Histone H2BK123 monoubiquitination is the critical determinant for H3K4 and H3K79 trimethylation by COMPASS and Dot1

Histone H2B monoubiquitination by Rad6/Bre1 is required for the trimethylation of both histone H3K4 and H3K79 by COMPASS and Dot1 methyltransferases, respectively. The dependency of methylation at H3K4 and H3K79 on the monoubiquitination of H2BK123 was recently challenged, and extragenic mutations in the strain background used for previous studies or epitope-tagged proteins were suggested to be the sources of this discrepancy. In this study, we show that H3K4 and H3K79 methylation is solely dependent on H2B monoubiquitination regardless of any additional alteration to the H2B sequence or genome. Furthermore, we report that Y131, one of the yeast histone H2A/H2B shuffle strains widely used for the last decade in the field of chromatin and transcription biology, carries a wild-type copy of each of the HTA2 and HTB2 genes under the GAL1/10 promoter on chromosome II. Therefore, we generated the entire histone H2A and H2B alanine-scanning mutant strains in another background, which does not express wild-type histones

ダイチョウキン ニ オケル ジスルフィド ケツゴウ ノ ソウセイ ニ カカワル キノン キョウヤク コウソ DsbB ノ セイカガクテキ カイセキ

京都大学0048新制・課程博士博士(理学)甲第12840号理博第3150号新制||理||1467(附属図書館)UT51-2007-H113京都大学大学院理学研究科化学専攻(主査)教授 伊藤 維昭, 教授 三木 邦夫, 教授 杉山 弘学位規則第4条第1項該当Doctor of ScienceKyoto UniversityDA

Data from: Set1/COMPASS and Mediator are repurposed to promote epigenetic transcriptional memory

In yeast and humans, previous experiences can lead to epigenetic transcriptional memory: repressed genes that exhibit mitotically heritable changes in chromatin structure and promoter recruitment of poised RNA polymerase II preinitiation complex (RNAPII PIC), which enhances future reactivation. Here, we show that INO1 memory in yeast is initiated by binding of the Sfl1 transcription factor to the cis-acting Memory Recruitment Sequence, targeting INO1 to the nuclear periphery. Memory requires a remodeled form of the Set1/COMPASS methyltransferase lacking Spp1, which dimethylates histone H3 lysine 4 (H3K4me2). H3K4me2 recruits the SET3C complex, which plays an essential role in maintaining this mark. Finally, while active INO1 is associated with Cdk8- Mediator, during memory, Cdk8+ Mediator recruits poised RNAPII PIC lacking the Kin28 CTD kinase. Aspects of this mechanism are generalizable to yeast and conserved in human cells. Thus, COMPASS and Mediator are repurposed to promote epigenetic transcriptional poising by a highly conserved mechanism

Regulation of H3K4 Trimethylation via Cps40 (Spp1) of COMPASS Is Monoubiquitination Independent: Implication for a Phe/Tyr Switch by the Catalytic Domain of Set1▿

The multiprotein complex Set1/COMPASS is the founding member of the histone H3 lysine 4 (H3K4) methyltransferases, whose human homologs include the MLL and hSet1 complexes. COMPASS can mono-, di-, and trimethylate H3K4, but transitioning to di- and trimethylation requires prior H2B monoubiquitination followed by recruitment of the Cps35 (Swd2) subunit of COMPASS. Another subunit, Cps40 (Spp1), interacts directly with Set1 and is only required for transitioning to trimethylation. To investigate how the Set1 and COMPASS subunits establish the methylation states of H3K4, we generated a homology model of the catalytic domain of Saccharomyces cerevisiae yeast Set1 and identified several key residues within the Set1 catalytic pocket that are capable of regulating COMPASS's activity. We show that Tyr1052, a putative Phe/Tyr switch of Set1, plays an essential role in the regulation of H3K4 trimethylation by COMPASS and that the mutation to phenylalanine (Y1052F) suppresses the loss of Cps40 in H3K4 trimethylation levels, suggesting that Tyr1052 functions together with Cps40. However, the loss of H2B monoubiquitination is not suppressed by this mutation, while Cps40 is stably assembled in COMPASS on chromatin, demonstrating that Tyr1052- and Cps40-mediated H3K4 trimethylation takes place following and independently of H2B monoubiquitination. Our studies provide a molecular basis for the way in which H3K4 trimethylation is regulated by Tyr1052 and the Cps40 subunit of COMPASS

Linking H3K79 trimethylation to Wnt signaling through a novel Dot1-containing complex (DotCom)

Epigenetic modifications of chromatin play an important role in the regulation of gene expression. KMT4/Dot1 is a conserved histone methyltransferase capable of methylating chromatin on Lys79 of histone H3 (H3K79). Here we report the identification of a multisubunit Dot1 complex (DotCom), which includes several of the mixed lineage leukemia (MLL) partners in leukemia such as ENL, AF9/MLLT3, AF17/MLLT6, and AF10/MLLT10, as well as the known Wnt pathway modifiers TRRAP, Skp1, and β-catenin. We demonstrated that the human DotCom is indeed capable of trimethylating H3K79 and, given the association of β-catenin, Skp1, and TRRAP, we investigated, and found, a role for Dot1 in Wnt/Wingless signaling in an in vivo model system. Knockdown of Dot1 in Drosophila results in decreased expression of a subset of Wingless target genes. Furthermore, the loss of expression for the Drosophila homologs of the Dot1-associated proteins involved in the regulation of H3K79 shows a similar reduction in expression of these Wingless targets. From yeast to human, specific trimethylation of H3K79 by Dot1 requires the monoubiquitination of histone H2B by the Rad6/Bre1 complex. Here, we demonstrate that depletion of Bre1, the E3 ligase required for H2B monoubiquitination, leads specifically to reduced bulk H3K79 trimethylation levels and a reduction in expression of many Wingless targets. Overall, our study describes for the first time the components of DotCom and links the specific regulation of H3K79 trimethylation by Dot1 and its associated factors to the Wnt/Wingless signaling pathway

yohheiBrickner.fastq

yohheiBrickner.fast

Enhancer-associated H3K4 monomethylation by trithorax-related, the drosophila homolog of mammalian MLL3/MLL4

Monomethylation of histone H3 on Lys 4 (H3K4me1) and acetylation of histone H3 on Lys 27 (H3K27ac) are histone modifications that are highly enriched over the body of actively transcribed genes and on enhancers. Although in yeast all H3K4 methylation patterns, including H3K4me1, are implemented by Set1/COMPASS (complex of proteins associated with Set1), there are three classes of COMPASS-like complexes in Drosophila that could carry out H3K4me1 on enhancers: dSet1, Trithorax, and Trithorax-related (Trr). Here, we report that Trr, the Drosophila homolog of the mammalian Mll3/4 COMPASS-like complexes, can function as a major H3K4 monomethyltransferase on enhancers in vivo. Loss of Trr results in a global decrease of H3K4me1 and H3K27ac levels in various tissues. Assays with the cut wing margin enhancer implied a functional role for Trr in enhancer-mediated processes. A genome-wide analysis demonstrated that Trr is required to maintain the H3K4me1 and H3K27ac chromatin signature that resembles the histone modification patterns described for enhancers. Furthermore, studies in the mammalian system suggested a role for the Trr homolog Mll3 in similar processes. Since Trr and mammalian Mll3/4 complexes are distinguished by bearing a unique subunit, the H3K27 demethylase UTX, we propose a model in which the H3K4 monomethyltransferases Trr/Mll3/Mll4 and the H3K27 demethylase UTX cooperate to regulate the transition from inactive/poised to active enhancers

The possible interplanetary transfer of microbes: assessing the viability of Deinococcus spp. under the ISS environmental conditions for performing exposure experiments of microbes in the Tanpopo mission

To investigate the possible interplanetary transfer of life, numerous exposure experiments have been carried out on various microbes in space since the 1960s. In the Tanpopo mission, we have proposed to carry out experiments on capture and space exposure of microbes at the Exposure Facility of the Japanese Experimental Module of the International Space Station (ISS). Microbial candidates for the exposure experiments in space include Deinococcus spp.: Deinococcus radiodurans, D. aerius and D. aetherius. In this paper, we have examined the survivability of Deinococcus spp. under the environmental conditions in ISS in orbit (i.e., long exposure to heavy-ion beams, temperature cycles, vacuum and UV irradiation). A One-year dose of heavy-ion beam irradiation did not affect the viability of Deinococcus spp. within the detection limit. Vacuum (10−1 Pa) also had little effect on the cell viability. Experiments to test the effects of changes in temperature from 80 °C to −80 °C in 90 min (±80 °C/90 min cycle) or from 60 °C to −60 °C in 90 min (±60 °C/90 min cycle) on cell viability revealed that the survival rate decreased severely by the ±80 °C/90 min temperature cycle. Exposure of various thicknesses of deinococcal cell aggregates to UV radiation (172 nm and 254 nm, respectively) revealed that a few hundred micrometer thick aggregate of deinococcal cells would be able to withstand the solar UV radiation on ISS for 1 year. We concluded that aggregated deinococcal cells will survive the yearlong exposure experiments. We propose that microbial cells can aggregate as an ark for the interplanetary transfer of microbes, and we named it ‘massapanspermia’

Resetting the epigenetic balance of Polycomb and COMPASS function at enhancers for cancer therapy

The lysine methyltransferase KMT2C (also known as MLL3), a subunit of the COMPASS complex, implements monomethylation of Lys4 on histone H3 (H3K4) at gene enhancers. KMT2C (hereafter referred to as MLL3) frequently incurs point mutations across a range of human tumor types, but precisely how these lesions alter MLL3 function and contribute to oncogenesis is unclear. Here we report a cancer mutational hotspot in MLL3 within the region encoding its plant homeodomain (PHD) repeats and demonstrate that this domain mediates association of MLL3 with the histone H2A deubiquitinase and tumor suppressor BAP1. Cancer-associated mutations in the sequence encoding the MLL3 PHD repeats disrupt the interaction between MLL3 and BAP1 and correlate with poor patient survival. Cancer cells that had PHD-associated MLL3 mutations or lacked BAP1 showed reduced recruitment of MLL3 and the H3K27 demethylase KDM6A (also known as UTX) to gene enhancers. As a result, inhibition of the H3K27 methyltransferase activity of the Polycomb repressive complex 2 (PRC2) in tumor cells harboring BAP1 or MLL3 mutations restored normal gene expression patterns and impaired cell proliferation in vivo. This study provides mechanistic insight into the oncogenic effects of PHD-associated mutations in MLL3 and suggests that restoration of a balanced state of Polycomb-COMPASS activity may have therapeutic efficacy in tumors that bear mutations in the genes encoding these epigenetic factors