Publications by authors named "Taiping Chen"

80 Publications

The Essential Function of SETDB1 in Homologous Chromosome Pairing and Synapsis during Meiosis.

Cell Rep 2021 Jan;34(1):108575

Yale Stem Cell Center and Department of Cell Biology, Yale University School of Medicine, New Haven, CT 06520, USA. Electronic address:

SETDB1 is a histone-lysine N-methyltransferase critical for germline development. However, its function in early meiotic prophase I remains unknown. Here, we report that Setdb1 null spermatocytes display aberrant centromere clustering during leptotene, bouquet formation during zygotene, and subsequent failure in pairing and synapsis of homologous chromosomes, as well as compromised meiotic silencing of unsynapsed chromatin, which leads to meiotic arrest before pachytene and apoptosis of spermatocytes. H3K9me3 is enriched in centromeric or pericentromeric regions and is present in many sites throughout the genome, with a subset changed in the Setdb1 mutant. These observations indicate that SETDB1-mediated H3K9me3 is essential for the bivalent formation in early meiosis. Transcriptome analysis reveals the function of SETDB1 in repressing transposons and transposon-proximal genes and in regulating meiotic and somatic lineage genes. These findings highlight a mechanism in which SETDB1-mediated H3K9me3 during early meiosis ensures the formation of homologous bivalents and survival of spermatocytes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.celrep.2020.108575DOI Listing
January 2021

The inactive Dnmt3b3 isoform preferentially enhances Dnmt3b-mediated DNA methylation.

Genes Dev 2020 Nov 1;34(21-22):1546-1558. Epub 2020 Oct 1.

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Houston, Texas 77030, USA.

The de novo DNA methyltransferases Dnmt3a and Dnmt3b play crucial roles in developmental and cellular processes. Their enzymatic activities are stimulated by a regulatory protein Dnmt3L (Dnmt3-like) in vitro. However, genetic evidence indicates that Dnmt3L functions predominantly as a regulator of Dnmt3a in germ cells. How Dnmt3a and Dnmt3b activities are regulated during embryonic development and in somatic cells remains largely unknown. Here we show that Dnmt3b3, a catalytically inactive Dnmt3b isoform expressed in differentiated cells, positively regulates de novo methylation by Dnmt3a and Dnmt3b with a preference for Dnmt3b. Dnmt3b3 is equally potent as Dnmt3L in stimulating the activities of Dnmt3a2 and Dnmt3b2 in vitro. Like Dnmt3L, Dnmt3b3 forms a complex with Dnmt3a2 with a stoichiometry of 2:2. However, rescue experiments in triple-knockout (TKO) mouse embryonic stem cells (mESCs) reveal that Dnmt3b3 prefers Dnmt3b2 over Dnmt3a2 in remethylating genomic sequences. Dnmt3a2, an active isoform that lacks the N-terminal uncharacterized region of Dnmt3a1 including a nuclear localization signal, has very low activity in TKO mESCs, indicating that an accessory protein is absolutely required for its function. Our results suggest that Dnmt3b3 and perhaps similar Dnmt3b isoforms facilitate de novo DNA methylation during embryonic development and in somatic cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/gad.341925.120DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7608744PMC
November 2020

Germline DNMT3A mutation in familial acute myeloid leukaemia.

Epigenetics 2020 Aug 28:1-10. Epub 2020 Aug 28.

Department of Genomic Medicine, The University of Texas MD Anderson Cancer Center , Houston, TX, USA.

Acute myeloid leukaemia (AML) is a heterogeneous myeloid malignancy characterized by recurrent clonal events, including mutations in epigenetically relevant genes such as , and . Next-generation sequencing analysis of a mother and son pair who both developed adult-onset diploid AML identified a novel germline missense mutation p.P709S. The p.P709S protein-altering variant resides in the highly conserved catalytic DNMT3A methyltransferase domain. Functional studies demonstrate that the p.P709S variant confers dominant negative effects when interacting with wildtype . LINE-1 pyrosequencing and reduced representation bisulphite sequencing (RBBS) analysis demonstrated global DNA hypomethylation in germline samples, not present in the leukaemic samples. Somatic acquisition of p.R172K mutations, in concert with additional acquired clonal DNMT3A events in both patients at the time of AML diagnosis, confirms the important pathogenic interaction of epigenetically active genes, and implies a strong selection and regulation of methylation in leukaemogenesis. Improved characterization of germline mutations may enable us to better predict malignant clonal evolution, improving our ability to provide customized treatment or future preventative strategies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/15592294.2020.1809871DOI Listing
August 2020

Author Correction: LRIG1 is a pleiotropic androgen receptor-regulated feedback tumor suppressor in prostate cancer.

Nat Commun 2020 06 4;11(1):2895. Epub 2020 Jun 4.

Department of Pharmacology and Therapeutics, Roswell Park Comprehensive Cancer Center, Buffalo, NY, 14263, USA.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-020-16615-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7272421PMC
June 2020

The acute myeloid leukemia variant DNMT3A Arg882His is a DNMT3B-like enzyme.

Nucleic Acids Res 2020 04;48(7):3761-3775

Department of Biochemistry, Purdue University, West Lafayette, IN 47907, USA.

We have previously shown that the highly prevalent acute myeloid leukemia (AML) mutation, Arg882His, in DNMT3A disrupts its cooperative mechanism and leads to reduced enzymatic activity, thus explaining the genomic hypomethylation in AML cells. However, the underlying cause of the oncogenic effect of Arg882His in DNMT3A is not fully understood. Here, we discovered that DNMT3A WT enzyme under conditions that favor non-cooperative kinetic mechanism as well as DNMT3A Arg882His variant acquire CpG flanking sequence preference akin to that of DNMT3B, which is non-cooperative. We tested if DNMT3A Arg882His could preferably methylate DNMT3B-specific target sites in vivo. Rescue experiments in Dnmt3a/3b double knockout mouse embryonic stem cells show that the corresponding Arg878His mutation in mouse DNMT3A severely impairs its ability to methylate major satellite DNA, a DNMT3A-preferred target, but has no overt effect on the ability to methylate minor satellite DNA, a DNMT3B-preferred target. We also observed a previously unappreciated CpG flanking sequence bias in major and minor satellite repeats that is consistent with DNMT3A and DNMT3B specificity suggesting that DNA methylation patterns are guided by the sequence preference of these enzymes. We speculate that aberrant methylation of DNMT3B target sites could contribute to the oncogenic potential of DNMT3A AML variant.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/nar/gkaa139DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7144950PMC
April 2020

Author Correction: De novo identification of essential protein domains from CRISPR-Cas9 tiling-sgRNA knockout screens.

Nat Commun 2020 Feb 25;11(1):1134. Epub 2020 Feb 25.

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Smithville, TX, 78957, USA.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-020-14940-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7042308PMC
February 2020

The ZBTB24-CDCA7 axis regulates HELLS enrichment at centromeric satellite repeats to facilitate DNA methylation.

Protein Cell 2020 03;11(3):214-218

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Smithville, TX, 78957, USA.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s13238-019-00682-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7026229PMC
March 2020

LRIG1 is a pleiotropic androgen receptor-regulated feedback tumor suppressor in prostate cancer.

Nat Commun 2019 12 2;10(1):5494. Epub 2019 Dec 2.

Department of Pharmacology and Therapeutics, Roswell Park Comprehensive Cancer Center, Buffalo, NY, 14263, USA.

LRIG1 has been reported to be a tumor suppressor in gastrointestinal tract and epidermis. However, little is known about the expression, regulation and biological functions of LRIG1 in prostate cancer (PCa). We find that LRIG1 is overexpressed in PCa, but its expression correlates with better patient survival. Functional studies reveal strong tumor-suppressive functions of LRIG1 in both AR and AR xenograft models, and transgenic expression of LRIG1 inhibits tumor development in Hi-Myc and TRAMP models. LRIG1 also inhibits castration-resistant PCa and exhibits therapeutic efficacy in pre-established tumors. We further show that 1) AR directly transactivates LRIG1 through binding to several AR-binding sites in LRIG1 locus, and 2) LRIG1 dampens ERBB expression in a cell type-dependent manner and inhibits ERBB2-driven tumor growth. Collectively, our study indicates that LRIG1 represents a pleiotropic AR-regulated feedback tumor suppressor that functions to restrict oncogenic signaling from AR, Myc, ERBBs, and, likely, other oncogenic drivers.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-019-13532-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6889295PMC
December 2019

De novo identification of essential protein domains from CRISPR-Cas9 tiling-sgRNA knockout screens.

Nat Commun 2019 10 4;10(1):4541. Epub 2019 Oct 4.

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Smithville, TX, 78957, USA.

High-throughput CRISPR-Cas9 knockout screens using a tiling-sgRNA design permit in situ evaluation of protein domain function. Here, to facilitate de novo identification of essential protein domains from such screens, we propose ProTiler, a computational method for the robust mapping of CRISPR knockout hyper-sensitive (CKHS) regions, which refer to the protein regions associated with a strong sgRNA dropout effect in the screens. Applied to a published CRISPR tiling screen dataset, ProTiler identifies 175 CKHS regions in 83 proteins. Of these CKHS regions, more than 80% overlap with annotated Pfam domains, including all of the 15 known drug targets in the dataset. ProTiler also reveals unannotated essential domains, including the N-terminus of the SWI/SNF subunit SMARCB1, which is validated experimentally. Surprisingly, the CKHS regions are negatively correlated with phosphorylation and acetylation sites, suggesting that protein domains and post-translational modification sites have distinct sensitivities to CRISPR-Cas9 mediated amino acids loss.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-019-12489-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6778102PMC
October 2019

Synthesis of N-Substituted Benzamide Derivatives and their Evaluation as Antitumor Agents.

Med Chem 2020 ;16(4):555-562

State Key Laboratory of Biotherapy, West China Hospital, West China Medical School, Chengdu 610041, Sichuan, China.

Background: Histone deacetylases inhibitors (HDACIs) with different chemical structures have been reported to play an important role in the treatment of cancer.

Objective: The study aims to modify the structure of Entinostat (MS-275) to discover new compounds with improved anti-proliferative activities and perform SAR studies on this class of bioactive compounds.

Methods: Fourteen N-substituted benzamide derivatives were synthesized and their antiproliferative activities were tested with four cancer cell lines (MCF-7, A549, K562 and MDA-MB- 231) by MTT assay.

Results: Compared with MS-275, six compounds exhibited comparable or even better antiproliferative activities against specific/certain cancer cell lines.

Conclusion: The preliminary SARs showed that (ⅰ) the 2-substituent of the phenyl ring in the R group and heteroatoms of amide which can chelate with zinc ion are critical to the antiproliferative activity and (ⅱ) chlorine atom or nitro-group on the same benzene ring largely decreases their anti-proliferative activity. Molecular docking study illustrated the interaction (binding affinity) between the synthesized compounds and HDAC2 was observed to be similar to that of MS-275.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.2174/1573406415666190712120611DOI Listing
January 2021

Systematic evaluation of RNA-Seq preparation protocol performance.

BMC Genomics 2019 Jul 11;20(1):571. Epub 2019 Jul 11.

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Science Park, Smithville, TX, 78957, USA.

Background: RNA-Seq is currently the most widely used tool to analyze whole-transcriptome profiles. There are numerous commercial kits available to facilitate preparing RNA-Seq libraries; however, it is still not clear how some of these kits perform in terms of: 1) ribosomal RNA removal; 2) read coverage or recovery of exonic vs. intronic sequences; 3) identification of differentially expressed genes (DEGs); and 4) detection of long non-coding RNA (lncRNA). In RNA-Seq analysis, understanding the strengths and limitations of commonly used RNA-Seq library preparation protocols is important, as this technology remains costly and time-consuming.

Results: In this study, we present a comprehensive evaluation of four RNA-Seq kits. We used three standard input protocols: Illumina TruSeq Stranded Total RNA and mRNA kits, a modified NuGEN Ovation v2 kit, and the TaKaRa SMARTer Ultra Low RNA Kit v3. Our evaluation of these kits included quality control measures such as overall reproducibility, 5' and 3' end-bias, and the identification of DEGs, lncRNAs, and alternatively spliced transcripts. Overall, we found that the two Illumina kits were most similar in terms of recovering DEGs, and the Illumina, modified NuGEN, and TaKaRa kits allowed identification of a similar set of DEGs. However, we also discovered that the Illumina, NuGEN and TaKaRa kits each enriched for different sets of genes.

Conclusions: At the manufacturers' recommended input RNA levels, all the RNA-Seq library preparation protocols evaluated were suitable for distinguishing between experimental groups, and the TruSeq Stranded mRNA kit was universally applicable to studies focusing on protein-coding gene profiles. The TruSeq protocols tended to capture genes with higher expression and GC content, whereas the modified NuGEN protocol tended to capture longer genes. The SMARTer Ultra Low RNA Kit may be a good choice at the low RNA input level, although it was inferior to the TruSeq mRNA kit at standard input level in terms of rRNA removal, exonic mapping rates and recovered DEGs. Therefore, the choice of RNA-Seq library preparation kit can profoundly affect data outcomes. Consequently, it is a pivotal parameter to consider when designing an RNA-Seq experiment.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s12864-019-5953-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6625085PMC
July 2019

Structural basis of specific DNA binding by the transcription factor ZBTB24.

Nucleic Acids Res 2019 09;47(16):8388-8398

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA.

ZBTB24, encoding a protein of the ZBTB family of transcriptional regulators, is one of four known genes-the other three being DNMT3B, CDCA7 and HELLS-that are mutated in immunodeficiency, centromeric instability and facial anomalies (ICF) syndrome, a genetic disorder characterized by DNA hypomethylation and antibody deficiency. The molecular mechanisms by which ZBTB24 regulates gene expression and the biological functions of ZBTB24 are poorly understood. Here, we identified a 12-bp consensus sequence [CT(G/T)CCAGGACCT] occupied by ZBTB24 in the mouse genome. The sequence is present at multiple loci, including the Cdca7 promoter region, and ZBTB24 binding is mostly associated with gene activation. Crystallography and DNA-binding data revealed that the last four of the eight zinc fingers (ZFs) (i.e. ZF5-8) in ZBTB24 confer specificity of DNA binding. Two ICF missense mutations have been identified in the ZBTB24 ZF domain, which alter zinc-binding cysteine residues. We demonstrated that the corresponding C382Y and C407G mutations in mouse ZBTB24 abolish specific DNA binding and fail to induce Cdca7 expression. Our analyses indicate and suggest a structural basis for the sequence specific recognition by a transcription factor centrally important for the pathogenesis of ICF syndrome.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/nar/gkz557DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6895263PMC
September 2019

DNA Methylation Reprogramming during Mammalian Development.

Genes (Basel) 2019 03 29;10(4). Epub 2019 Mar 29.

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, 1808 Park Road 1C, Smithville, TX 78957, USA.

DNA methylation (5-methylcytosine, 5mC) is a major form of DNA modification in the mammalian genome that plays critical roles in chromatin structure and gene expression. In general, DNA methylation is stably maintained in somatic tissues. However, DNA methylation patterns and levels show dynamic changes during development. Specifically, the genome undergoes two waves of global demethylation and remethylation for the purpose of producing the next generation. The first wave occurs in the germline, initiated with the erasure of global methylation in primordial germ cells (PGCs) and completed with the establishment of sex-specific methylation patterns during later stages of germ cell development. The second wave occurs after fertilization, including the erasure of most methylation marks inherited from the gametes and the subsequent establishment of the embryonic methylation pattern. The two waves of DNA methylation reprogramming involve both distinct and shared mechanisms. In this review article, we provide an overview of the key reprogramming events, focusing on the important players in these processes, including DNA methyltransferases (DNMTs) and ten-eleven translocation (TET) family of 5mC dioxygenases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/genes10040257DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6523607PMC
March 2019

Lysine-specific demethylase-2 is distinctively involved in brown and beige adipogenic differentiation.

FASEB J 2019 04 25;33(4):5300-5311. Epub 2019 Jan 25.

Department of Medical Cell Biology, Institute of Molecular Embryology and Genetics, Kumamoto University, Kumamoto, Japan.

Transcriptional and epigenetic regulation is fundamentally involved in initiating and maintaining progression of cellular differentiation. The 2 types of thermogenic adipocytes, brown and beige, are thought to be of different origins but share functionally similar phenotypes. Here, we report that lysine-specific demethylase 2 (LSD2) regulates the expression of genes associated with lineage identity during the differentiation of brown and beige adipogenic progenitors in mice. In HB2 mouse brown preadipocytes, short hairpin RNA-mediated knockdown (KD) of LSD2 impaired formation of lipid droplet-containing adipocytes and down-regulated brown adipogenesis-associated genes. Transcriptomic analysis revealed that myogenesis-associated genes were up-regulated in LSD2-KD cells under adipogenic induction. In addition, loss of LSD2 during later phases of differentiation had no obvious influence on adipogenic traits, suggesting that LSD2 functions during earlier phases of brown adipocyte differentiation. Using adipogenic cells from the brown adipose tissues of LSD2-knockout (KO) mice, we found reduced expression of brown adipogenesis genes, whereas myogenesis genes were not affected. In contrast, when LSD2-KO cells from inguinal white adipose tissues were subjected to beige induction, these cells showed a dramatic rise in myogenic gene expression. Collectively, these results suggest that LSD2 regulates distinct sets of genes during brown and beige adipocyte formation.-Takase, R., Hino, S., Nagaoka, K., Anan, K., Kohrogi, K., Araki, H., Hino, Y., Sakamoto, A., Nicholson, T. B., Chen, T., Nakao, M. Lysine-specific demethylase-2 is distinctively involved in brown and beige adipogenic differentiation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1096/fj.201801422RRDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6436657PMC
April 2019

DNMT3L facilitates DNA methylation partly by maintaining DNMT3A stability in mouse embryonic stem cells.

Nucleic Acids Res 2019 01;47(1):152-167

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Smithville, TX 78957, USA.

DNMT3L (DNMT3-like), a member of the DNMT3 family, has no DNA methyltransferase activity but regulates de novo DNA methylation. While biochemical studies show that DNMT3L is capable of interacting with both DNMT3A and DNMT3B and stimulating their enzymatic activities, genetic evidence suggests that DNMT3L is essential for DNMT3A-mediated de novo methylation in germ cells but is dispensable for de novo methylation during embryogenesis, which is mainly mediated by DNMT3B. How DNMT3L regulates DNA methylation and what determines its functional specificity are not well understood. Here we show that DNMT3L-deficient mouse embryonic stem cells (mESCs) exhibit downregulation of DNMT3A, especially DNMT3A2, the predominant DNMT3A isoform in mESCs. DNA methylation analysis of DNMT3L-deficient mESCs reveals hypomethylation at many DNMT3A target regions. These results confirm that DNMT3L is a positive regulator of DNA methylation, contrary to a previous report that, in mESCs, DNMT3L regulates DNA methylation positively or negatively, depending on genomic regions. Mechanistically, DNMT3L forms a complex with DNMT3A2 and prevents DNMT3A2 from being degraded. Restoring the DNMT3A protein level in DNMT3L-deficient mESCs partially recovers DNA methylation. Thus, our work uncovers a role for DNMT3L in maintaining DNMT3A stability, which contributes to the effect of DNMT3L on DNMT3A-dependent DNA methylation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/nar/gky947DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6326784PMC
January 2019

Deficient LRRC8A-dependent volume-regulated anion channel activity is associated with male infertility in mice.

JCI Insight 2018 08 23;3(16). Epub 2018 Aug 23.

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Smithville, Texas, USA.

Ion channel-controlled cell volume regulation is of fundamental significance to the physiological function of sperm. In addition to volume regulation, LRRC8A-dependent volume-regulated anion channel (VRAC) activity is involved in cell cycle progression, insulin signaling, and cisplatin resistance. Nevertheless, the contribution of LRRC8A and its dependent VRAC activity in the germ cell lineage remain unknown. By utilizing a spontaneous Lrrc8a mouse mutation (c.1325delTG, p.F443*) and genetically engineered mouse models, we demonstrate that LRRC8A-dependent VRAC activity is essential for male germ cell development and fertility. Lrrc8a-null male germ cells undergo progressive degeneration independent of the apoptotic pathway during postnatal testicular development. Lrrc8a-deficient mouse sperm exhibit multiple morphological abnormalities of the flagella (MMAF), a feature commonly observed in the sperm of infertile human patients. Importantly, we identified a human patient with a rare LRRC8A hypomorphic mutation (c.1634G>A, p.Arg545His) possibly linked to Sertoli cell-only syndrome (SCOS), a male sterility disorder characterized by the loss of germ cells. Thus, LRRC8A is a critical factor required for germ cell development and volume regulation in the mouse, and it might serve as a novel diagnostic and therapeutic target for SCOS patients.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1172/jci.insight.99767DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6141173PMC
August 2018

DNMT3A and TET1 cooperate to regulate promoter epigenetic landscapes in mouse embryonic stem cells.

Genome Biol 2018 07 12;19(1):88. Epub 2018 Jul 12.

Stem Cells and Regenerative Medicine Center, Baylor College of Medicine, Houston, TX, 77030, USA.

Background: DNA methylation is a heritable epigenetic mark, enabling stable but reversible gene repression. In mammalian cells, DNA methyltransferases (DNMTs) are responsible for modifying cytosine to 5-methylcytosine (5mC), which can be further oxidized by the TET dioxygenases to ultimately cause DNA demethylation. However, the genome-wide cooperation and functions of these two families of proteins, especially at large under-methylated regions, called canyons, remain largely unknown.

Results: Here we demonstrate that DNMT3A and TET1 function in a complementary and competitive manner in mouse embryonic stem cells to mediate proper epigenetic landscapes and gene expression. The longer isoform of DNMT3A, DNMT3A1, exhibits significant enrichment at distal promoters and canyon edges, but is excluded from proximal promoters and canyons where TET1 shows prominent binding. Deletion of Tet1 increases DNMT3A1 binding capacity at and around genes with wild-type TET1 binding. However, deletion of Dnmt3a has a minor effect on TET1 binding on chromatin, indicating that TET1 may limit DNA methylation partially by protecting its targets from DNMT3A and establishing boundaries for DNA methylation. Local CpG density may determine their complementary binding patterns and therefore that the methylation landscape is encoded in the DNA sequence. Furthermore, DNMT3A and TET1 impact histone modifications which in turn regulate gene expression. In particular, they regulate Polycomb Repressive Complex 2 (PRC2)-mediated H3K27me3 enrichment to constrain gene expression from bivalent promoters.

Conclusions: We conclude that DNMT3A and TET1 regulate the epigenome and gene expression at specific targets via their functional interplay.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13059-018-1464-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6042404PMC
July 2018

Identification of Rpl29 as a major substrate of the lysine methyltransferase Set7/9.

J Biol Chem 2018 08 29;293(33):12770-12780. Epub 2018 Jun 29.

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Smithville, Texas 78957; Center for Cancer Epigenetics, The University of Texas MD Anderson Cancer Center, Smithville, Texas 78957; Program in Genetics and Epigenetics, The University of Texas MD Anderson Cancer Center UT Health Graduate School of Biomedical Sciences, Houston, Texas 77030. Electronic address:

Set7/9 (also known as Set7, Set9, Setd7, and Kmt7) is a lysine methyltransferase that catalyzes the methylation of multiple substrates, including histone H3 and non-histone proteins. Although not essential for normal development and physiology, Set7/9-mediated methylation events play important roles in regulating cellular pathways involved in various human diseases, making Set7/9 a promising therapeutic target. Multiple Set7/9 inhibitors have been developed, which exhibit varying degrees of potency and selectivity However, validation of these compounds has been hampered by the lack of a reliable cellular biomarker for Set7/9 activity. Here, we report the identification of Rpl29, a ribosomal protein abundantly expressed in all cell types, as a major substrate of Set7/9. We show that Rpl29 lysine 5 (Rpl29K5) is methylated exclusively by Set7/9 and can be demethylated by Lsd1 (also known as Kdm1a). Rpl29 is not a core component of the ribosome translational machinery and plays a regulatory role in translation efficiency. Our results indicate that Rpl29 methylation has no effect on global protein synthesis but affects Rpl29 subcellular localization. Using an Rpl29 methylation-specific antibody, we demonstrate that Rpl29K5 methylation is present ubiquitously and validate that ()-PFI-2, a Set7/9 inhibitor, efficiently reduces Rpl29K5 methylation in cell lines. Thus, Rpl29 methylation can serve as a specific cellular biomarker for measuring Set7/9 activity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/jbc.RA118.002890DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6102145PMC
August 2018

The Arginine Methyltransferase PRMT6 Regulates DNA Methylation and Contributes to Global DNA Hypomethylation in Cancer.

Cell Rep 2017 Dec;21(12):3390-3397

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Smithville, TX 78957, USA; Center for Cancer Epigenetics, The University of Texas MD Anderson Cancer Center, Smithville, TX 78957, USA; Program in Genetics and Epigenetics, The University of Texas MD Anderson Cancer Center, UTHealth Graduate School of Biomedical Sciences, Houston, TX 77030, USA. Electronic address:

DNA methylation plays crucial roles in chromatin structure and gene expression. Aberrant DNA methylation patterns, including global hypomethylation and regional hypermethylation, are associated with cancer and implicated in oncogenic events. How DNA methylation is regulated in developmental and cellular processes and dysregulated in cancer is poorly understood. Here, we show that PRMT6, a protein arginine methyltransferase responsible for asymmetric dimethylation of histone H3 arginine 2 (H3R2me2a), negatively regulates DNA methylation and that PRMT6 upregulation contributes to global DNA hypomethylation in cancer. Mechanistically, PRMT6 overexpression impairs chromatin association of UHRF1, an accessory factor of DNMT1, resulting in passive DNA demethylation. The effect is likely due to elevated H3R2me2a, which inhibits the interaction between UHRF1 and histone H3. Our work identifies a mechanistic link between protein arginine methylation and DNA methylation, which is disrupted in cancer.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.celrep.2017.11.082DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5753604PMC
December 2017

Zscan4 Inhibits Maintenance DNA Methylation to Facilitate Telomere Elongation in Mouse Embryonic Stem Cells.

Cell Rep 2017 Aug;20(8):1936-1949

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Smithville, TX 78957, USA; Center for Cancer Epigenetics, The University of Texas MD Anderson Cancer Center, Smithville, TX 78957, USA; The University of Texas MD Anderson Cancer Center UTHealth Graduate School of Biomedical Sciences, Houston, TX 77030, USA. Electronic address:

Proper telomere length is essential for embryonic stem cell (ESC) self-renewal and pluripotency. Mouse ESCs (mESCs) sporadically convert to a transient totipotent state similar to that of two-cell (2C) embryos to recover shortened telomeres. Zscan4, which exhibits a burst of expression in 2C-like mESCs, is required for telomere extension in these cells. However, the mechanism by which Zscan4 extends telomeres remains elusive. Here, we show that Zscan4 facilitates telomere elongation by inducing global DNA demethylation through downregulation of Uhrf1 and Dnmt1, major components of the maintenance DNA methylation machinery. Mechanistically, Zscan4 recruits Uhrf1 and Dnmt1 and promotes their degradation, which depends on the E3 ubiquitin ligase activity of Uhrf1. Blocking DNA demethylation prevents telomere elongation associated with Zscan4 expression, suggesting that DNA demethylation mediates the effect of Zscan4. Our results define a molecular pathway that contributes to the maintenance of telomere length homeostasis in mESCs.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.celrep.2017.07.070DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5595351PMC
August 2017

DNA methylation of intragenic CpG islands depends on their transcriptional activity during differentiation and disease.

Proc Natl Acad Sci U S A 2017 09 21;114(36):E7526-E7535. Epub 2017 Aug 21.

Division of Medical Sciences and Graduate Entry Medicine, School of Medicine, University of Nottingham, Royal Derby Hospital, Derby DE22 3DT, United Kingdom;

The human genome contains ∼30,000 CpG islands (CGIs). While CGIs associated with promoters nearly always remain unmethylated, many of the ∼9,000 CGIs lying within gene bodies become methylated during development and differentiation. Both promoter and intragenic CGIs may also become abnormally methylated as a result of genome rearrangements and in malignancy. The epigenetic mechanisms by which some CGIs become methylated but others, in the same cell, remain unmethylated in these situations are poorly understood. Analyzing specific loci and using a genome-wide analysis, we show that transcription running across CGIs, associated with specific chromatin modifications, is required for DNA methyltransferase 3B (DNMT3B)-mediated DNA methylation of many naturally occurring intragenic CGIs. Importantly, we also show that a subgroup of intragenic CGIs is not sensitive to this process of transcription-mediated methylation and that this correlates with their individual intrinsic capacity to initiate transcription in vivo. We propose a general model of how transcription could act as a primary determinant of the patterns of CGI methylation in normal development and differentiation, and in human disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.1703087114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5594649PMC
September 2017

Correction for Yu et al., "A Mouse Null Allele Defines an Essential Role for Arginine Methylation in Genome Maintenance and Cell Proliferation".

Mol Cell Biol 2017 09 11;37(17). Epub 2017 Aug 11.

Terry Fox Molecular Oncology Group and Bloomfield Center for Research on Aging, Segal Cancer Centre, Lady Davis Institute for Medical Research, Sir Mortimer B. Davis Jewish General Hospital, and Departments of Oncology and Medicine, McGill University, Montréal, Québec H3T 1E2, Canada.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/MCB.00298-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5559678PMC
September 2017

Transcription and chromatin determinants of de novo DNA methylation timing in oocytes.

Epigenetics Chromatin 2017 12;10:25. Epub 2017 May 12.

Epigenetics Programme, Babraham Institute, Cambridge, CB22 3AT UK.

Background: Gametogenesis in mammals entails profound re-patterning of the epigenome. In the female germline, DNA methylation is acquired late in oogenesis from an essentially unmethylated baseline and is established largely as a consequence of transcription events. Molecular and functional studies have shown that imprinted genes become methylated at different times during oocyte growth; however, little is known about the kinetics of methylation gain genome wide and the reasons for asynchrony in methylation at imprinted loci.

Results: Given the predominant role of transcription, we sought to investigate whether transcription timing is rate limiting for de novo methylation and determines the asynchrony of methylation events. Therefore, we generated genome-wide methylation and transcriptome maps of size-selected, growing oocytes to capture the onset and progression of methylation. We find that most sequence elements, including most classes of transposable elements, acquire methylation at similar rates overall. However, methylation of CpG islands (CGIs) is delayed compared with the genome average and there are reproducible differences amongst CGIs in onset of methylation. Although more highly transcribed genes acquire methylation earlier, the major transitions in the oocyte transcriptome occur well before the de novo methylation phase, indicating that transcription is generally not rate limiting in conferring permissiveness to DNA methylation. Instead, CGI methylation timing negatively correlates with enrichment for histone 3 lysine 4 (H3K4) methylation and dependence on the H3K4 demethylases KDM1A and KDM1B, implicating chromatin remodelling as a major determinant of methylation timing. We also identified differential enrichment of transcription factor binding motifs in CGIs acquiring methylation early or late in oocyte growth. By combining these parameters into multiple regression models, we were able to account for about a fifth of the variation in methylation timing of CGIs. Finally, we show that establishment of non-CpG methylation, which is prevalent in fully grown oocytes, and methylation over non-transcribed regions, are later events in oogenesis.

Conclusions: These results do not support a major role for transcriptional transitions in the time of onset of DNA methylation in the oocyte, but suggest a model in which sequences least dependent on chromatin remodelling are the earliest to become permissive for methylation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13072-017-0133-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5429541PMC
July 2017

DNA methylation intratumor heterogeneity in localized lung adenocarcinomas.

Oncotarget 2017 Mar;8(13):21994-22002

Department of Thoracic/Head and Neck Medical Oncology, The University of Texas, MD Anderson Cancer Center, Houston, TX 77030, USA.

Cancers are composed of cells with distinct molecular and phenotypic features within a given tumor, a phenomenon termed intratumor heterogeneity (ITH). Previously, we have demonstrated genomic ITH in localized lung adenocarcinomas; however, the nature of methylation ITH in lung cancers has not been well investigated. In this study, we generated methylation profiles of 48 spatially separated tumor regions from 11 localized lung adenocarcinomas and their matched normal lung tissues using Illumina Infinium Human Methylation 450K BeadChip array. We observed methylation ITH within the same tumors, but to a much less extent compared to inter-individual heterogeneity. On average, 25% of all differentially methylated probes compared to matched normal lung tissues were shared by all regions from the same tumors. This is in contrast to somatic mutations, of which approximately 77% were shared events amongst all regions of individual tumors, suggesting that while the majority of somatic mutations were early clonal events, the tumor-specific DNA methylation might be associated with later branched evolution of these 11 tumors. Furthermore, our data showed that a higher extent of DNA methylation ITH was associated with larger tumor size (average Euclidean distance of 35.64 (> 3cm, median size) versus 27.24 (<= 3cm), p = 0.014), advanced age (average Euclidean distance of 34.95 (above 65) verse 28.06 (below 65), p = 0.046) and increased risk of postsurgical recurrence (average Euclidean distance of 35.65 (relapsed patients) versus 29.03 (patients without relapsed), p = 0.039).
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.18632/oncotarget.15777DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5400640PMC
March 2017

PRMT5 C-terminal Phosphorylation Modulates a 14-3-3/PDZ Interaction Switch.

J Biol Chem 2017 02 28;292(6):2255-2265. Epub 2016 Dec 28.

From the Department of Epigenetics and Molecular Carcinogenesis, University of Texas M.D. Anderson Cancer Center, Smithville, Texas 78957,

PRMT5 is the primary enzyme responsible for the deposition of the symmetric dimethylarginine in mammalian cells. In an effort to understand how PRMT5 is regulated, we identified a threonine phosphorylation site within a C-terminal tail motif, which is targeted by the Akt/serum- and glucocorticoid-inducible kinases. While investigating the function of this posttranslational modification, we serendipitously discovered that its free C-terminal tail binds PDZ domains (when unphosphorylated) and 14-3-3 proteins (when phosphorylated). In essence, a phosphorylation event within the last few residues of the C-terminal tail generates a posttranslational modification-dependent PDZ/14-3-3 interaction "switch." The C-terminal motif of PRMT5 is required for plasma membrane association, and loss of this switching capacity is not compatible with life. This signaling phenomenon was recently reported for the HPV E6 oncoprotein but has not yet been observed for mammalian proteins. To investigate the prevalence of PDZ/14-3-3 switching in signal transduction, we built a protein domain microarray that harbors PDZ domains and 14-3-3 proteins. We have used this microarray to interrogate the C-terminal tails of a small group of candidate proteins and identified ERBB4, PGHS2, and IRK1 (as well as E6 and PRMT5) as conforming to this signaling mode, suggesting that PDZ/14-3-3 switching may be a broad biological paradigm.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/jbc.M116.760330DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5313098PMC
February 2017

Genetic Studies on Mammalian DNA Methyltransferases.

Adv Exp Med Biol 2016 ;945:123-150

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, 1808 Park Road 1C, Smithville, TX, 78957, USA.

Cytosine methylation at the C5-position, generating 5-methylcytosine (5mC), is a DNA modification found in many eukaryotic organisms, including fungi, plants, invertebrates, and vertebrates, albeit its levels vary greatly in different organisms. In mammals, cytosine methylation occurs predominantly in the context of CpG dinucleotides, with the majority (60-80 %) of CpG sites in their genomes being methylated. DNA methylation plays crucial roles in the regulation of chromatin structure and gene expression and is essential for mammalian development. Aberrant changes in DNA methylation levels and patterns are associated with various human diseases, including cancer and developmental disorders. DNA methylation is mediated by three active DNA methyltransferases (Dnmts), namely, Dnmt1, Dnmt3a, and Dnmt3b, in mammals. Over the last two decades, genetic manipulations of these enzymes, as well as their regulators, in mice have greatly contributed to our understanding of the biological functions of DNA methylation in mammals. In this chapter, we discuss genetic studies on mammalian Dnmts, focusing on their roles in embryogenesis, cellular differentiation, genomic imprinting, and X-chromosome inactivation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-3-319-43624-1_6DOI Listing
June 2017

Histone Demethylase LSD1 Promotes Adipocyte Differentiation through Repressing Wnt Signaling.

Cell Chem Biol 2016 Oct 15;23(10):1228-1240. Epub 2016 Sep 15.

China Novartis Institutes for BioMedical Research, 4218 Jinke Road, Zhangjiang Hi-Tech Park, Pudong New Area, Shanghai 201203, China. Electronic address:

Adipose tissue plays important roles in animals. White fat stores energy in lipids, while brown fat is responsible for nonshivering thermogenesis through UCP1-mediated energy dissipation. Although epigenetic mechanisms modulate differentiation in multiple lineages, the epigenetic regulation of brown adipocyte differentiation is poorly understood. By screening a collection of epigenetic compounds, we found that Lysine-Specific Demethylase 1 (LSD1) inhibitors repress brown adipocyte differentiation. RNAi-mediated Lsd1 knockdown causes a similar effect, which can be rescued by expression of wild-type but not catalytic-inactive LSD1. Mechanistically, LSD1 promotes brown adipogenesis by demethylating H3K4 on promoter regions of Wnt signaling components and repressing the Wnt pathway. Furthermore, deletion of Lsd1 in mice leads to inhibition of brown adipogenesis, validating the pivotal role of LSD1 in brown fat development in vivo. Our work identifies LSD1 as a key epigenetic regulator in brown adipogenesis. The link between LSD1 and the Wnt pathway provides potential opportunities to modulate brown fat differentiation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.chembiol.2016.08.010DOI Listing
October 2016

Negative regulation of DNMT3A de novo DNA methylation by frequently overexpressed UHRF family proteins as a mechanism for widespread DNA hypomethylation in cancer.

Cell Discov 2016 12;2:16007. Epub 2016 Apr 12.

Shanghai Key Laboratory of Regulatory Biology, the Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China; Collaborative Innovation Center for Cancer Medicine, Sun Yat-Sen University Cancer Center, Guangzhou, China.

Global DNA hypomethylation is a most common epigenetic alteration in cancer, but the mechanism remains elusive. Previous studies demonstrate that UHRF1 but not UHRF2 is required for mediating DNA maintenance methylation by DNMT1. Here we report unexpectedly a conserved function for UHRF1 and UHRF2: inhibiting de novo DNA methylation by functioning as E3 ligases promoting DNMT3A degradation. UHRF1/2 are frequently overexpressed in cancers and we present evidence that UHRF1/2 overexpression downregulates DNMT3A proteins and consequently leads to DNA hypomethylation. Abrogating this negative regulation on DNMT3A or overexpression of DNMT3A leads to increased DNA methylation and impaired tumor growth. We propose a working model that UHRF1/2 safeguards the fidelity of DNA methylation and suggests that UHRF1/2 overexpression is likely a causal factor for widespread DNA hypomethylation in cancer via suppressing DNMT3A.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/celldisc.2016.7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4849474PMC
July 2016

Maternal Setdb1 Is Required for Meiotic Progression and Preimplantation Development in Mouse.

PLoS Genet 2016 Apr 12;12(4):e1005970. Epub 2016 Apr 12.

Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Smithville, Texas, United States of America.

Oocyte meiotic progression and maternal-to-zygote transition are accompanied by dynamic epigenetic changes. The functional significance of these changes and the key epigenetic regulators involved are largely unknown. Here we show that Setdb1, a lysine methyltransferase, controls the global level of histone H3 lysine 9 di-methyl (H3K9me2) mark in growing oocytes. Conditional deletion of Setdb1 in developing oocytes leads to meiotic arrest at the germinal vesicle and meiosis I stages, resulting in substantially fewer mature eggs. Embryos derived from these eggs exhibit severe defects in cell cycle progression, progressive delays in preimplantation development, and degeneration before reaching the blastocyst stage. Rescue experiments by expressing wild-type or inactive Setdb1 in Setdb1-deficient oocytes suggest that the catalytic activity of Setdb1 is essential for meiotic progression and early embryogenesis. Mechanistically, up-regulation of Cdc14b, a dual-specificity phosphatase that inhibits meiotic progression, greatly contributes to the meiotic arrest phenotype. Setdb1 deficiency also leads to derepression of transposons and increased DNA damage in oocytes, which likely also contribute to meiotic defects. Thus, Setdb1 is a maternal-effect gene that controls meiotic progression and is essential for early embryogenesis. Our results uncover an important link between the epigenetic machinery and the major signaling pathway governing meiotic progression.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.pgen.1005970DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4829257PMC
April 2016