
h
ost p
l
ant (Rogers an
d
Rogers 1992). T
h
e
f

oreign DN
A
DNA Meth
y
lation in Plants
97
intro
d
uce
d
into
b
ar
l
ey ce
ll
s was a
bl
e to persist t
h
roug
h
at
l
east two p
l
an
t
g
enerations. Transformation of barle

y
cells was defined b
y
showin
g
initiation
o
f transcription at the proper site on the barle
y
promoter for the chimeric
g
ene in a
l
eurone tissue
f
rom
b
ot
h
a primary trans
f
ormant an
d
its progeny
,
and b
y
tissue-specific expression (aleurone
g
reater than leaf) in the pro

g
en
y.
T
h
is persistence t
h
roug
h
many mu
l
tip
l
es o
f
ce
ll d
ivision is consi
d
ere
d
a
s
f
ormall
y
equivalent to transformation, re
g
ardless of whether the DNA wa
s

c
h
ro mosoma
ll
yintegrate
d
or carrie
d
as an episome,
b
ut
d
oes not necessar-
il
y
rep resent stable inte
g
ration into the
g
enome, since the forei
g
nDNAwa
s
f
requentl
y
rearran
g
ed or lost (Ro
g

ers and Ro
g
ers 1992). The forei
g
nDN
A
w
as most sta
bl
ew
h
en
pl
asmi
d
DNA use
d
in trans
f
ormation
l
ac
k
e
d
a
d
enin
e
m

eth
y
lation but had complete meth
y
lation of c
y
tosine residues in the CG at
Hpa II sites; a
d
enine met
h
y
l
ation a
l
one was associate
d
wit
h
mar
k
e
df
oreig
n
DNA instabilit
y
. Thus, barle
y
cellshaveas

y
stem that identifies DNA lackin
g
th
e proper met
h
y
l
ation pattern an
d
causes its
l
oss
f
rom active
l
y
d
ivi
d
ing c e
ll
s
(Ro
g
ers and Ro
g
ers 1992). These intri
g
uin

g
data on forei
g
n DNA meth
y
latio
n
in plant cells ma
y
resemble a host restriction-modification phenomenon tha
t
is common in prokar
y
otes
.
3
Adenine DNA Methylation
3
.1
N
6
-Meth
y
ladenine in DNA of Eukar
y
ote
s
N
6
-

Meth
y
ladenine (m
6
A
) occurs as aminor base in DNAof various or
g
anisms
.
It was
fi
rst
d
etecte
d
in
E
.co
li
D
NA 50 years ago (Dunn an
d
Smit
h
1955).
Then it was shown to be obvious in most bacterial DNA (Van
y
ushin et al.
1968; Barras an
d

Marinus 1989). It
h
as a
l
so
b
een
f
oun
d
in DNA o
f
a
l
gae
(Pa
kh
omova et a
l
. 1968; Hattman et a
l
. 1978; Ba
b
in
g
er et a
l
. 2001) an
d
t

h
eir
viruses (Que et al. 1997; Nelson et al. 1998), fun
g
i(Bur
y
anov et al. 1970;
R
o
g
ers et a
l
. 1986), an
d
protozoa (Gutierrez et a
l
. 2000) inc
l
u
d
in
g
T
etra
hy
men
a
(Gorovsk
y
et al. 1973; Kirnos et al. 1980; Pratt and Hattman 1981)

,
Crithidia
(Zaitseva et a
l
. 1974)
,
Paramecium (Cummings et a
l
. 1974),
O
xytric
h
a
(
Ra
e
and S
p
ear 1978),
T
rypanosoma cruz
i
(
Ro
j
as and Galanti 1990), and Stylonychia
(Ammermann et a
l
. 1981). In DNA o
f

various a
l
gae,
N
6
-d
imet
h
ya
d
enine wa
s
d
etecte
d(
Pa
kh
omova 1974
)
.A
b
out 0.8% o
f
a
d
enine resi
d
ues are
f
oun

d
a
s
m
6
AinDNAo
f
t
h
e transcriptiona
ll
y active macronuc
l
ei o
f
T
etra
h
ymena
(Gorovs
ky
et a
l
. 1973; Kirnos et a
l
. 1980). A met
hyl
ation site is
5


-
NA T-3

(Bromber
g
et al. 1982), and about 3% meth
y
lation sites are GATC (Harrison
et a
l
. 1986; Karrer an
d
Van Nu
l
an
d
1998)
.
9
8B.F.Van
y
ushin
T
h
ea
d
enine met
h
y
l

ate
d
GATC sites are pre
f
erentia
ll
y
l
ocate
d
in
l
in
k
e
r
D
NA, unmeth
y
lat ed sites are
g
enerall
y
in DNA of nucleosome cores, and
h
istone H1 is not required for the maintenance of normal meth
y
lation pat-
terns (Karrer an
d

Van Nu
l
an
d
2002). It was suggeste
d
t
h
at met
h
y
l
ate
d
sites
ma
y
reflect a distribution of nucleosome positions, onl
y
some of which pro
-
vi
d
e accessi
b
i
l
ity to a
d
enine DNA met

h
y
l
trans
f
erase (Karrer an
d
Van Nu
-
l
and 2002). However, the enz
y
me meth
y
latin
g
adenine residues i
n
T
etrah
y
-
men
a
DNA
h
as not yet
b
een iso
l

ate
d
an
d
its amino aci
d
sequence is un-
kn
o
wn
.
DN A
o
f
t
h
es
lim
e
m
ou
l
d
P
h
y
sarum flavicomum
beco
m
es se

n
s
i
t
iv
e
to t
h
e
D
p
nI restriction endonuclease durin
g
enc
y
stment. This ma
y
be du
e
to t
h
ea
pp
earance o
fm
6
A resi
d
ues in GATC se
q

uences in t
h
is DNA (Z
hu
and Henne
y
1990). Earl
y
data on the presence of m
6
A
in mammalian s
p
erm
D
NA were am
b
iguous (Unger an
d
Venner 1966), an
d
attempts to
d
etect an
d
iso
l
ate t
h
is minor

b
ase
f
rom DNA o
f
man
y
inverte
b
rates an
d
verte
b
rates
were unsuccess
f
u
l
(Vanyus
h
in et a
l
. 1970; Law
l
ey et a
l
. 1972; Fantappie e
t
a
l

. 2001). Nevert
h
e
l
ess, it was
j
u
dg
e
df
rom t
h
e
d
i
ff
erent resistance o
f
ani
-
mal DNA to r estriction endonucleases sensitive to meth
y
lation of adenine
resi
d
ues
(
Ta
q
I,

M
bo
I
an
d
S
au3AI) t
h
at some
g
enes
(
Myo
-
D1) (Ka
y
et a
l
.
1
994)—steroid-5
-
α
-reductase
g
enes 1 and 2 (Re
y
es et al. 1997)—of mam
-
ma

l
s (mouse, rat) mig
h
t con tain
m
6
A
resi
d
ues. T
h
is in
d
irect
l
y suggests t
h
at
anima
l
sma
yh
ave a
d
enine DNA met
hyl
trans
f
erases. It is interestin
g

t
h
at a
d-
d
ition o
f
N
6
-met
h
y
ld
eoxya
d
enosine (Me
d
A
d
o ) to C6.9 g
l
ioma ce
ll
s triggers
a
d
i
ff
erentiation process an
d

t
h
e expression o
f
t
h
eo
l
i
g
o
d
en
d
ro
gl
ia
l
mar
k
er
2

,3

-c
y
clic nucleotide 3

-

phosphor
y
lase. The differentiation induced b
y
N
6
-
met
hyld
eox
y
a
d
enosine was a
l
so o
b
serve
d
on p
h
eoc
h
romoc
y
toma an
d
ter-
atocarcinoma cell lines and on d
y

sembr
y
oplastic neuroepithelial tumou
r
ce
ll
s (Rate
l
et a
l
. 2001). T
h
e precise mec
h
anism
b
yw
h
ic
h
mo
d
i
fi
e
d
nu-
cleoside induces cell differentia tion is still unclear, but it is consider ed to
b
ere

l
ate
d
to ce
ll
cyc
l
emo
d
i
fi
cations. T
h
e incu
b
ation o
f
C2C12 myo
bl
ast
s
in the presence of MedAdo induces m
y
o
g
enesis (Charles et al. 2004). It
i
s
r
e

m
a
rk
ab
l
et
h
at m
6
A was detected b
y
a method based on HPLC cou
-
p
l
e
d
to e
l
ectrospra
y
ionization tan
d
em mass spectrometr
y
in t
h
eDNA
f
ro

m
M
edAdo-treated cells (it remains undetectable in D NA from control cells)
.
F
urt
h
ermore, Me
d
A
d
oregu
l
ates t
h
e expression o
f
p21, myogenin, mTOR an
d
M
HC. Interestin
g
l
y
, in the pluripotent C2C12 cell line, MedAdo drives the
d
i
ff
erentiation towar
d

s myogenesis on
l
y(C
h
ar
l
es et a
l
. 2004). T
h
ese resu
l
ts
point to
N
6
-met
hyld
eox
y
a
d
enosine as a nove
l
in
d
ucer o
f
m
y

o
g
enesis an
d
f
urt
h
er exten
d
st
h
e
d
i
ff
erentiation potentia
l
ities o
f
t
h
is met
h
y
l
ate
d
nuc
l
eo

-
s
i
de.
m
6
A has been f ound in DNA of hi
g
her plants (Van
y
ushin et al. 1971;
Buryanov et a
l
. 1972). It may
b
e present in p
l
asti
d
(amy
l
op
l
ast) DNA (Ngern-
DNA Meth
y
lation in Plants
99
p
rasirtsiri et a

l
. 1988). In w
h
eat see
dl
ings it is present in
h
eavy
(
ρ
= 1.718 g
/
cm
3
)
mitoc
h
on
d
ria
l
DN A (Van
y
us
h
in et a
l
. 1988; A
l
e

k
san
d
rus
hk
ina et a
l
.
1990; Kirnos et al. 1992a, b). Similar mtDNA co ntainin
gm
6
A
w
e
r
ea
l
so
f
ou
n
d
in many ot
h
er
h
ig
h
er p
l

ants inc
l
u
d
ing various arc
h
egoniates (mosses,
f
erns,
and others) and an
g
iosperms (monocots, dicots; Kirnos et al. 1992a). Th
e
synt
h
esis o
f
t
h
is unusua
l
DN A ta
k
es p
l
ace main
l
y in speci
fi
cvacuo

l
ar vesi-
c
l
es containin
g
mitoc
h
on
d
ria, an
d
it is a sort o
f
a
g
in
g
in
d
ex in w
h
eat an
d
o
t
h
er p
l
ants (Kirnos et a

l
. 1992
b
;Ba
k
eeva et a
l
. 1999; Vanyus
h
in et a
l
. 2004)
.
T
h
ereissomein
d
irect evi
d
ence (
b
ase
d
on t
h
ecomparisono
f
pro
d
ucts o

f
DNA h
y
drol
y
sis with restriction endonucleases
Mbo
I
a
n
d
S
a
u
3A) that some
a
d
enine resi
d
ues in zein genes o
f
corn can
b
e met
h
y
l
ate
d
(Pintor-Toro 1987)

.
Th
e
D
R
M2
g
ene
in
Arabido
p
sis
w
as found to be meth
y
lated at both adenin
e
resi
d
ues in some GATC sequences an
d
at t
h
e int erna
l
cytosine resi
d
ues in
C
CGG sites (Ashapkin et al. 2002). Thus, two differen t s

y
stems of the
g
enome
m
o
d
i
fi
cation exist in
h
ig
h
er p
l
ants. It is a
b
so
l
ute
l
yun
k
nown
h
ow t
h
ese sys
-
t

ems ma
y
interact and to what de
g
ree the
y
are interdependent. It appear
s
t
hat adenine meth
y
lation ma
y
influence the c
y
tosine modification and vic
e
versa. In terestin
g
l
y
, the adenine meth
y
lation of th
e
DRM2
g
ene observed i
s
m

ost prominent in wild-t
y
pe plants and appears to be diminished b
y
the
p
resence o
f
antisens
e
METI
transgenes. Since METI
d
oes not possess a
d
enine
D
NA meth
y
ltransferase activit
y
, its action on adenine meth
y
lation is evi-
d
ent
l
y a secon
d
ary e

ff
ect me
d
iate
d
t
h
roug
h
a
d
enine DNA met
h
y
l
trans
f
erase
or some other factors. An
y
wa
y
, we have to keep in mind the idea that ther
e
may exist a new sophisticated type of interdependent regulation of gene func
-
tionin
g
in plants, based on the combinator
y

hierarch
y
of certain chemicall
y
and biolo
g
icall
y
different meth
y
lation s of the
g
enome
.
3
.
2
A
den
i
ne DNA Meth
y
ltransferases
m
6
A is formed in DNA due to the reco
g
nition and meth
y
lation of respec

-
t
ive a
d
enine resi
d
ues in certain sequences
by
speci
fi
ca
d
enine DNA met
hyl-
t
ransferases. Adenine DNA meth
y
ltransferases of b acterial ori
g
in can als
o
m
et
h
y
l
ate cytosine resi
d
ues in DNA wit
h

t
h
e
f
ormation o
fm
4
C(Je
l
tsc
h
2001).
The com
p
arison of
p
rotein structures
p
rovides evidence for an evolution-
ary
l
in
kb
etween wi
d
e
l
y
d
iverge

d
su
bf
ami
l
ies o
fb
acteria
l
DN
A
N
6
-a
d
enin
e
m
et
hyl
trans
f
erases an
d
ar
g
ues a
g
ainst t
h

ec
l
ose
h
omo
l
o
gy
o
f
N
6
-
a
d
enine an
d
N
4
-
cytosine met
h
y
l
trans
f
erases (Bujnic
k
i 1999–2000).
Enz

y
matic DNA met
hyl
ationinpro
k
ar
y
otes an
d
eu
k
ar
y
otes p
l
a
y
sanim-
p
ortant role in the re
g
ulation of man
yg
enetic processes includin
g
transcrip-
t
ion, rep
l
ication, DNA repair an

d
gene transposition (Razin an
d
Riggs 1980).
100
B
.F.Van
y
ushi
n
It is a
l
so an integrative e
l
ement o
fh
ost restriction-mo
d
i
fi
cation system in
bacteria and some lower eukar
y
otes (Arber 1974).
A
denine DNA meth
y
ltransferases of eukar
y
otes could be inherited fro

m
s
ome pro
k
aryotic ancestor. T
h
ey may
b
e
h
omo
l
ogous to
k
nown pro
k
ary
-
otic DNA-(amino)meth
y
ltransferases due to the ver
y
conservative nature of
D
NA met
h
y
l
trans
f

erases in genera
l
. ORFs
f
or putative a
d
enine DNA met
h
y
l-
transferases were found in nuclear but not mitochondrial DNA of protozo
a
(
L
eis
h
mania major),
f
ungi (
S
acc
h
aromyces cerevisiae
,
Sc
h
izosacc
h
aromyce
s

p
ombe), hi
g
her plants(A
. thaliana
)
, and animals
(
Droso
p
hila melano
g
aster,
Caenorhabditis ele
g
ans
,
H
omo sa
p
ien
s
; Shornin
g
and Van
y
ushin 2001)
.
T
h

ere is not
h
ing current
l
y
k
nown a
b
out t
h
e ORF expression
d
etecte
d
or
activit
y
of respective eukar
y
otic proteins encoded in these or
g
anisms. Th
e
enzymatic activity o
f
t
h
ese DNA met
h
y

l
trans
f
erases may
b
every
l
imite
d
as is
true, for example, with the transcription of the Droso
p
hila melano
g
aster
C
5
-
cytosine-DNA met
h
y
l
trans
f
erase gene [t
h
is insect DNA contains an extreme
l
y
l

ow amount o
f
5-met
hyl
c
y
tosine (Gow
h
er et a
l
. 2000), an
d
t
h
e DNA met
hyl
-
transferase
g
ene is a component of a transposon-similar element expresse
d
on
ly
in t
h
e ear
ly
sta
g
es o

f
em
b
r
y
onic
d
eve
l
opment] (L
yk
oeta
l
. 2000)
.
The amino acid sequenc es of putative eukar
y
otic DNA-(amino)meth
y
l-
trans
f
erases (S
h
orning an
d
Vanyus
h
in 2001) are very
h

omo
l
ogous to eac
h
other, as well as to real DNA-(amino)meth
y
ltransferases of eubacteria, h
y
po-
t
h
etica
l
met
h
y
l
trans
f
erases o
f
arc
h
ae
b
acteria an
d
puta tive HemK-proteins o
f
eu

k
ar
y
otes (Bu
j
nic
k
ian
d
Ra
dl
ins
k
a 1999). T
h
ese putative eu
k
ar
y
otic a
d
enin
e
D
NA meth
y
ltransferases (ORF) share conservative motifs (I, IV) specific fo
r
D
NA-(amino)meth

y
ltransferases and motifs II, III, V, VI and X. Motif I (it
takes part in bindin
g
of the methionine part of the
S
-adenos
y
lmethionine
mo
l
ecu
l
ean
d
is speci
fi
c
f
or a
ll
A
d
oMet-
d
epen
d
ent met
h
y

l
trans
f
erases) was
detected in all eukar
y
otic ORFs found. The amino acid composition of the cat-
a
l
ytic centre in a
ll
putative DNA-(amino)met
h
y
l
trans
f
erases is practica
ll
yt
h
e
s
ame; it is extremel
y
conservative and does not have an
y
mutations. It seem
s
that if mutations in the catal

y
tic centre of these enz
y
mes occurred, the
y
eithe
r
would be effectivel
y
repaired or the mutants would be lethal. Motifs V, VI
and X in eukar
y
otic ORFs detected are more similar to analo
g
ous motifs i
n
D
NA-(amino)-met
h
y
l
trans
f
erases
f
rom grou
p
g
. In most ORFs
d

etecte
d,
t
he
conservative motifs specific for DNA-(amino)meth
y
ltransferases occup
y
less
t
h
an
h
a
lf
o
f
t
h
etota
l
amino aci
d
sequence. Six o
f
t
h
ese ORFs
h
ave a re

l
ative
l
y
l
ar
g
e N-terminal part (about 170–200 amino acid residues) located in fron
t
o
f
t
h
e conservative moti
f
s
.
It cannot be ruled out that the
g
ene of the putative DNA-(amino)meth
y
l
-
transferase is located in a block of genes regulating the replication of mi-
toc
h
on
d
ria
l

DNA. In
f
u
ll
y sequence
d
mitoc
h
on
d
ria
l
genomes o
f
eu
k
aryote
s
DNA Meth
y
lation in Plant
s
1
01
(t
h
e
l
iverwor
t

Marc
h
antia po
l
ymorp
ha
,
A
ra
b
i
d
opsis t
h
a
l
ian
a
,sugar
b
eet, t
he
al
g
a Chr
y
sodid
y
mus s
y

nuroideus
)
the nucleotide sequences with si
g
nificant
homolo
gy
to
g
enes of prokar
y
otic DNA-(amino)meth
y
ltransferases were no
t
o
b
serve
d
(S
h
orning an
d
Vanyus
h
in 2001). It is most pro
b
a
bl
et

h
at an enzyme
encoded in the nucleus is trans
p
orted somehow into mitochondria. Putativ e
p
roteins AAF52125 o
f
Drosop
h
i
l
ame
l
anogaster
a
n
d
BAB02202 o
f
A
ra
b
i
d
opsi
s
thaliana
m
i

g
ht have a si
g
nal peptide for mitochondrial transportation o
n
th
e N-en
d
.Ot
h
er ORFs
f
or
h
ypot
h
etica
l
DNA-(amino)met
h
y
l
trans
f
erases o
f
eukar
y
otes do not have distinct si
g

nal peptides on the N-end; but, in fact, thi
s
d
oes not mean that the
y
do no t hav e them. Si
g
nal peptides ma
y
be present o
n
th
e C-en
d
an
dd
i
ff
erent
f
rom
k
nown N-termina
l
signa
l
s may occur (DeLa
b
r
e

et al. 1999).
Th
e
fi
rst eu
k
aryotic (p
l
ant
)
N
6
-a
d
enine DNA met
h
y
l
trans
f
erase
(
wa
d
m
-
t
as
e
)

iso
l
ate
d
was
f
rom t
h
e vacuo
l
ar vesic
l
e
f
raction o
f
a
g
in
g
w
h
eat co
l
eopti
l
e
(Fe
d
oreyeva an

d
Vanyus
h
in 2002). T
h
e vesic
l
es appear in p
l
ant apoptoti
c
ce
ll
s, are enric
h
e
d
wit
h
Ca
2+
an
d
contain active
ly
rep
l
icatin
g
mitoc

h
on
-
d
ria (Bakeeva et al. 1999; Van
y
ushin 2004). In the presence of
S
-
adenos
y
l
-
l
-
met
h
ionine, t
h
e enz
y
me
d
enovomet
hyl
ates t
h
e
fi
rst a

d
enine resi
d
ue in
t
he TG
A
TCA sequence in the sin
g
le-stranded (ss)DNA or dsDNA substrates,
b
ut it pre
f
ers sing
l
e-stran
d
e
d
structures. W
h
eat a
d
enine DNA met
h
y
l
trans
-
f

erase is a M
g
2+
-
or
C
a
2
+
-d
epen
d
ent enz
y
me wit
h
a maximum activit
y
at pH
7
.5–8.0. A
b
out 2–3 mM CaC
l
2
o
r MgC
l
2
i

nt
h
e reaction mixture is nee
d
e
df
or
th
e maxima
l
DNA met
hyl
ation activit
y
.T
h
e enz
y
me is stron
gly
in
h
i
b
ite
dby
eth
y
lenediaminetetraacetate (EDTA). The optimal concentration of AdoMe
t

in DNA met
hyl
ation wit
h
w
a
d
mtas
e
is a
b
out 10
µ
M. Wa
d
mtase
e
nco
d
e
d
in t
he
w
heat nuclear DNA ma
y
be homolo
g
ous to th
e

A
. thaliana
ORF (GenBank
,
B
AB02202.1), w
h
ic
h
mig
h
t
b
e ascri
b
e
d
to putative a
d
enine DNA met
h
y
l
trans-
ferases (Shornin
g
and Van
y
ushin 2001). The meth
y

lated adenine residues
f
oun
d
in Gm
6
ATC site s o
f
a
DRM2
gene inanuc
l
ear DNA o
f
A. t
h
a
l
iana(As
h
a
p-
k
in et a
l
. 2002) cou
ld b
e a constituent part o
f
a sequence TGATCA reco

g
nize
d
and meth
y
lated b
y
wheat adenine DNA meth
y
ltransferase. Unfortunatel
y
,w
e
d
onot
k
now w
h
et
h
er a
d
enine DNA met
hyl
trans
f
erase i
n
A
ra

b
i
d
o
p
sis ce
ll
s
h
a
s
t
he same site specificit
y
as it has in wheat plants
.
S
ince
w
a
d
mtas
e
is
f
oun
d
in vesic
l
es wit

h
mitoc
h
on
d
ria
l
active
l
y-rep
l
icatin
g
DNA, its maximal activit
y
is associated with mtDNA replication and it prefer
s
t
o met
h
y
l
ate ssDNA, t
h
is enzyme seems to operate main
l
ywit
h
rep
l

icatin
g
m
tDNA. Simi
l
ar to t
h
e
k
nown
d
am enz
y
me contro
ll
in
g
p
l
asmi
d
rep
l
ication
in
b
acteria
,
w
a

d
mtas
e
seems to contro
l
re
pl
ication o
f
mtDNA t
h
at are re
p
re-
sente
d
main
ly by
circu
l
ar mo
l
ecu
l
es in w
h
eat see
dl
in
g

s (Kirnos et a
l
. 1992a,
b
)
.
A
s mitochondria could be evolutionaril
y
of bacterial ori
g
in, the bacterial con
-
t
ro
lf
or p
l
asmi
d
rep
l
ication
b
ya
d
enine DNA met
h
y
l

ation seems to
b
e acquire
d
102
B
.F.Van
y
ushi
n
b
yp
l
ant ce
ll
s, an
d
it is pro
b
a
bl
y use
df
or t
h
econtro
l
o
f
mitoc

h
on
d
ria rep
l
ica-
tion.
3
.
3
Putat
i
ve Role of Aden
i
ne DNA Meth
y
lat
i
on
i
n Plants
Un
f
ortunate
ly
,t
h
e
f
unctiona

l
ro
l
eo
f
a
d
enine DNA met
hyl
ation in p
l
ants an
d
ot
h
er
h
ig
h
er eu
k
aryotes is un
k
nown. T
h
ere are some
d
ata avai
l
a

bl
es
h
owin
g
t
h
at t
h
ec
h
aracter o
f
transcription o
f
man
y
p
l
ant
g
enes an
d
t
h
emorp
h
o
l
o

gy
and development of transformed plant cells and the plants are drasticall
y
c
h
ange
d
a
f
ter intro
d
uction into t
h
em o
f
genetic constructs wit
h
expresse
d
g
enes of prokar
y
otic adenine DNA meth
y
ltransferases. For example, in tro-
d
uction an
d
expression o
f

t
h
e
b
acteria
l
a
d
enine DNA met
h
y
l
trans
f
erase
(
d
am
)
g
ene is accompanie
dby
GATC sequence met
hyl
ation in DNA o
f
trans
g
enic
to

b
acco p
l
ants an
d
c
h
anges in t
h
e
l
ea
f
an
d
in
fl
oresc ence morp
h
o
l
ogy. T
h
ee
f
-
fi
cienc
y
o

f
a
d
enine DNA met
hyl
ation was
d
irect
ly
proportiona
l
to expression
le
v
e
l
so
f
t
h
e
dam
construct, and meth
y
lation of all GATC sites was observed
i
na
h
i
ghly

expressin
gl
ine
.
Increasin
g
expression levels of the enz
y
me in different plants correlated
wit
h
increasing
l
ya
b
norma
l
p
h
enotypes a
ff
ecting
l
ea
f
pigmentation, apica
l
dominance and leaf and floral structure
(
van Blokland et al. 1998

)
.More
-
over
,
d
a
m
-met
h
y
l
ation o
f
promoter regions in constructs wit
h
p
l
ant gene
s
for alcohol deh
y
dro
g
enase, ubiquitin and actin r esults in an increase in the
transcription of these
g
enes in tobacco and wheat tissues (Graham and Larki
n
1

995). T
h
is pre
l
iminar
y
met
hyl
ation o
f
promoters is a
l
so important
f
or tran
-
s
cri
p
tion of
P
R
1
a
n
d
PR
2
g
enes in constructs introduced into tobacco proto

-
p
l
asts
b
ye
l
ectroporation (Bro
d
zi
k
an
d
Hennig 1998). A
d
enine met
h
y
l
ation o
f
the AG-motif sequence AGATCCAA in the promoter of NtM
y
b2 (a re
g
ulato
r
o
f
t

h
eto
b
acco retrotransposon Tto1)
b
y
b
acteria
ld
am met
h
y
l
ase en
h
ance
s
a
ctivit
y
of the AG-motif-bindin
g
pr otein (AGP1) in tobacco cells (Su
g
imoto et
a
l. 2003). The presence of meth
y
lated adenine residues in the sequence GAT
C

s
cattered in the reporter plasmid introduced into intact barle
y
aleurone la
y
er
s
by
a particle bombardment increased transcription from hormone-re
g
ulated
α
-
amy
l
ase promoters two- to
fi
ve
f
o
ld
, regar
dl
ess o
f
t
h
e promot er strengt
h,
andproperhormonalre

g
ulation of transcription was maintained (Ro
g
ers an
d
Rogers 1995). T
h
e met
h
y
l
ate
d
a
d
enine e
ff
ect was simi
l
ar w
h
en t
h
e amount o
f
reporter construct DNA used was varied over a 20-fold ran
g
e, be
g
innin

g
with
an amount t
h
at gave on
l
y a sma
ll
increment o
f
expression.
Simi
l
ar transcription-en
h
ancin
g
e
ff
ects
f
or met
hyl
ate
d
a
d
enine resi
d
ues

in DNA were seen with the CaMV 35S, maize Adh1 and maize ubi
q
uitin
p
ro-
moters (Rogers an
d
Rogers 1995). It was s
h
own t
h
at some proteins present in
DNA Meth
y
lation in Plant
s
1
03
wh
eat germ nuc
l
ear extracts
b
oun
d
pre
f
erentia
ll
ytoa

d
enine-met
h
y
l
ate
d
DNA
rather than c
y
tosine-meth
y
lated DNA. It seems that enhanced transcriptio
n
o
f nuclear
g
enes in barle
y
due to the presence of
m
6
A residues in the vicinit
y
of
active promoters may
b
eme
d
iate

db
ym
6
ADNA-
b
in
d
ing protein (Roger
s
and Ro
g
ers 1995).
H
ence, met
h
y
l
ation o
f
a
d
enine resi
d
ues in DNA may contro
l
gene expre s
-
sion in plants. This all means that adenine DNA meth
y
lation in plants i

s
not an inci
d
enta
l
or unexpecte
d
event, an
d
it may p
l
ay a signi
fi
cant p
h
ysio
-
lo
g
ical role. It was h
y
pothesized that modulation of meth
y
lation of adenin
e
residues b
y
incorporation of c
y
tokinins (N

6
-deriva tives of adenine) into DN
A
m
ay serve as a mec
h
anism o
f
p
h
yto
h
ormona
l
regu
l
ation o
f
gene expres-
sion and cellular differentiation in plants (Van
y
ushin 1984). C
y
tokinins (6
-
b
enzy
l
aminopurine) can incorporate into t
h

eDNAo
f
p
l
ants (Ku
d
ryas
h
ov
a
an
d
Van
y
us
h
in 1986) an
d
Tet ra
hy
mena p
y
ri
f
ormi
s
(
Mazin an
d
Van

y
us
h
in
1986). In
f
act, 6-
b
enzy
l
aminopurine in
h
i
b
its p
l
asti
d
DNA met
h
y
l
ation i
n
s
y
camore ce
ll
cu
l

ture an
d
in
d
uces in t
h
ese ce
ll
st
h
e exp ression o
f
enz
y
me
s
i
nvolved in photos
y
nthesis (N
g
ernprasirtsiri and Akazawa 1990). It cannot b
e
r
u
l
e
d
out t
h

at in t
h
is particu
l
ar case, c
y
to
k
inin ma
yb
einvo
l
ve
d
in re
g
u
l
atio
n
o
f adenine DNA meth
y
lation in a plastid
.
T
h
e
d
ata s

h
owing t
h
at a
d
enine DNA met
h
y
l
ation may
b
einvo
l
ve
d
in a con
-
t
ro
lf
or persistence o
ff
orei
g
nDNAinap
l
ant ce
ll
is o
f

specia
l
interest. Un
-
l
i
k
e cytosine met
h
y
l
ation, t
h
ea
d
enine met
h
y
l
ation a
l
one is associate
d
wit
h
m
ar
k
e
df

orei
g
n DNA insta
b
i
l
it
y
(Ro
g
ers an
d
Ro
g
ers 1992). P
l
ant ce
ll
s seem t
o
haveas
y
stem discriminatin
g
between adenine and c
y
tosine DNA modifica
-
t
ions, an

d
t
h
e speci
fi
c enz
y
mes resem
bl
in
g
to some extent
b
acteria
l
restrictio
n
e
ndonucleases could be res
p
onsible for selective elimina tion of im
p
ro
p
riate
ad
enine met
h
y
l

ate
d
DNA. Recent
l
ywe
h
ave iso
l
ate
df
rom w
h
eat see
dl
ing
s
a
few s
p
ecific AdoMet-, C
a
2
+
,M
g
2
+
-de
p
endent endonucleases discriminat-

ing
b
etween met
h
y
l
ate
d
an
d
unmet
h
y
l
ate
d
DNAs (Fe
d
oreyeva an
d
Vanyus
h
in
2
004; B.F. Van
y
ushin, unpublished). This ma
y
also indicate on the presence of
R

-M s
y
stem in hi
g
her plants.
4
C
o
ncl
u
s
io
ns
DNA met
hyl
ation contro
l
sp
l
ant
d
eve
l
opment an
d
is invo
l
ve
d
in

g
ene si-
l
encing an
d
parenta
l
imprinting. It ta
k
es part in contro
lf
or transgenes an
d
f
orei
g
n DNA. Severe distortions in DNA meth
y
lation are accompanied b
y
es
-
sential chan
g
es in plant
g
rowth a nd morpholo
gy
. But unlike animals where
d

mt1
k
noc
k
out resu
l
ts in a
bl
oc
k
o
fd
eve
l
opment an
d
is most
l
y
l
et
h
a
l
,p
l
ant
s
104
B

.F.Van
y
ushi
n
l
ac
k
ing ana
l
ogous enzyme MET1 survive. It seems t
h
at ot
h
er,
l
ess-speci
fic
D
NA meth
y
ltransferases or specific modifications of proteins surroundin
g
the DNA meth
y
lation site ma
y
compensate for the absence of MET1. Plants
h
ave a system o
f

siRNA gene si
l
encing conjugate
d
wit
h
aRNA-
d
irecte
d
DNA
meth
y
lation carried out b
y
enz
y
mes capable of performin
g
CNG and un-
con v en tiona
l
met
h
y
l
ations. T
h
is system is consi
d

ere
d
amec
h
anism
f
or t
he
control of viral infections and even for plant immunit
y
to viral infections, but
t
h
e exact mec
h
anisms o
f
t
h
ese events nee
d
to
b
einvestigate
d
muc
hf
urt
h
er

.
Therei sno doubtthat DNA meth
y
lationisonl
y
an inte
g
ral part of acomple
x
sy
stem in an ensemble of unique structures that control
g
ene activit
y
mostl
y
carrie
d
out in c
h
romatin, w
h
i
l
e
b
eing c
l
ose
l

yinter
d
epen
d
ent on t
h
e
h
iston
e
code. The control of DNA meth
y
lation in a cell ma
y
exist at least at three levels
:
(
1) enzyme(s) activity, (2) CH
3
-
d
onors an
d
(3) avai
l
a
b
i
l
ity o

f
t
h
esu
b
strat
e
D
NA to
b
emo
d
i
fi
e
d
in a
fl
uctuatin
g
c
h
romatin structure.
Some p
l
ant DNA met
h
y
l
trans

f
erases are unique, t
h
ey contain t
h
e conser
-
vative u
b
iquitin association (UBA)
d
omain an
d
seem to
b
econtro
ll
e
d
in a ce
ll
c
y
cle b
y
ubiquitin-mediated protein de
g
radation or (and) the ubiquitiniza-
tion ma
y

a
l
ter t
h
ece
ll
u
l
ar
l
oca
l
ization o
f
t
h
ese enz
y
mes
d
ue to respectiv
e
external si
g
nals, the cell c
y
cle or transposon (or retroviral) activit
y
.
Al

ong wit
h
cytosine met
h
y
l
ation, t
h
e met
h
y
l
ation o
f
a
d
enine in p
l
ant DN
A
was observed and specific adenine DNA meth
y
ltransferase was described. The
s
ame p
l
ant gene ma y
b
e met
h

y
l
ate
d
at
b
ot
h
t
h
ea
d
enine an
d
cytosine resi
d
ues.
The functional role of adenine DNA meth
y
lation is still unknown. An
y
wa
y
,
two different s
y
stems of the
g
enome modification based on meth
y

lation o
f
adenines and c
y
tosines exist in hi
g
her plants. It is
y
et unknown how thes
e
sy
stems ma
y
interact and to what de
g
ree the
y
are interdependent. It appear
s
t
h
at a
d
enine met
h
y
l
ation ma y in
fl
uence cytosine mo

d
i
fi
cation an
d
vice versa,
and mutual control for these
g
enome modifications ma
y
be a part of th
e
epigenetic contro
l
o
f
gene activity in p
l
ants
.
The specific endonucleases discriminatin
g
between DNA meth
y
lated and
unmethylated at adenine and cytosine residues seem to be present in plants
.
It means that plants ma
y
have a restriction-modification s

y
stem
.
Further investigation of chromatin and the interaction of DNA-modifyin
g
enzymes wit
h
various
f
actors or proteins, inc
l
u
d
ing
h
ormone-receptor co m-
p
lexes, is a most im
p
ortant task towards the resolution of the
p
roblem of time
,
p
l
ace an
d
ro
l
eo

f
DNA met
h
y
l
ations in a p
l
ant ce
ll.
A
c
k
now
l
e
d
gement
s
T
h
is wor
k
was supporte
d
in part
b
yt
h
e Russian Foun
d

ation
f
or
B
asic Researc
h
(grant No. 02-04-48005) an
d
Ministry o
f
In
d
ustry, Science an
d
Tec
h
-
n
o
l
ogies (grant No. NSH-1019.2003.4.).
DNA Meth
y
lation in Plant
s
1
05
Re
f
erences

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y
ashova IB, Kirnos MD, Van
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ushin BF (1990) S
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h
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asmi
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l
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ria
l
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d
its a
d
enine met

h
y
l
ation
in co
l
eopti
l
ean
d
initia
ll
ea
f
o
f
w
h
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dl
ings: in
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uence o
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p
h
yto
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b
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h
eci
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h
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h
i
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awa I (2001) Surveying CpG met
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l
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k
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rus
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yu
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