Abstract
Yeast disruptor of telomeric silencing-1 (DOT1) is involved in gene silencing and in the pachytene checkpoint during meiotic cell cycle. Here we show that the Dot1 protein possesses intrinsic histone methyltransferase (HMT) activity. When compared with Rmt1, another putative yeast HMT, Dot1 shows very distinct substrate specificity. While Rmt1 methylates histone H4, Dot1 targets histone H3. In contrast to Rmt1, which can only modify free histones, Dot1 activity is specific to nucleosomal substrates. This was also confirmed using native chromatin purified from yeast cells. We also demonstrate that, like its mammalian homolog PRMT1, Rmt1 specifically dimethylates an arginine residue at position 3 of histone H4 N-terminal tail. In surprising contrast, methylation by Dot1 occurs in the globular domain of nucleosomal histone H3. Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) analysis suggests that H3 lysine 79 is trimethylated by Dot1. The intrinsic nucleosomal histone H3 methyltransferase activity of Dot1 is certainly a key aspect of its function in gene silencing at telomeres, most likely by directly modulating chromatin structure and Sir protein localization. In agreement with a role in regulating localization of histone deacetylase complexes like SIR, an increase of bulk histone acetylation is detected in dot1- cells.
Highlights
From the ‡Laval University Cancer Research Center, Hotel-Dieu de Quebec (CHUQ), 9 McMahon Street, Quebec City, Quebec G1R 2J6, Canada and the ¶Eastern Quebec Proteomic Center, CHUL Research Center, 2705 boulevard Laurier, Quebec City, Quebec G1V 4G2, Canada
We show that the transcription silencing regulator Dot1 contains intrinsic histone methyltransferase (HMT) activity that targets the globular domain of nucleosomal histone H3
Dot1 Methylates Nucleosomal Histone H3 but Not Free Histones, While Rmt1 Only Methylates Free Histone H4 —Initially searching for histone methyltransferases that could modulate subsequent acetylation of histone H4 tail by histone acetyltransferase (HAT) in S. cerevisiae, we produced recombinant proteins corresponding to two putative yeast HMTs, Rmt1 and Dot1
Summary
From the ‡Laval University Cancer Research Center, Hotel-Dieu de Quebec (CHUQ), 9 McMahon Street, Quebec City, Quebec G1R 2J6, Canada and the ¶Eastern Quebec Proteomic Center, CHUL Research Center, 2705 boulevard Laurier, Quebec City, Quebec G1V 4G2, Canada. In contrast to Rmt1, which can only modify free histones, Dot1 activity is specific to nucleosomal substrates. We show that the transcription silencing regulator Dot1 contains intrinsic HMT activity that targets the globular domain of nucleosomal histone H3.
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