Identification of long range regulatory elements of mouse alpha-globin gene cluster by quantitative associated chromatin trap (QACT).
Chromatin from different regions of the genome frequently forms steady associations that play important roles in regulating gene expression. The widely used chromatin conformation capture (3C) assay allows determination of the in vivo structural organization of an active endogenous locus. However, unpredicted chromatin associations within a given genomic locus can not be identified by 3C. Here, we describe a new strategy, quantitative associated chromatin trap (QACT), which incorporates a modified 3C method and a quantitative assay tool, to capture and quantitatively analyzes all possible associated chromatin partners (ACPs) of a given chromatin fragment. Using QACT, we have analyzed the chromatin conformation of the mouse alpha-globin gene cluster and proved the extensive interaction between HS26 and alpha-globin genes. In addition, we have identified a candidate alpha1-globin gene specific silencer 475A8 which shows the differentiation-stage specific DNase I hypersensitivity. Functional analysis suggests that 475A8 may regulate the alpha1-globin gene during terminal differentiation of committed erythroid progenitor cells. ChIP (chromatin immunoprecipitation) and cotransfection assays demonstrate that GATA-1, a hemopoietic specific transcriptional factor, may increase alpha1-globin gene expression by suppressing the function of 475A8 in terminally differentiated erythroid cells.
Di
Li-Jun
LJ
National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100005, P.R. China.
Wang
Li
L
Zhou
Guo-Ling
GL
Wu
Xue-Song
XS
Guo
Zhi-Chen
ZC
Ke
Xi-Song
XS
Liu
De-Pei
DP
Liang
Chih-Chuan
CC
D000107:Acetylation; D000818:Animals; D002460:Cell Line; D002843:Chromatin / Q000235:genetics*; D003850:Deoxyribonuclease I / Q000378:metabolism; D050982:GATA1 Transcription Factor / Q000378:metabolism; D005914:Globins / Q000235:genetics*; D006657:Histones / Q000378:metabolism; D051379:Mice; D005810:Multigene Family / Q000235:genetics*; D011485:Protein Binding; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics*; D013379:Substrate Specificity
D002843:Chromatin; D050982:GATA1 Transcription Factor; C494689:Gata1 protein, mouse; D006657:Histones; D005914:Globins; D003850:Deoxyribonuclease I