Presentation MolB

8/7/2019 Presentation MolB

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Long Noncoding RNA as Modular

Scaffold of Histone Modification

Complexes

Miao-Chih Tsai, Ohad Manor, Yue

Wan, Nima Mosammaparast, Jordon

K. Wang, Fei Lan, Yang Shi, Eran Segaland Howard Y. Chang

Published Online 8 July 2010

Science 6 August 2010:

Vol. 329 no. 5992 pp. 689-693


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General Hypothesis

lincRNA can coordinate histone modifications by

binding to multiple histone modificationenzymes

This is established using lincRNA HOTAIR


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HOX-HOTAIR

The HOX gene cluster serves as a classic model

of gene regulation during embryonic

development.

The 39 human HOX genes are contained

within four clusters, called HOXAD, on four

independent chromosomes.

HOTAIR is transcribed from HOXC complex and

acts in trans to repress HOXD


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Specific Hypothesis

HOTAIR may coordinately interact with

both PRC2 and LSD1

HOTAIR

Methylation

of H3K27

Demethylation

of H3K4


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Specific Aims

1. To find out the interaction between LSD1 andHOTAIR

2. To find out the binding domains of PRC2 andLSD1

3. To establish the bridging role of HOTAIRbetween PRC2 and LSD1

4. HOTAIR mediated bridging of PRC2 and LSD1enables their coordinate binding to targetgenes on chromatin


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Specific Aim 1

The authors did RNA immuno precipitation ofLSD1 in foreskin fibroblast and HELA cells and

retrieved endogenous HOTAIR RNA


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M Tsai et al. Science 2010;329:689-693


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Effector Arms of REST Complex

REST

Co-RESTLSD1

CDYLG9a


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Negative Control-

biotinylated GFP and

antisense HOTAIR


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Specific Aims


2. To find out the binding domains of PRC2 andLSD1




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Specific Aim 2



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Specific Aims


2. To find out the binding domains of PRC2 and

LSD1




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STRATEGY

Before and after HOTAIR silencing, RIP was done

IP EZH2 IP LSD1

WB LSD1 WB EZH2

siGFP - negative control for RNA silencingIgG - negative control for IP


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Specific Aims


2. To find out the binding domains of PRC2 and

LSD1




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For every protein studied,

siHOTAIR silencing

chIP followed by microarray (ChIP-chip)

Mapping of protein occupancy acrossgenome

The proteins studied were H3K4me2, H3K27me3,LSD1 and SUZ12

In the presence of HOTAIR, it represses HOXDresulting in the decrease of H3K4me2, increase ofH3K27me3, LSD1 and SUZ12


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Correlation studies



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CONCLUSION


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