Immunoglobulin V regions diversify by gene conversion in DT40 cells

Able to assay of sIgM+status by flow cytometry

V V V V V J C1

V V V V V* J C1

V

V

V*

Chromatin features of pseudogene donors

Histone acetylation

H3.3 variant

10000

1000

100

10

0

0 400 600 800 1000200

10000

1000

100

10

0

0 400 600 800 1000200

Forward scatter Forward scatter

s IG

M

s IG

M

GFP-LacI HIRA-LacI

Effects on diversification

Med

ian

sIg

M

loss

1.0 1.7 11.0 8.4

0

2

4

6

8

10

12

14

GFP-LacI HIRA Cl18 HIRA-LacI VP16-LacI

Long tract GC77%

Short tract GC3%

Point Mutation20%

Insertion0%

Deletion0%

GFP-LacI

71

Long tract GC66%

Short tract GC21%

Point Mutation11%

Insertion2%

Deletion0%

HIRA-LacI

137

VP16-LacI

Long-tract GC45%

Short-tract GC29%

Point Mutation13%

Deletion0%

Insertion13%

31

LacI-HP1Long tract GC

3%Short tract GC

17%

Point Mutation58%

Insertion8%

Deletion14%

36

Long tract GC77%

Short tract GC3%

Point Mutation20%

Insertion0%

Deletion0%

GFP-LacI

71

Long tract GC66%

Short tract GC21%

Point Mutation11%

Insertion2%

Deletion0%

HIRA-LacI

137

VP16-LacI

Long-tract GC45%

Short-tract GC29%

Point Mutation13%

Deletion0%

Insertion13%

31

LacI-HP1Long tract GC

3%Short tract GC

17%

Point Mutation58%

Insertion8%

Deletion14%

36

GFP-LacI-11 HIRA-LacI-19

Effect of HIRA on chromatin

Cell 1

Cell 2

nucleosomedeposition

Cell 1

Cell 2

~150 bp

MNase

VP16-LacI-10GFP-LacI-11

AcH4 IgG AcH4 IgG

V17

Ova

Enrichment: 4.5 12.4

GFP-LacI VP16-LacI

Effect of VP16 on chromatin