Isolation and characterization of proteins associated with histone H3 tails in vivo.
The histone H3 amino-terminal tails play an important role in regulating chromatin transcription. Although the mechanisms by which the H3 tail modulates transcription are not well understood, recent discoveries of specific interactions of regulatory factors with H3 tails suggest that H3 tails are a key player in the precise regulation of transcription activity. To investigate the recruitment-based action of H3 tails in chromatin transcription, we purified H3 tail-associated proteins from HeLa cells that stably express epitope-tagged H3 tails. This approach resulted in the identification of multiple histone methyltransferase activities and transcription regulatory factors that are specifically associated with expressed H3 tail domains. Point mutations of Lys-9 and Lys-27 to block cellular modifications of the tail domains completely abolished the association of specific factors, including HP1 and several repressors. Importantly, our transcription analysis revealed that the purified factors can significantly stimulate p300-mediated transcription from chromatin templates. These results implicate that the H3 tail, when accessible in relaxed chromatin, acts as a transcriptional regulator by mediating recruitment of specific sets of cofactors.
Heo
Kyu
K
Department of Biochemistry and Molecular Biology, University of Southern California/Norris Comprehensive Cancer Center, University of Southern California Keck School of Medicine, Los Angeles, California 90033, USA.
Kim
Bong
B
Kim
Kyunghwan
K
Choi
Jongkyu
J
Kim
Hyunjung
H
Zhan
Yuxia
Y
Ranish
Jeffrey A
JA
An
Woojin
W
D019943:Amino Acid Substitution; D002474:Cell-Free System / Q000737:chemistry / Q000378:metabolism; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism*; D050881:E1A-Associated p300 Protein / Q000096:biosynthesis; D006367:HeLa Cells; D000076983:Histone Methyltransferases; D011495:Histone-Lysine N-Methyltransferase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D006657:Histones / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D020125:Mutation, Missense; D011485:Protein Binding; D011496:Protein Methyltransferases; D011499:Protein Processing, Post-Translational / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000235:genetics; D014157:Transcription Factors / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D014158:Transcription, Genetic / Q000502:physiology*
D002843:Chromatin; D006657:Histones; D014157:Transcription Factors; D000076983:Histone Methyltransferases; D011496:Protein Methyltransferases; D011495:Histone-Lysine N-Methyltransferase; D050881:E1A-Associated p300 Protein; C494360:EP300 protein, human