Ezh2 requires PHF1 to efficiently catalyze H3 lysine 27 trimethylation in vivo.
The mammalian Polycomblike protein PHF1 was previously shown to interact with the Polycomb group (PcG) protein Ezh2, a histone methyltransferase whose activity is pivotal in sustaining gene repression during development and in adulthood. As Ezh2 is active only when part of the Polycomb Repressive Complexes (PRC2-PRC4), we examined the functional role of its interaction with PHF1. Chromatin immunoprecipitation experiments revealed that PHF1 resides along with Ezh2 at Ezh2-regulated genes such as the HoxA loci and the non-Hox MYT1 and WNT1 genes. Knockdown of PHF1 or of Ezh2 led to up-regulated HoxA gene expression. Interestingly, depletion of PHF1 did correlate with reduced occupancy of Bmi-1, a PRC1 component. As expected, knockdown of Ezh2 led to reduced levels of its catalytic products H3K27me2/H3K27me3. However, reduced levels of PHF1 also led to decreased global levels of H3K27me3. Notably, the levels of H3K27me3 decreased while those of H3K27me2 increased at the up-regulated HoxA loci tested. Consistent with this, the addition of PHF1 specifically stimulated the ability of Ezh2 to catalyze H3K27me3 but not H3K27me1/H3K27me2 in vitro. We conclude that PHF1 modulates the activity of Ezh2 in favor of the repressive H3K27me3 mark. Thus, we propose that PHF1 is a determinant in PcG-mediated gene repression.
Sarma
Kavitha
K
Howard Hughes Medical Institute, NYU School of Medicine-Smilow Research Center, Biochemistry Department, 522 First Avenue, 2nd Floor, Room 211, New York, NY 10016, USA.
Margueron
Raphael
R
Ivanov
Alexey
A
Pirrotta
Vincenzo
V
Reinberg
Danny
D
D002384:Catalysis; D002460:Cell Line; D049109:Cell Proliferation; D048430:Cell Shape; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D000071221:Enhancer of Zeste Homolog 2 Protein; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008239:Lysine / Q000378:metabolism; D008745:Methylation; D063151:Polycomb Repressive Complex 2; D063146:Polycomb-Group Proteins; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000235:genetics; D012097:Repressor Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000235:genetics
D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; C112309:PHF1 protein, human; D063146:Polycomb-Group Proteins; D034741:RNA, Small Interfering; D012097:Repressor Proteins; D014157:Transcription Factors; C498409:EZH2 protein, human; D000071221:Enhancer of Zeste Homolog 2 Protein; D063151:Polycomb Repressive Complex 2; D008239:Lysine