Structural and Functional Insights into the Human Borjeson- Forssman- Lehmann Syndrome- associated Protein PHF6*

Background:PHF6 gene is mutated in BFLS and adult acute myeloid and T-cell acute lymphoblastic leukemias. Results: Crystal structure of the second extended PHD domain of PHF6 was solved. Conclusion: PHF6-ePHD2 is a novel structural module and binds dsDNA. Significance: PHF6 may function as a transcriptional repressor using its ePHD domains binding to DNA and recruiting NuRD complex through its NoLS region to regulate gene transcription. The plant homeodomain finger 6 (PHF6) was originally identified as the gene mutated in the X-linked mental retardation disorder Borjeson-Forssman-Lehmann syndrome. Mutations in the PHF6 gene have also been associated with T-cell acute lymphoblastic leukemia and acute myeloid leukemia. Approximately half of the disease-associated mutations are distributed in the second conserved extended plant homeodomain (ePHD2) of PHF6, indicating the functional importance of the ePHD2 domain. Here, we report the high resolution crystal structure of the ePHD2 domain of PHF6, which contains an N-terminal pre-PHD (C2HC zinc finger), a long linker, and an atypical PHD finger. PHF6-ePHD2 appears to fold as a novel integrated structural module. Structural analysis of PHF6-ePHD2 reveals pathological implication of PHF6 gene mutations in Borjeson-Forssman-Lehmann syndrome, T-cell acute lymphoblastic leukemia, and acute myeloid leukemia. The binding experiments show that PHF6-ePHD2 can bind dsDNA but not histones. We also demonstrate PHF6 protein directly interacts with the nucleosome remodeling and deacetylation complex component RBBP4. Via this interaction, PHF6 exerts its transcriptional repression activity. Taken together, these data support the hypothesis that PHF6 may function as a transcriptional repressor using its ePHD domains binding to the promoter region of its repressed gene, and this process was regulated by the nucleosome remodeling and deacetylation complex that was recruited to the genomic target site by NoLS region of PHF6.