Mammalian ASH1L Is a Histone Methyltransferase That Occupies the Transcribed Region of Active Genes

Abstract

Histone lysine methylation regulates genomic functions, including gene transcription. Previous reports found various degrees of methylation at H3K4, H3K9, and H4K20 within the transcribed region of active mammalian genes. To identify the enzymes responsible for placing these modifications, we examined ASH1L, the mammalian homolog of the Drosophila melanogaster Trithorax group (TrxG) protein Ash1. Drosophila Ash1 has been reported to methylate H3K4, H3K9, and H4K20 at its target sites. Here we demonstrate that mammalian ASH1L associates with the transcribed region of all active genes examined, including Hox genes. The distribution of ASH1L in transcribed chromatin strongly resembles that of methylated H3K4 but not that of H3K9 or H4K20. Accordingly, the SET domain of ASH1L methylates H3K4 in vitro, and knockdown of ASH1L expression reduced H3K4 trimethylation at HoxA10 in vivo. Notably, prior methylation at H3K9 reduced ASH1L-mediated methylation at H3K4, suggesting cross-regulation among these marks. Drosophila ash1 and trithorax interact genetically, and the mammalian TrxG protein MLL1 and ASH1L display highly similar distributions and substrate specificities. However, by using MLL null cell lines we found that their recruitments occur independently of each other. Collectively, our data suggest that ASH1L occupies most, if not all, active genes and methylates histone H3 in a nonredundant fashion at a subset of genes.

H3K36 Methylation Antagonizes PRC2-mediated H3K27 Methylation

“…5C), indicating that Ash1 is an H3K36-specific dimethylase. Our results are in agreement with one previous report (20) but disagree with three other reports (19,21,22).…”

“…Because the enzymatic activity of Ash1 is highly controversial, we examined the experimental details of the four previous related reports (19 -22) for potential explanations for the discrepancies. In the two studies that reported Ash1 as an H3K4-specific methyltransferase, GST-tagged H3 tail (22) or recombinant histone H3 (21) were used as substrates for determining site specificity. However, those substrates are not the native substrates of Ash1.…”

“…Ash1 was also reported to be a histone methyltransferase specific for H3K36 (20). In addition, two other independent studies reported Ash1 to be a histone H3K4-specific methyltransferase (21,22). Therefore, the exact chromatin modification that Ash1 generates and thereby potentially antagonizes PRC2-mediated H3K27 methylation remains controversial.…”

“…Thus, we decided to turn to the controversial histone methyltransferase Ash1, which is a well known Trithorax group protein that antagonizes Polycomb silencing (15,16). Although several groups reported that Ash1 is specific for H3K4 (19,21,22) or even H3K9 and H4K20 (19), one independent study reported that Ash1 is a H3K36-specific methyltransferase (20). Moreover, the Set domain of Ash1 is much more closely related to H3K36-specific methyltransferases (including HYPB and Mes4) than H3K4-specific methyltransferases such as Set1 or Trithorax (supplemental Fig.…”

The bromodomain interaction module

“…TAF1 has also been shown to bind di-acetylated p53 (K373 ac /K382 ac [85]) resulting in recruitment to the p21 promoter. The last subfamily of human bromodomains (VIII) contains the methyl-transferase ash1 (absent, small, or homeotic)-like (ASH1L) [28], the chromatin remodelling factors SWI/SNF-related matrixassociated actin-dependent regulator of chromatin a2 (SMARCA2) [86] and SWI/SNF-related matrix-associated actin-dependent regulator of chromatin a4 (SMARCA4) [87] as well as the Polybromo 1 (PB1) [31]. The BRD of SMARCA2 has been shown to bind to histone H3 (K9 ac [43], K14 ac [33,43] and K9 ac /K14 ac [43]) as well as histone H4 (K8 ac , K12 ac [43], K16 ac [33] and K5 ac /K8 ac /K12 ac /K16 ac [43]).…”