Science - USA (2021-11-12)

enriched in proximity to centromeres, and
these regions are relatively GC-rich compared
with the AT-rich chromosome arms (Fig. 4A).
Gene density drops as the centromeres are ap-
proached, whereas transposon density increases,
until they are replaced byCEN180(Fig. 4A). Gene
and transposon densities are tracked closely by

H3K4me3andH3K9me2ChIP-seqenrichment,

respectively (Fig. 4A). H3K9me2 enrichment is

observed within the centromere, although there

is a reduction in the center coincident with

CENH3 enrichment (Fig. 4A), consistent with

reduced H3 occupancy caused by CENH3 re-

placement. A slight increase in H3K4me3 en-

richment is observed within the centromeres,

relative to the flanking pericentromeres (Fig. 4A).

Using our ONT reads with the DeepSignal-

plant algorithm ( 20 ), we observed dense DNA

methylation across the centromeres in CG,

CHG, and CHH contexts (Fig. 4, A and B). How-

ever, CHG DNA methylation shows relatively

Naishet al.,Science 374 , eabi7489 (2021) 12 November 2021 6of9

Fig. 4. Epigenetic organization
and meiotic recombination within
the centromeres.(A) Quantification
of genomic features plotted along
chromosome arms that were propor-
tionally scaled between telomeres
(TEL) and centromere midpoints
(CEN) [defined by maximum CENH3
ChIP-seq log 2 (ChIP/input) enrich-
ment]. Data analyzed were gene,
transposon, andCEN180density;
CENH3, H3K4me3, H3K9me2,
H2A.W6, H2A.W7, H2A.Z, H3K27me1,
H3K27me3, REC8, and ASY1 log 2
(ChIP/input); and percentage of AT/GC
base composition, DNA methylation,
SPO11-1-oligonucleotides (in wild type
andmet1), and crossovers (table S7).
(B) Plot quantifying crossovers (red),
percentage of CG DNA methylation
(pink), CENH3 (blue), SPO11-1-
oligonucleotides in wild type andmet1,
andCEN180density along centromere

2. (C) An interphase nucleus immu-
   nostained for H3K9me2 (magenta)
   and CENH3-GFP (green) is shown at
   the top. The white line indicates the
   confocal section used for the intensity
   plot shown on the right; the region
   outlined by the white dashed line
   shows a magnified image of a centro-
   mere. The scale bar represents 5mM. At
   the bottom is a male meiocyte (early
   prophase I) immunostained for CENH3
   (red) and V5-DMC1 (green). The region
   outlined by the white line indicates the
   magnified region shown in the lower row
   of images. Scale bars are 10mM (upper)
   and 1mM(lower).(D) Plots of CENH3
   ChIP enrichment (gray), DNA methyla-
   tion in CG (blue), CHG (green) and CHH
   (red) contexts, andCEN180variants
   (purple), averaged over windows
   centered onCEN180starts. The red
   dashed lines show 178-bp increments.
   (E) Metaprofiles of CG-context DNA
   methylation, RNA-seq, and siRNA-seq in
   wild type (green) ormet1(pink and
   purple) ( 29 ) aroundCEN180(n=
   66,131), centromeric intactATHILA
   (n= 53),ATHILAlocated outside the
   centromeres (n= 58),GYPSY(n=
   3979), and random positions (n=
   66,131). Shaded ribbons represent 95%
   confidence intervals for windowed mean values.

D

C

E

B

A

SPO11-1 log2(oligos/gDNA)

Crossovers/10 kbCENH3 log2(ChIP/input)

REC8 ASY1 CENH3 log2(ChIP/input)

SPO11-1 log2(oligos/gDNA)

SPO11-1 log2(oligos/gDNA): Colmet1

CENH3 log2(ChIP/input) kyp suvh5 suvh6

Scaled Coordinates Scaled Coordinates

Coordinates (Mbp)

CEN2

DNA methylation % CG

SPO11-1-oligos log2(oligos/gDNA)

CEN180

/ 10 kb

H2A.W6 H2A.W7H2A.Z

H3K9me2 CENH3 log2(ChIP/input)

H3K27me1 H3K27me3

H3K9me2CENH3 log2(ChIP/input)

Scaled Coordinates

2468

2 468

2468

(^012345) 0 0.5 1 1.5 2 2.5
3
0
10 20 30 40 50
0
2
4
6
012345
-3
-2
-1
0
-1
0
1
2
62012345
63
64
65
35
36
37
38
012345
0
1
-1
-0.5
0
-1
0
1
2
012345
0
0.4
0.8
-1
0
1
2
-2
-1.5
-1
-0.5
(^012345) -0.2
0
0.2
0.4 0.6 0.8
-0.8 -0.6 -0.4
-0.2
0
0.2
012345
-0.8 -0.6 -0.4 -0.2
0
0.2
012345
-0.8 -0.6 -0.4 -0.2
0
0.2
0
0.1
0.2
20
40
60
80
(^01234501234)
-1.0
-0.6
-0.2
0.2
01234
0
20
40
60
01 2 3
4
Crossovers/10 kb
Crossovers/10 kb
Crossovers/10 kb
Colmet1
log2(ChIP/input) CENH3
CEN180
(n=66,131)
Centromeric
ATHILA
(n=53)
Random
(n=66,131)
GYPSY
(n=3,979)
Non-centromeric
ATHILA
(n=58)
Coordinates Coordinates Coordinates Coordinates Coordinates
Genes/10 kbTransposons/10 kb
CEN180/10 kbCENH3 log 2 (ChIP/input)
TEL 0.5 CEN 0.5 TEL
H3K4me3H3K9me2
CENH3 log2(ChIP/input)
%AT %GC
CENH3 log2(ChIP/input)
TEL 0.5 CEN 0.5 TEL
TEL 0.5 CEN 0.5 TEL
TEL 0.5 CEN 0.5 TEL
TEL 0.5 CEN 0.5 TEL
TEL 0.5 CEN 0.5 TEL
TEL 0.5 CEN 0.5 TEL
TEL 0.5 CEN 0.5 TEL
0 0.5 1 1.5
250
500
750
1000 H3K9me2CENH3
Distance (μm)
Intensity
DMC1-V5
H3K9me2
CENH 3
CENH3 Merge
DAPI
0
(^0123450)
20
40
60
80
-1
0
1
2
H3K9me2 CENH3 log2(ChIP/input)
DNA methylation %CG %CHG %CHH
TEL 0.5 CEN 0.5 TEL
6
7
0 10
20 30
0.05 0.15 0.25
-500 0 500 1000
-1000 -500 0 500 1000
-1000 -500 0 500 1000
Coordinates (bp)
CG
CHG
CHH
CEN180
CENH3
% DNA methylation
CEN180
variants
-1000
−1kb Start End +1kb−2kb Start End+2kb −2kbStart End+2kb −2kbStart End+2kb−1kb Start End +1kb
sRNAs (TPM)
RNA−seq (TPM)
CG
DNA
methylation 0
25
50
75
100
0
2
4
6
0
0.5
1
1.5
−1kb Start End +1kb−2kbStart End+2kb −2kbStart End+2kb −2kbStart End+2kb−1kb Start End +1kb
Col Rep1Col Rep2
met1 Rep1 Rep2
Col Rep1Col Rep2
met1 Rep3
Col Rep3
met1 Rep2
Col 21 nt
Col 24 nt
Rep1
21 nt
met1 24 nt
Col 22 ntmet1 22 nt
0
0.5
1
1.5
0
0.5
1
1.5
0
0.5
1
1.5
0
0.5
1
1.5
−1kb Start End +1kb−2kb Start End+2kb −2kbStart End+2kb −2kbStart End+2kb−1kb Start End +1kb
0
25
50
75
100
0
2
4
6
0
25
50
75
100
0
2
4
6
0
25
50
75
100
0
2
4
6
0
25
50
75
100
0
2
4
6
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