Supplementary Table and Figures    

PcG complex Drosophila Human RING1A

Ring/Sex Combs Extra (Sce) RING1B CBX2 CBX4 CBX6 CBX7

Polycomb (Pc)

CBX8 PCGF1/NSPC1 PCGF2/MEL18

PCGF3 PCGF4/BMI1

PCGF5

Posterior Sex Combs (Psc)

PCGF6/MBLR PHC1

PRC1

Polyhomeotic (Ph) PHC2 EZH1

Enhancer of Zeste (E(z)) EZH2

Suppressor of Zeste 12 (Suz12) SUZ12 Extra Sex combs (Esc) EED variants

RBAP46

PRC2

Nucleosome remodeling factor 55 (p55) RBAP48 SFMBT1 SFMBT2 L3MBTL2

Scm with Four MBT Domains (Sfmbt)

MBTD1 PhoRC

Pleiohomeotic (Pho) YY1

   

Number of MBT domains Drosophila Human

SCMH1 2 Sex Comb on Midleg (Scm)

SCML2 L3MBTL1 L3MBTL3 3

lethal (3) malignant brain tumor (L(3)mbt)

L3MBTL4 SFMBT1 SFMBT2 L3MBTL2

4 Scm with Four MBT Domains (Sfmbt)

MBTD1

 Table S1. PcG complexes (top) and MBT domain-containing proteins (bottom) in

flies and humans.

nor

mal

ized

den

sity

SFMBT1LSD1CoRESTINPUT

SFMBT1LSD1

CoRESTCTCF

H3K27ac

H3K27me3

POL IIH3K36me3

H3K4me1

H3K4me2

H3K4me3

H3K79me2

H3K9acH4K20me1

Scalechr1:

20 kb hg19149790000 149800000 149810000 149820000 149830000 149840000 149850000 149860000

HIST2H2BFHIST2H2BFHIST2H2BFHIST2H3D

HIST2H4AHIST2H4ABC019846

HIST2H3CHIST2H2AA3

HIST2H2BCHIST2H2AA3

HIST2H3C

BC019846

HIST2H4A

HIST2H4AHIST2H2BE

HIST2H2ACHIST2H2AB

40 -

0 40 -

0 40 -

0 40 -

0

INPUT

SFMBT1

LSD1

CoREST

A

Figure S1

-10 -8 -6 -4 -2 0 2 4 6 8 10

1

2

3

4

Distance from TSS (kbp)

B

C

Figure S1, related to Figure 2, ChIP-seq read density profiles and correlation with

histone PTMs.

A. SFMBT1 (blue), LSD1 (orange), CoREST (violet) and input (grey) ChIP-seq read

distribution on ± 10kb from TSSs. The x axis represents the distance from the TSS (kbp)

and the y axis indicates the normalized read density.

B. High-density map of all the SFMBT1 ERs that are also bound by LSD1 or

CoREST. Each horizontal line represents a separate TSS (± 5kb region), with the

position of the TSS marked by the vertical green midline. The position of ERs for the

indicated protein is represented by horizontal segments.

C. Read density profile for input, SFMBT1, LSD1, and CoREST at the HIST2

cluster. The x axis corresponds to the genomic location and the y axis corresponds to the

normalized ChIP-seq signal density. Location of the histone genes are represented at the

bottom.

 

unsynch

ronized

early S

late SG2/M

G1

synchronized

SFMBT1

LSD1

CoREST

Cyclin E2

p-H3S10

ß-tubulin0 200400

600 8001000PI

unsynchronized

early S

late S

G2/M

G1

−600 −400 −200 0 200 400 600

−20

020

4060

SFMBT1

bp from TSS

Ant

ibod

y −

inpu

t (R

P10

M) G1

S

−600 −400 −200 0 200 400 600

050

100

150

200 LSD1

bp from TSS

Ant

ibod

y −

inpu

t (R

P10

M) G1

S

−600 −400 −200 0 200 400 600

−20

020

4060

CoREST

bp from TSS

Ant

ibod

y −

inpu

t (R

P10

M) G1

S

−600 −400 −200 0 200 400 600

−20

020

4060

IgG

bp from TSS

Ant

ibod

y −

inpu

t (R

P10

M) G1

S

Scalechr5:

1 kb hg19

PWWP2A

6 -

-6 _10 -

-10 _10 -

-10 _6 -

-6 _10 -

-10 _10 -

-10 _

SFMBT1

LSD1

CoREST

SFMBT1

LSD1

CoREST

S phase

G1 phase

Figure S2

A B

C

D

Figure S2, related to Figure 5, Components of the SLC complex are enriched at the

replication-dependent histone loci during G1 phase but not S phase.

 

A. Cell cycle phases of HeLa S3 cells released after double thymidine

synchronization (Figure 5B-D) as confirmed by FACS analysis. The x axis represents the

propidium iodine (PI) reading. The cell population in different cell cycle stage is

represented by a histogram, with its correlated cell cycle phase indicated next to it.

B. SFMBT1, LSD1, and CoREST proteins are not differentially expressed during the

cell cycle. HeLa S3 cells were released after double thymidine synchronization and four

population of cells were collected: cells in early S, late S, G2/M, and G1 phase. Proteins

analyzed are marked on the right side of each panel.

C. Enrichment of SFMBT1, LSD1, CoREST, and IgG at all histone genes within

HIST1 and HIST2 loci. The x axis corresponds to number of base pair from TSS and the y

axis to read per 10 million reads (RP10M) after input subtraction.

D. Read density profile for SFMBT1, LSD1, and CoREST at the PWWP2A promoter

during two cell cycle stages: S and G1. The x axis corresponds to the genomic location

and the y axis to the normalized ChIP-seq signal density of each antibody after

subtraction of the IgG signal.

                             

D10D12D14D16D18

D20D22D24D26six weeksmSFMBT1 mSFMBT2 mLSD1 mCoREST

mRN

A a

bund

ance

(rel

ativ

e to

β-a

cin)

15%

10%

5%

0%

SFMBT1

β-tubulin

D10 D14 D20 MiceMice PND Adult

RNA binding

RNA processing

ncRNA metabolic process

ncRNA processing

mRNA metabolic process

GO Molecular Function

GO Biological Process

1.0E-1 1.0E-5 1.0E-9 1.0E-13 1.0E-17 1.0E-21p-value

Figure S3

A B

C D

SFMBT1

LSD1

538 15965696

Figure S3. The SLC complex in mouse testes.

A. mRNA abundance of SFMBT1, SFMBT2, LSD1, and CoREST in mice testis.

The y axis represents the mRNA abundance (in percentages relative to β-actin). Bars

represent the protein expression levels in mouse testis every other day from postnatal day

10 (D10) to day 26 (D26) mice, and from six-week-old adult mice.

B. Western blot analysis of SFMBT1 protein levels in day 10/14/20 mice and in

adult mice. Equal loading was confirmed by blotting for β-actin.

C. Venn diagram showing the overlap of ERs for SFMBT1 (blue) and LSD1

(orange), as determined by ChIP-seq in pachytene stage spermatocytes.

D. GO molecular function and biological process analysis of the highly enriched

regions targeted by both SFMBT1 and LSD1 using GREAT. The x axis corresponds to

the binomial raw p-value (in logarithmic scale).

                                               

dSfmbt_MBT1

dSfmbt_MBT4dSfmbt_MBT3dSfmbt_MBT2

SFMBT2_MBT1

SFMBT2_MBT4SFMBT2_MBT3SFMBT2_MBT2

SFMBT1_MBT1

SFMBT1_MBT4SFMBT1_MBT3SFMBT1_MBT2

L3MBTL1_MBT1

L3MBTL1_MBT3L3MBTL1_MBT2

dSfmbt_MBT1

dSfmbt_MBT4dSfmbt_MBT3dSfmbt_MBT2

SFMBT2_MBT1

SFMBT2_MBT4SFMBT2_MBT3SFMBT2_MBT2

SFMBT1_MBT1

SFMBT1_MBT4SFMBT1_MBT3SFMBT1_MBT2

L3MBTL1_MBT1

L3MBTL1_MBT3L3MBTL1_MBT2

Figure S4

Figure S4, related to discussion and Figure 3. Amino acid sequence alignment of

MBT domains from dSfmbt, human L3MBTL1, SFMBT1, and SFMBT2.

The predicted secondary structure is represented at the bottom. Residues important for

binding, as determined byseveral structural studies, are indicated with red stars. Three

residues aligning the aromatic cage located at the loop region between β3 and β4 (as

identified in the fourth MBT domain of dSfmbt and the second MBT domain of

L3MBTL1) are highlighted in yellow. The four residues in the MBT domains of

SFMBT1 that are mutated in this study are highlighted in blue.