Regulation of mouse steroidogenesis by WHISTLE and JMJD1C through histone methylation balance.

The dynamic exchange of histone lysine methylation status by histone methyltransferases and demethylases has been previously implicated as an important factor in chromatin structure and transcriptional regulation. Using immunoaffinity TAP analysis, we purified the WHISTLE-interacting protein complexes, which include the heat shock protein HSP90α and the jumonji C-domain harboring the histone demethylase JMJD1C. In this study, we demonstrate that JMJD1C specifically demethylates histone H3K9 mono- and di-methylation, and mediates transcriptional activation. We also provide evidence suggesting that both WHISTLE and JMJD1C performs functions in the development of mouse testes by regulating the expression of the steroidogenesis marker, p450c17, via SF-1-mediated transcription. Furthermore, we demonstrate that WHISTLE is recruited to the p450c17 promoter via SF-1 and represses the transcription of prepubertal stages of steroidogenesis, after which JMJD1C replaces WHISTLE and activates the expression of target genes via SF-1-mediated interactions. Our results demonstrate that the histone methylation balance mediated by HMTase WHISTLE and demethylase JMJD1C perform a transcriptional regulatory function in mouse testis development.

Multiple-myeloma-related WHSC1/MMSET isoform RE-IIBP is a histone methyltransferase with transcriptional repression activity.

Histone methylation is crucial for transcriptional regulation and chromatin remodeling. It has been suggested that the SET domain containing protein RE-IIBP (interleukin-5 [IL-5] response element II binding protein) may perform a function in the carcinogenesis of certain tumor types, including myeloma. However, the pathogenic role of RE-IIBP in these diseases remains to be clearly elucidated. In this study, we have conducted an investigation into the relationship between the histone-methylating activity of RE-IIBP and transcriptional regulation. Here, we report that RE-IIBP is up-regulated in the blood cells of leukemia patients, and we characterized the histone H3 lysine 27 (H3-K27) methyltransferase activity of RE-IIBP. Point mutant analysis revealed that SET domain cysteine 483 and arginine 477 are critical residues for the histone methyltransferase (HMTase) activity of RE-IIBP. RE-IIBP also represses basal transcription via histone deacetylase (HDAC) recruitment, which may be mediated by H3-K27 methylation. In the chromatin immunoprecipitation assays, we showed that RE-IIBP overexpression induces histone H3-K27 methylation, HDAC recruitment, and histone H3 hypoacetylation on the IL-5 promoter and represses expression. Conversely, short hairpin RNA-mediated knockdown of RE-IIBP reduces histone H3-K27 methylation and HDAC occupancy around the IL-5 promoter. These data illustrate the important regulatory role of RE-IIBP in transcriptional regulation, thereby pointing to the important role of HMTase activity in carcinogenesis.

Characterization of a novel WHSC1-associated SET domain protein with H3K4 and H3K27 methyltransferase activity.

Evolutionary conserved SET domains were originally identified in three Drosophila proteins: suppressor of variegation (Su (var) 3-9), enhancer of zeste (E(z)), and the trithorax. Some of the SET-domain containing proteins have been known to elicit methylation of histone lysine residues. Based on a search for SET-domain containing proteins using bioinformatic tools, we identified and subsequently named a novel SET domain as WHISTLE, that has histone methyltransferase (HMTase) activity. To characterize WHISTLE, we performed an HMTase assay, mass spectrometric analysis, lysine specificity, and transfection assays. Mass spectrometric and immunoblot analysis revealed that WHISTLE di-methylates H3K4 and di-, and tri-methylates H3K27 of histones. Overexpression of WHISTLE repressed transcription of the SV40 promoter. Our results suggest that WHISTLE is a novel SET domain containing a protein with specific H3K4 and H3K27 HMTase activity.