X-inactivation in female human embryonic stem cells is in a nonrandom pattern and prone to epigenetic alterations

Shen et al. 10.1073/pnas.0712018105.

Supporting Information

Files in this Data Supplement:

SI Figure 5
SI Figure 6
SI Table 2
SI Table 3
SI Figure 7




Fig. 5. Normal karyotyping and the presence of two X chromosomes in XIST- female hESCs. A and B show two X chromosomes by FISH with X chromosome-specific probe DAZ1 in XIST+ HSF6 and XIST- HSF6 derived neural progenitor cells (NPCs). (C) Normal karyotyping of XIST- HSF6 cells at P149.





Fig. 6. Association of abnormal nuclear morphology of hESCs with the loss XCI markers. (A) HSF6 cells with abnormal nuclear morphology can lose the XCI marker H3K27me3. (B) hESCs with or without H3K27me3 (red) punctate staining are found in the same colony. (C) H9 cells with abnormal nuclear morphology can lose the XCI marker H3K27me3. (D-F) Hoechst dye (blue) for nuclear staining. The arrows point to hESCs positive for punctate H3K27me3 staining, and the arrowheads point to hESCs without punctate H3K27me3 staining.





Fig. 7. Comparison of mitotic index of XIST+ and XIST- HSF6 hESCs. (A) Immunostaining of phosphor-H3-ser10 (green) and H3K27me3 of hESCs (red). (B) Statistical analysis of the difference of percentage of hESCs undergoing mitosis with or without XCI markers (t test, P = 0.678).





Table 2. List of X-linked genes that exhibit DNA methylation changes in promoter/first exon CpG islands in HSF6 in the absence of XCI markers

Gene name

Type

Z score

Probes, no.

Abs log ratio difference

BCOR

PROMOTER

10.180274

134

0.033315

GPC4

PROMOTER

8.817685

29

0.08067

BCORL1

PROMOTER

8.641933

39

0.068445

CXorf15

PROMOTER

8.226892

18

0.075031

RBBP7

PROMOTER

7.003296

14

0.072584

TRPC5

1st exon

6.421868

2

0.091118

UTX

PROMOTER

5.914889

17

0.054812

APLN

PROMOTER

5.847277

12

0.039811

TSC22D3

PROMOTER-2

5.786264

11

0.083968

MGC4825-FLJ25444

DIVERGENT_PROMOTER

4.974233

7

0.088701

AMOT

PROMOTER

4.658881

13

0.046675

hsa-mir-106a

PROMOTER

4.341017

14

0.049007

RP13-360B22.2

PROMOTER

4.277274

8

0.049273

SPET6

PROMOTER

4.209038

19

0.030608

ARX

PROMOTER-3

4.010452

4

0.09512

HCFC1-CXorf12

DIVERGENT_PROMOTER

3.905944

23

0.028566

ARX

PROMOTER-4

3.605972

12

0.051407

CXorf39

PROMOTER

3.416025

7

0.048319

TSC22D3

PROMOTER-1

3.186377

5

0.046768

DOCK11

PROMOTER-2

3.13456

12

0.044552

SYTL4

PROMOTER

3.033793

9

0.028558

CTPS2

PROMOTER

2.986476

10

0.033103

CITED1

PROMOTER

2.876459

22

0.036119

RNF128

PROMOTER

2.868216

9

0.032638

FGF13

PROMOTER

2.868042

9

0.036501

PLS3

PROMOTER

2.732271

8

0.034033

ARSD

PROMOTER

2.368144

12

0.02953

DLG3

First exon

2.312427

2

0.042552

IL13RA1

PROMOTER

2.299099

10

0.025346

FLJ10178

PROMOTER

2.259634

2

0.055944

GABRQ

PROMOTER

2.23286

8

0.036454

DOCK11

PROMOTER-1

2.212205

2

0.076996

MSN

First exon

2.20674

1

0.054918

IL1RAPL2

First exon

2.200505

5

0.04631

UBE1

PROMOTER-1

2.118992

4

0.05154

FLJ30058

PROMOTER

2.030117

7

0.038729

GPR101

PROMOTER-1

1.871797

7

0.033697

CHRDL1

PROMOTER

1.661521

4

0.034332

FLJ31132

PROMOTER

1.638484

5

0.04067

XKRX

PROMOTER

1.566734

5

0.03336

CACNA1F-CCDC22

DIVERGENT_PROMOTER

1.466447

4

0.02602

POLA

PROMOTER

1.454166

7

0.026836

RS1

PROMOTER

1.442102

5

0.025668

AFF2

PROMOTER

1.391821

2

0.027578

EFHC2

PROMOTER

1.344258

3

0.025909

ZCCHC12

PROMOTER

1.295133

4

0.031534

COVA1

PROMOTER

1.215483

4

0.045605

CUL4B

PROMOTER

1.140789

3

0.034281

NONO

PROMOTER

1.139628

8

0.030337

ARX

PROMOTER-2

1.114482

3

0.036424

SLC16A2

First exon

1.077044

5

0.03558

UTP14A

PROMOTER

1.027599

4

0.039731

ARMCX5

First exon

−1.392925

3

0.025802

PRPS1

PROMOTER

−1.398631

2

0.030214

SLC38A5-FTSJ1

DIVERGENT_PROMOTER

−2.856949

9

0.027639

DIAPH2

PROMOTER

−3.059579

9

0.027627

NAP1L3-RP1-32F7.2

DIVERGENT_PROMOTER

−4.085932

2

0.212526

IL1RAPL2

PROMOTER

−4.242745

5

0.065035

HTR2C

PROMOTER-2

−5.65394

10

0.038434

A Z score of >1 means lower methylation level in XIST- HSF6 hESCs (in black), whereas a Z score of less than −1 means higher methylation level in XIST- HSF6 hESCs (highlighted in blue).





Table 3. List of X-linked genes have at least 1.5-fold up-regulation in XIST− HSF6 hESCs from gene expression array

Gene name

Chromosome location

ARMCX1

chrX:100615561-100615620

ASMTL

chrX:1566007-1565964

BC039399

chrX:40354030-40354089

BCOR

chrX:39667584-39667525

BM928197

chrX:76968577-76968636

COL4A5

chrX:107746725-107746784

CR593492

chrX:118675925-118675984

CR601260

chrX:131229086-131229027

CR627122

chrX:131236518-131236459

CTPS2

chrX:16448393-16445479

CXorf6

chrX:149352785-149352844

CXorf15

chrX:16619656-16619715

CXorf39

chrX:103242155-103242214

DOCK11

chrX:117601834-117601893

ENST00000328046

chrX:23761772-23761713

ENST00000333023

chrX:135598936-135598995

FLJ31204

chrX:57398467-57398526

GPC3

chrX:132521288-132456099

HEPH

chrX:65269349-65269408

HNRPH2

chrX:100474831-100474890

IL13RA1

chrX:117710288-117710347

LOC159090

chrX:133631719-133631660

MGC16121

chrX:133404070-133404011

MUM1L1

chrX:105258691-105258750

NAP1L3

chrX:92732701-92732642

PIM2

chrX:48527078-48527019

PCTK1

chrX:46844220-46844279

PLS3

chrX:114707296-114707355

PSMD10

chrX:107134354-107134295

RAB39B

chrX:154053836-154053777

RBBP7

chrX:16622726-16622667

RP13-360B22.2

chrX:109226927-109226986

RPA4

chrX:95946417-95946476

S71486

chrX:32232361-32232302

SAT

chrX:23563123-23563182

SH3KBP1

chrX:19313800-19313741

SLC16A2

chrX:73536535-73536594

SMARCA1

chrX:128306260-128306201

THC2295157

chrX:51380358-51380417

USP11

chrX:46863306-46863458

USP9X

chrX:40848196-40848255

UTX

chrX:44727602-44727661

ZCCHC12

chrX:117742147-117742206

ZNF75

chrX:134145468-134145409

Note that those genes that also exhibited demethylation in CpG islands in XIST- hESCs are highlighted in red.

This Article

  1. Published online before print March 13, 2008, doi: 10.1073/pnas.0712018105 PNAS March 25, 2008 vol. 105 no. 12 4709-4714
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