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Albert Jeltsch - One of the best experts on this subject based on the ideXlab platform.
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The SUV39H1 Protein Lysine Methyltransferase Methylates Chromatin Proteins Involved in Heterochromatin Formation and VDJ Recombination
ACS chemical biology, 2017Co-Authors: Srikanth Kudithipudi, Maren Kirstin Schuhmacher, Adam Fiseha Kebede, Albert JeltschAbstract:SUV39H1 is an H3K9 methyltransferase involved in the formation of heterochromatin. We investigated its substrate specificity profile and show recognition of H3 residues between K4 and G12 with highly specific readout of R8. The specificity profile of SUV39H1 is distinct from its paralog SUV39H2, indicating that they can have different additional substrates. Using the specificity profile, several novel SUV39H1 candidate substrates were identified. We observed methylation of 19 novel substrates at the peptide level and for six of them at the protein level. Methylation of RAG2, SET8, and DOT1L was confirmed in cells, which all have important roles in chromatin regulation. Methylation of SET8 allosterically stimulates its H4K20 monomethylation activity connecting SUV39H1 to the generation of increased H4K20me3 levels, another heterochromatic modification. Methylation of RAG2 alters its subnuclear localization, indicating that SUV39H1 might regulate VDJ recombination. Taken together, our results indicate that ...
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Clr4 specificity and catalytic activity beyond H3K9 methylation.
Biochimie, 2017Co-Authors: Denis Kusevic, Srikanth Kudithipudi, Nahid Iglesias, Danesh Moazed, Albert JeltschAbstract:In fission yeast, the catalytic activity of the protein lysine methyltransferase (PKMT) Clr4, the sole homolog of the mammalian SUV39H1 and SUV39H2 enzymes, majorly contributes to the formation of heterochromatin. The enzyme introduces histone 3 lysine 9 (H3K9) di- and tri-methylation, a central heterochromatic histone modification, and later it was also found to methylate the Mlo3 protein, which has a role in heterochromatin formation as well. Herein, we have investigated the substrate specificity of Clr4 using custom made mutational scanning peptide arrays. Our data show, that Clr4 recognizes an RK core motif, showing high preference for R8. In addition, it exhibits specific contacts at the S10, T11, G12 and G13 positions of the H3 peptide recognizing an R-K-SKRT-TCS-G sequence. Based on the specificity profile and in vitro methyltransferase assay targeted searches, 11 putative methylation sites in S. pombe proteins were identified from reported Clr4 interacting proteins including Mlo3. Peptide methylation was observed on Mlo3 and 7 novel target sites with strongest methylation signals on Spbc28F2.11 (HMG box-containing protein) at lysine 292 and Hrp3 (Chromodomain ATP-dep DNA helicase) at lysine 89. These data suggest that Clr4 has additional methylation substrates and it will be important to study the biological function of these novel methylation events. Furthermore, the specificity profile of Clr4 has been used to develop a quantitative method to compare and cluster specificity profiles of PKMTs. It shows that the specificity profile of Clr4 is most similar to that of the SUV39H2 enzyme, one of its human homologs. This approach will be helpful in the comparison of the recognition profiles of other families of PKMTs as well.
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Investigation of H2AX methylation by the SUV39H2 protein lysine methyltransferase.
FEBS letters, 2016Co-Authors: Maren Kirstin Schuhmacher, Srikanth Kudithipudi, Albert JeltschAbstract:The H3K9 protein lysine methyltransferase SUV39H2 was reported to methylate K134 of H2AX and stimulate H2AX phosphorylation during DNA damage response [Sone K et al. (2014) Nat Commun 5, 5691]. However, the sequence context of H2AX-K134 differs from the specificity of SUV39H2. We performed in vitro methylation reactions with SUV39H2 (and its homolog SUV39H1) using H2AX protein and peptides, but no methylation at K134 or any other lysine in H2AX was detected. Positive controls demonstrated the functionality of the assays. While our data cannot finally exclude H2AX methylation of SUV39H2 in cells, additional experimental evidence is required to validate this claim.
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Activity and specificity of the human SUV39H2 protein lysine methyltransferase.
Biochimica et biophysica acta, 2014Co-Authors: Maren Kirstin Schuhmacher, Srikanth Kudithipudi, Denis Kusevic, Sara Weirich, Albert JeltschAbstract:Abstract The SUV39H1 and SUV39H2 enzymes introduce H3K9me3, which is essential for the viability of mammalian cells. It was the aim of the present work to investigate the substrate specificity and product pattern of SUV39H2. Methylation of peptide SPOT arrays showed that SUV39H2 recognizes a long motif on H3 comprising T6–K14, with highly specific readout of R8, S10, T11 and G12 and partial specificity at T6, A7, G13 and K14. Modification of R8 and phosphorylation of S10 or T11 lead to a reduction or loss of SUV39H2 activity towards H3K9. The specificity of SUV39H2 differs from other H3K9 PKMTs, like Dim-5 or G9a, and these biochemical differences can be explained by the structures of the corresponding enzymes. Based on the specificity profile we identified additional non-histone candidate substrates in human proteins, but all of them were only weakly methylated by SUV39H2 at the peptide level. We conclude that SUV39H2 displays a high preference for the methylation of H3. Using the catalytic SET domain we show here that the enzyme prefers H3K9me0 as a substrate over H3K9me1 and H3K9me2 and it introduces the first two methyl groups into H3K9me0 in a processive reaction. SUV39H2 can transfer up to three methyl groups to lysine 9 of histone H3 but the last methylation reaction is much slower than the first two steps. We also demonstrate that the N324K mutant in the SET domain of SUV39H2 that has been shown to cause an inherited nasal skin disease in Labrador Retrievers renders SUV39H2 inactive. Differences in the circular dichroism spectra of wild type and mutant proteins indicated that the mutation causes slight structural changes.
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Non-radioactive protein lysine methyltransferase microplate assay based on reading domains.
ChemMedChem, 2013Co-Authors: Srikanth Kudithipudi, Denis Kusevic, Albert JeltschAbstract:New protein lysine methyltransferase (PKMT) assays are needed to facilitate screening for improved PKMT inhibitors, because PKMTs are mutated or overexpressed in several cancers. In cells, methylated lysine residues are recognized by reading domains such as the chromodomain of HP1β, which bind to target proteins in a lysine-methylation-specific manner. Herein we describe a sensitive, robust, and non-radioactive high-throughput PKMT assay that employs the HP1β chromodomain to detect the methylation of peptide substrates by the human SUV39H1 and SUV39H2 PKMTs. The assay has a very good dynamic range and high signal-to-noise ratio. It can be used to screen for PKMT inhibitors, as illustrated by analyzing the inhibition of SUV39H1 by chaetocin. The IC50 value of this inhibition was found to be 480 nM, which is close to its published value. Our data indicate that natural reading domains can be used as alternates to methyl-specific antibodies in PKMT assays. Reading domains can be produced recombinantly in E. coli at low cost and consistent quality, and they are accessible to protein design.
J. S. Lee - One of the best experts on this subject based on the ideXlab platform.
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Novel polymorphisms in the SUV39H2 histone methyltransferase and the risk of lung cancer
Carcinogenesis, 2006Co-Authors: Kyong Ah Yoon, B. Hwangbo, I.-j. Kim, H. J. Kee, H. S. Kim, Y. K. Jang, J.-g. Park, Sohee Park, Jong Eun Lee, J. S. LeeAbstract:Histone H3 lysine 9 (H3-K9) methylation and DNA methylation are important features of mammalian heterochromatin. Suppressor of variegation 3-9 homolog 2 (SUV39H2) is the histone methyltransferase that is required to methylate H3-K9, leading to transcriptional repression or silencing of target genes. In this study, we investigated the association of SUV39H2 polymorphisms and the risk of lung cancer. From the results of PCR direct sequencing, eight single nucleotide polymorphisms (SNPs) of SUV39H2 were identified in Korean population. In a hospital-based study of 346 lung cancer patients and 423 healthy controls, a novel SNP in the 3'-UTR of SUV39H2 (1624 G-->C) was associated with a statistically significant increase in lung cancer risk. Compared to the G/G genotype, genotypes with 1624C allele (G/C + C/C) significantly increased the susceptibility to lung cancer with adjusted odds ratio (AOR) of 2.63 (95% confidence interval (CI)= 1.10-6.29) for ever-smokers, especially in the older age group (age >or=55 years). Specifically, the variant genotype of 1624SNP was significantly associated with an increased risk of squamous cell carcinoma (AOR, 3.52; 95% CI = 1.13-9.45) in the older age group, while no significant association was found in patients with other histology. This study provided the first evidence that a novel SUV39H2 polymorphism may be an important predictive marker for lung cancer susceptibility for the smokers.
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SUV39H2 polymorphisms and the risk of lung cancer
Journal of Clinical Oncology, 2005Co-Authors: B. Hwangbo, I.-j. Kim, H. J. Kee, H. S. Kim, C. S. Park, J. M. Kim, C. G. Park, Y. K. Jang, J.-g. Park, J. S. LeeAbstract:9582 Background: Histone methylation is an important step for forming heterochromatin and the regulation of gene functions. SUV39H methyltrasferase, which targets histone H3 lysine 9, has two types of genes in human, SUV39H1 and SUV39H2. SUV39H1 was found to interact with Rb protein and Suv39h1/h2 double knockout mice had an increased risk of lymphoma. We investigated the relation of SUV39H2 polymorphisms and lung cancer. Methods: Among 8 novel SNPs identified on the SUV39H2 gene through direct PCR sequence, we examined 6004G/C (3’ UTR) polymorphism on DNA samples from 348 (M/F= 237/111) lung cancer (LC) cases and 423 (M/F=242/181) controls without cancer (NC) using an automatic direct sequencing and a real-time PCR (TaqMan assay). Results: The minor allele frequency was 9.8% and 7.1% for LC and NC, respectively. There was no effect of gender or smoking status on genotype distribution. The distribution of non-wild genotype (GC + CC) by smoking status and age group (<55 vs. ≥55 years) is shown in the table...
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SUV39H2 polymorphisms and the risk of lung cancer
Journal of Clinical Oncology, 2005Co-Authors: B. Hwangbo, I.-j. Kim, H. J. Kee, H. S. Kim, C. S. Park, J. M. Kim, C. G. Park, Y. K. Jang, J.-g. Park, J. S. LeeAbstract:9582 Background: Histone methylation is an important step for forming heterochromatin and the regulation of gene functions. SUV39H methyltrasferase, which targets histone H3 lysine 9, has two types of genes in human, SUV39H1 and SUV39H2. SUV39H1 was found to interact with Rb protein and Suv39h1/h2 double knockout mice had an increased risk of lymphoma. We investigated the relation of SUV39H2 polymorphisms and lung cancer. Methods: Among 8 novel SNPs identified on the SUV39H2 gene through direct PCR sequence, we examined 6004G/C (3’ UTR) polymorphism on DNA samples from 348 (M/F= 237/111) lung cancer (LC) cases and 423 (M/F=242/181) controls without cancer (NC) using an automatic direct sequencing and a real-time PCR (TaqMan assay). Results: The minor allele frequency was 9.8% and 7.1% for LC and NC, respectively. There was no effect of gender or smoking status on genotype distribution. The distribution of non-wild genotype (GC + CC) by smoking status and age group (
Rafael C B Sampaio - One of the best experts on this subject based on the ideXlab platform.
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21 the contrasting role of histone methyltransferases during nuclear reprogramming suv39h knockdown improves bovine somatic cell nuclear transfer while the absence of ehmt2 hampers it
Reproduction Fertility and Development, 2020Co-Authors: Rafael C B Sampaio, Dewison Ricardo Ambrizi, Ricardo P Nociti, J Pinzon, Juliano Rodrigues Sangalli, T H C De Bem, K Takahashi, J Cruz, Lawrence C SmithAbstract:The persistence of somatic epigenetic memory is indicated as the main barrier for efficient nuclear reprogramming. The augmented levels of histone methylation on somatic nuclei have been shown as the major regulator of this aberrant remodelling. Although they occupy the same site, H3K9me2 and H3K9me3 are catalysed by different histone methyltransferases (HMTs), EHMT2 and SUV39H, respectively. However, the influence of these HMTs during nuclear reprogramming is unclear. Hence, the main goal of this project was to investigate the role of HMTs responsible for H3K9 methylation during nuclear reprogramming and its consequences on embryo development. For this, we employed a small interfering RNA (siRNA)-mediated knockdown approach targeting EHMT2, SUV39H1, and SUV39H2 in bovine fetal fibroblasts. The RT-qPCR analyses showed ~80% reduction in total RNA after siRNA treatments for the target genes when compared with the control vector. We then quantified the H3K9me2 and H3K9me3 levels by immunostaining. The analysis displayed that H3K9me2 levels were diminished ~50% compared with control, whereas the reduction of H3K9me3 levels was only ~25%. Cells transfected with siRNA targeting EHMT2, SUV39H1, and SUV39H2 (All-siRNA) or control vector (control) were used as a nuclear donor on somatic cell nuclear transfer (SCNT) in five biological replicates. The IVF embryos were used as a biological control for immunostaining analysis. Embryos at both 8- to 16-cell and blastocysts stage (n = 10 from 5 replicates) were collected to evaluate the effect of HMT knockdown on H3K9me2 and H3K9me3 levels by immunostaining. We found a reduction of blastocyst rates in the treatment 28 ± 6.3 (mean ± s.e.m.) when compared with control 60 ± 4.8 (P = 0.004). The immunostaining analysis showed that the levels of H3K9me2 and H3K9me3 were higher in cloned (control) than IVF embryos (P < 0.05). Moreover, the All-siRNA group displayed a reduction in H3K9me2 levels compared with the control group and IVF through developmental stages analysed (P < 0.05). Differently, H3K9me3 levels were higher in the All-siRNA group at the 8- to 16-cell stage, but no difference was found between treated and control groups at the blastocyst stage. We, therefore, decided to test whether individual knockdown would display a different result. We then used cells transfected with siRNA targeting only EHMT2 (EHMT2-siRNA), targeting SUV39H1 + SUV39H2 (SUV-siRNA), or control vector (control) as a nuclear donor on SCNT in five biological replicates. Surprisingly, the SUV-siRNA group increased blastocyst production 38 ± 4.4 when compared with the control group 29 ± 4.4 (P = 0.01), whereas the EHMT2-siRNA showed a reduction in blastocyst rates: 21 ± 5.6 (P = 0.04). Our results indicate that EHMT2 has a key role during SCNT, possibly by its crosstalk with other modifications. Even though the SUV39H knockdown induced a small reduction in H3K9me3 levels in the nuclear donor, it was enough to increase the blastocyst rates by 10%. These results will allow us to better understand the complex mechanisms involved in the persistent epigenetic memory during nuclear reprogramming.
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21 The contrasting role of histone methyltransferases during nuclear reprogramming: SUV39H knockdown improves bovine somatic cell nuclear transfer, while the absence of EHMT2 hampers it
Reproduction Fertility and Development, 2020Co-Authors: Rafael C B Sampaio, Dewison Ricardo Ambrizi, Ricardo P Nociti, J Pinzon, Juliano Rodrigues Sangalli, K Takahashi, J Cruz, Lawrence C Smith, Pablo J. RossAbstract:The persistence of somatic epigenetic memory is indicated as the main barrier for efficient nuclear reprogramming. The augmented levels of histone methylation on somatic nuclei have been shown as the major regulator of this aberrant remodelling. Although they occupy the same site, H3K9me2 and H3K9me3 are catalysed by different histone methyltransferases (HMTs), EHMT2 and SUV39H, respectively. However, the influence of these HMTs during nuclear reprogramming is unclear. Hence, the main goal of this project was to investigate the role of HMTs responsible for H3K9 methylation during nuclear reprogramming and its consequences on embryo development. For this, we employed a small interfering RNA (siRNA)-mediated knockdown approach targeting EHMT2, SUV39H1, and SUV39H2 in bovine fetal fibroblasts. The RT-qPCR analyses showed ~80% reduction in total RNA after siRNA treatments for the target genes when compared with the control vector. We then quantified the H3K9me2 and H3K9me3 levels by immunostaining. The analysis displayed that H3K9me2 levels were diminished ~50% compared with control, whereas the reduction of H3K9me3 levels was only ~25%. Cells transfected with siRNA targeting EHMT2, SUV39H1, and SUV39H2 (All-siRNA) or control vector (control) were used as a nuclear donor on somatic cell nuclear transfer (SCNT) in five biological replicates. The IVF embryos were used as a biological control for immunostaining analysis. Embryos at both 8- to 16-cell and blastocysts stage (n = 10 from 5 replicates) were collected to evaluate the effect of HMT knockdown on H3K9me2 and H3K9me3 levels by immunostaining. We found a reduction of blastocyst rates in the treatment 28 ± 6.3 (mean ± s.e.m.) when compared with control 60 ± 4.8 (P = 0.004). The immunostaining analysis showed that the levels of H3K9me2 and H3K9me3 were higher in cloned (control) than IVF embryos (P
Yusuke Nakamura - One of the best experts on this subject based on the ideXlab platform.
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development of novel SUV39H2 inhibitors that exhibit growth suppressive effects in mouse xenograft models and regulate the phosphorylation of h2ax
Oncotarget, 2018Co-Authors: Theodore Vougiouklakis, Yusuke Nakamura, Vassiliki Saloura, Jaehyun Park, Naofumi Takamatsu, Takashi Miyamoto, Yo MatsuoAbstract:Protein methyltransferase SUV39H2 was reported to methylate histone H2AX at lysine 134 and enhance the formation of phosphorylated H2AX (γ-H2AX), which causes chemoresistance of cancer cells. We found that a series of imidazo[1,2-a]pyridine compounds that we synthesized could inhibit SUV39H2 methyltransferase activity. One of the potent compounds, OTS193320, was further analyzed in in vitro studies. The compound decreased global histone H3 lysine 9 tri-methylation levels in breast cancer cells and triggered apoptotic cell death. Combination of OTS193320 with doxorubicin (DOX) resulted in reduction of γ-H2AX levels as well as cancer cell viability compared to a single agent OTS193320 or DOX. Further optimization of inhibitors and their in vivo analysis identified a compound, OTS186935, which revealed significant inhibition of tumor growth in mouse xenograft models using MDA-MB-231 breast cancer cells and A549 lung cancer cells without any detectable toxicity. Our results suggest that the SUV39H2 inhibitors sensitize cancer cells to DOX by reduction of γ-H2AX levels in cancer cells, and collectively demonstrate that SUV39H2 inhibition warrants further investigation as a novel anti-cancer therapy.
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Development of novel SUV39H2 inhibitors that exhibit growth suppressive effects in mouse xenograft models and regulate the phosphorylation of H2AX.
Oncotarget, 2018Co-Authors: Theodore Vougiouklakis, Yusuke Nakamura, Vassiliki Saloura, Jaehyun Park, Naofumi Takamatsu, Takashi Miyamoto, Yo MatsuoAbstract:// Theodore Vougiouklakis 1 , Vassiliki Saloura 1 , Jae-Hyun Park 1 , Naofumi Takamatsu 2 , Takashi Miyamoto 2 , Yusuke Nakamura 1, 3 and Yo Matsuo 2 1 Section of Hematology/Oncology, Department of Medicine, The University of Chicago, Chicago, IL, USA 2 OncoTherapy Science Inc., Kawasaki, Japan 3 Department of Surgery, The University of Chicago, Chicago, IL, USA Correspondence to: Yusuke Nakamura, email: ynakamura@bsd.uchicago.edu Yo Matsuo, email: y-matsuo@oncotherapy.co.jp Keywords: methyltransferase; small-molecule inhibitor; SUV39H2; γ-H2AX Received: May 05, 2018 Accepted: June 01, 2018 Published: August 07, 2018 ABSTRACT Protein methyltransferase SUV39H2 was reported to methylate histone H2AX at lysine 134 and enhance the formation of phosphorylated H2AX (γ-H2AX), which causes chemoresistance of cancer cells. We found that a series of imidazo[1,2- a ]pyridine compounds that we synthesized could inhibit SUV39H2 methyltransferase activity. One of the potent compounds, OTS193320, was further analyzed in in vitro studies. The compound decreased global histone H3 lysine 9 tri-methylation levels in breast cancer cells and triggered apoptotic cell death. Combination of OTS193320 with doxorubicin (DOX) resulted in reduction of γ-H2AX levels as well as cancer cell viability compared to a single agent OTS193320 or DOX. Further optimization of inhibitors and their in vivo analysis identified a compound, OTS186935, which revealed significant inhibition of tumor growth in mouse xenograft models using MDA-MB-231 breast cancer cells and A549 lung cancer cells without any detectable toxicity. Our results suggest that the SUV39H2 inhibitors sensitize cancer cells to DOX by reduction of γ-H2AX levels in cancer cells, and collectively demonstrate that SUV39H2 inhibition warrants further investigation as a novel anti-cancer therapy.
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Abstract 4041: SUV39H2 inhibition enhances sensitivity of breast cancer cells to doxorubicin through downregulation of γ-H2AX production
Experimental and Molecular Therapeutics, 2017Co-Authors: Theodore Vougiouklakis, Vassiliki Saloura, Yo Matsuo, Yusuke NakamuraAbstract:Accumulating evidence has demonstrated the biological importance of protein methyltransferases in human tumorigenesis, and several small molecular inhibitors targeting these enzymes have been developed. Moreover, inhibiting pathways involved in DNA repair has been shown to enhance the cytotoxicity of DNA-damaging agents. Herein, we report the development of a potent SUV39H2 inhibitor (OTS193320) that decreases global histone H3 lysine 9 tri-methylation (H3K9me3) levels in cancer cells and attenuates cancer cell proliferation, and suppresses the tumor growth in mouse xenograft models. SUV39H2 (Suppressor of variegation 3-9 homolog 2), is a protein methyltransferase known to methylate histone H3 at lysine 9 (H3K9), and recently reported to methylate histone H2AX at lysine 134, which enhances the accumulation of phosphorylated H2AX (γ-H2AX) and regulates the DNA repair pathway in human cancer. SUV39H2 is highly expressed in many cancer types, including lung and breast cancers, while its expression levels are restricted to testis in normal adult tissues. Exposure of OTS193320 to two triple negative breast cancer cell lines attenuated H3K9me3 levels in a dose-dependent manner and triggered apoptotic cell death. Combination of OTS193320 and doxorubicin (DOX) resulted in reduction of γ-H2AX levels compared to single agent DOX, as visualized on western blot and immunocytochemical analysis. Furthermore, combination therapy attenuated the levels of p53-binding protein 1, which is reported to co-localize with γ-H2AX foci, compared to single agent DOX. Cell viability assays demonstrated a significant growth suppressive effect when OTS193320 was combined with DOX, compared to single agent treatment of either drug, suggesting chemosensitization to DOX. Importantly, in a mouse xenograft model of A549 lung cancer cells, we observed a tumor growth inhibition of 60.8% at day 14 with 25mg/kg intravenous administration of the inhibitor, without significant body weight loss or toxicity. Immunohistochemical staining of tumors treated with the inhibitor demonstrated a significantly lower number of Ki-67 positive cells and attenuated distribution patterns of H3K9me3 compared to the control tumors. Collectively, our results demonstrate that SUV39H2 inhibition may be a promising approach to develop a novel class of anti-cancer treatment. Citation Format: Theodore Vougiouklakis, Vassiliki Saloura, Yo Matsuo, Yusuke Nakamura. SUV39H2 inhibition enhances sensitivity of breast cancer cells to doxorubicin through downregulation of γ-H2AX production [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4041. doi:10.1158/1538-7445.AM2017-4041
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Automethylation of SUV39H2, an oncogenic histone lysine methyltransferase, regulates its binding affinity to substrate proteins
Oncotarget, 2016Co-Authors: Lianhua Piao, Takehiro Suzuki, Naoshi Dohmae, Yusuke Nakamura, Makoto Nakakido, Ryuji HamamotoAbstract:// Lianhua Piao 1 , Makoto Nakakido 1 , Takehiro Suzuki 2 , Naoshi Dohmae 2 , Yusuke Nakamura 1 , Ryuji Hamamoto 1, 3 1 Section of Hematology/Oncology, Department of Medicine, The University of Chicago, Chicago, IL 60637, USA 2 Biomolecular Characterization Unit, RIKEN Center for Sustainable Resource Science, Wako, Saitama 351-0198, Japan 3 Division of Molecular Modification and Cancer Biology, National Cancer Center, Chuo-ku, Tokyo 104-0045, Japan Correspondence to: Ryuji Hamamoto, e-mail: ryujihamamoto@gmail.com Keywords: SUV39H2, oncogene, automethylation, SET domain Received: November 12, 2015 Accepted: February 25, 2016 Published: March 14, 2016 ABSTRACT We previously reported that the histone lysine methyltransferase SUV39H2, which is overexpressed in various types of human cancer, plays a critical role in the DNA repair after double strand breakage, and possesses oncogenic activity. Although its biological significance in tumorigenesis has been elucidated, the regulatory mechanism of SUV39H2 activity through post-translational modification is not well known. In this study, we demonstrate in vitro and in vivo automethylation of SUV39H2 at lysine 392. Automethylation of SUV39H2 led to impairment of its binding affinity to substrate proteins such as histone H3 and LSD1. Furthermore, we observed that hyper-automethylated SUV39H2 reduced methylation activities to substrates through affecting the binding affinity to substrate proteins. Our finding unveils a novel autoregulatory mechanism of SUV39H2 through lysine automethylation.
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SUV39H2 methylates and stabilizes LSD1 by inhibiting polyubiquitination in human cancer cells
Oncotarget, 2015Co-Authors: Lianhua Piao, Takehiro Suzuki, Naoshi Dohmae, Yusuke Nakamura, Ryuji HamamotoAbstract:// Lianhua Piao 1 , Takehiro Suzuki 2 , Naoshi Dohmae 2 , Yusuke Nakamura 1 and Ryuji Hamamoto 1 1 Section of Hematology/Oncology, Department of Medicine, The University of Chicago, Chicago, IL, USA 2 Biomolecular Characterizaion Unit, RIKEN Center for Sustainable Resource Science, Wako, Saitama, Japan Correspondence to: Ryuji Hamamoto, email: // Keywords : SUV39H2, carcinogenesis, LSD1, non-histone protein methylation Received : May 02, 2015 Accepted : June 26, 2015 Published : July 03, 2015 Abstract LSD1 is a histone lysine demethylase, which is highly expressed in multiple types of human cancer. Although its roles in transcriptional regulation have been well-studied, functional regulation of LSD1 by post-translational modifications still remains unknown. Here, we demonstrate that the histone lysine methyltransferase SUV39H2 trimethylated LSD1 on lysine 322. Knockdown of SUV39H2 resulted in a decrease of LSD1 protein even though the mRNA levels were unchanged. SUV39H2-induced LSD1 methylation suppresses LSD1 polyubiquitination and subsequent degradation. In addition, we also observed indirect effect of SUV39H2 overexpression on LSD1-target genes. Our results reveal the regulatory mechanism of LSD1 protein through its lysine methylation by SUV39H2 in human cancer cells.
Manfred Schmid - One of the best experts on this subject based on the ideXlab platform.
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Isolation and Characterization of SUV39H2, a Second Histone H3 Methyltransferase Gene That Displays Testis-Specific Expression
Molecular and cellular biology, 2000Co-Authors: Dónal O'carroll, Stephen Rea, Susanne Opravil, Antoine H. F. M. Peters, Manfred Schmid, Harry Scherthan, Andrew R. Haynes, Götz Laible, Angelika Lebersorger, Martin JerratschAbstract:Higher-order chromatin has been implicated in epigenetic gene control and in the functional organization of chromosomes. We have recently discovered mouse (Suv39h1) and human (SUV39H1) histone H3 lysine 9-selective methyltransferases (Suv39h HMTases) and shown that they modulate chromatin dynamics in somatic cells. We describe here the isolation, chromosomal assignment, and characterization of a second murine gene, SUV39H2. Like Suv39h1, SUV39H2 encodes an H3 HMTase that shares 59% identity with Suv39h1 but which differs by the presence of a highly basic N terminus. Using fluorescent in situ hybridization and haplotype analysis, the SUV39H2 locus was mapped to the subcentromeric region of mouse chromosome 2, whereas the Suv39h1 locus resides at the tip of the mouse X chromosome. Notably, although both Suv39h loci display overlapping expression profiles during mouse embryogenesis, SUV39H2 transcripts remain specifically expressed in adult testes. Immunolocalization of SUV39H2 protein during spermatogenesis indicates enriched distribution at the heterochromatin from the leptotene to the round spermatid stage. Moreover, SUV39H2 specifically accumulates with chromatin of the sex chromosomes (XY body) which undergo transcriptional silencing during the first meiotic prophase. These data are consistent with redundant enzymatic roles for Suv39h1 and SUV39H2 during mouse development and suggest an additional function of the SUV39H2 HMTase in organizing meiotic heterochromatin that may even impart an epigenetic imprint to the male germ line.
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Regulation of chromatin structure by site-specific histone H3 methyltransferases
Nature, 2000Co-Authors: Stephen Rea, Dónal O'carroll, Zu Wen Sun, Karl Mechtier, Susanne Opravil, Chris P Ponting, Brian D. Strahl, Frank Eisenhaber, Manfred Schmid, C D AllisAbstract:The organization of chromatin into higher-order structures influences chromosome function and epigenetic gene regulation. Higher-order chromatin has been proposed to be nucleated by the covalent modification of histone tails and the subsequent establishment of chromosomal subdomains by non-histone modifier factors. Here we show that human SUV39H1 and murine Suv39h1--mammalian homologues of Drosophila Su(var)3-9 and of Schizosaccharomyces pombe clr4--encode histone H3-specific methyltransferases that selectively methylate lysine 9 of the amino terminus of histone H3 in vitro. We mapped the catalytic motif to the evolutionarily conserved SET domain, which requires adjacent cysteine-rich regions to confer histone methyltransferase activity. Methylation of lysine 9 interferes with phosphorylation of serine 10, but is also influenced by pre-existing modifications in the amino terminus of H3. In vivo, deregulated SUV39H1 or disrupted Suv39h activity modulate H3 serine 10 phosphorylation in native chromatin and induce aberrant mitotic divisions. Our data reveal a functional interdependence of site-specific H3 tail modifications and suggest a dynamic mechanism for the regulation of higher-order chromatin.
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functional mammalian homologues of the drosophila pev modifier su var 3 9 encode centromere associated proteins which complex with the heterochromatin component m31
The EMBO Journal, 1999Co-Authors: Louise Aagaard, Manfred Schmid, Götz Laible, Philipp Selenko, Gunna Schotta, Steffi Kuhfittig, Andrea Wolf, Angelika Lebersorge, Prim Singh, Gunte ReuteAbstract:The chromo and SET domains are conserved sequence motifs present in chromosomal proteins that function in epigenetic control of gene expression, presumably by modulating higher order chromatin. Based on sequence information from the SET domain, we have isolated human (SUV39H1) and mouse (Suv39h1) homologues of the dominant Drosophila modifier of position-effect-variegation (PEV) Su(var)3-9. Mammalian homologues contain, in addition to the SET domain, the characteristic chromo domain, a combination that is also preserved in the Schizosaccharyomyces pombe silencing factor clr4. Chromatin-dependent gene regulation is demonstrated by the potential of human SUV39H1 to increase repression of the pericentromeric white marker gene in transgenic flies. Immunodetection of endogenous Suv39h1/SUV39H1 proteins in a variety of mammalian cell lines reveals enriched distribution at heterochromatic foci during interphase and centromere-specific localization during metaphase. In addition, Suv39h1/SUV39H1 proteins associate with M31, currently the only other characterized mammalian SU(VAR) homologue. These data indicate the existence of a mammalian SU(VAR) complex and define Suv39h1/SUV39H1 as novel components of mammalian higher order chromatin.
