Characterizing DNA Methyltransferases With An Ultrasensitive Luciferase-Linked Continuous Assay

Abstract Strong evidence suggests that genetic variations in DNA methyltransferases (DNMTs) may alter the downstream expression and DNA methylation patterns of neuronal genes and influence cognition. This study investigates the association between a DNMT1 polymorphism, rs2162560, and chemotherapy-associated cognitive impairment (CACI) in a cohort of breast cancer patients. This is a prospective, longitudinal cohort study. From 2011 to 2017, 351 early-stage breast cancer patients receiving chemotherapy were assessed at baseline, the midpoint, and the end of chemotherapy. DNA was extracted from whole blood, and genotyping was performed using Sanger sequencing. Patients’ self-perceived cognitive function and cognitive performance were assessed at three different time points using FACT-Cog (v.3) and a neuropsychological battery, respectively. The association between DNMT1 rs2162560 and cognitive function was evaluated using logistic regression analyses. Overall, 33.3% of the patients reported impairment relative to baseline in one or more cognitive domains. Cognitive impairment was observed in various objective cognitive domains, with incidences ranging from 7.2% to 36.9%. The DNMT1 rs2162560 A allele was observed in 21.8% of patients and this was associated with lower odds of self-reported cognitive decline in the concentration (OR = 0.45, 95% CI: 0.25–0.82, P = 0.01) and functional interference (OR = 0.48, 95% CI: 0.24–0.95, P = 0.03) domains. No significant association was observed between DNMT1 rs2162560 and objective cognitive impairment. This is the first study to show a significant association between the DNMT1 rs2162560 polymorphism and CACI. Our data suggest that epigenetic processes could contribute to CACI, and further studies are needed to validate these findings.

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Neuroblastoma and the epigenome

Cancer and Metastasis Reviews ◽

10.1007/s10555-020-09946-y ◽

2021 ◽

Author(s):

Irfete S. Fetahu ◽

Sabine Taschner-Mandl

Keyword(s):

Dna Methylation ◽

Histone Modifications ◽

Disease Diagnosis ◽

Dna Methyltransferases ◽

Epigenetic Alterations ◽

Aberrant Expression ◽

Tet Proteins ◽

Relative Paucity ◽

Underlying Causes ◽

Aberrant Dna Methylation

AbstractNeuroblastoma (NB) is a pediatric cancer of the sympathetic nervous system and one of the most common solid tumors in infancy. Amplification of MYCN, copy number alterations, numerical and segmental chromosomal aberrations, mutations, and rearrangements on a handful of genes, such as ALK, ATRX, TP53, RAS/MAPK pathway genes, and TERT, are attributed as underlying causes that give rise to NB. However, the heterogeneous nature of the disease—along with the relative paucity of recurrent somatic mutations—reinforces the need to understand the interplay of genetic factors and epigenetic alterations in the context of NB. Epigenetic mechanisms tightly control gene expression, embryogenesis, imprinting, chromosomal stability, and tumorigenesis, thereby playing a pivotal role in physio- and pathological settings. The main epigenetic alterations include aberrant DNA methylation, disrupted patterns of posttranslational histone modifications, alterations in chromatin composition and/or architecture, and aberrant expression of non-coding RNAs. DNA methylation and demethylation are mediated by DNA methyltransferases (DNMTs) and ten-eleven translocation (TET) proteins, respectively, while histone modifications are coordinated by histone acetyltransferases and deacetylases (HATs, HDACs), and histone methyltransferases and demethylases (HMTs, HDMs). This article focuses predominately on the crosstalk between the epigenome and NB, and the implications it has on disease diagnosis and treatment.

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The Temporal Order of DNA Replication Shaped by Mammalian DNA Methyltransferases

Cells ◽

10.3390/cells10020266 ◽

2021 ◽

Vol 10 (2) ◽

pp. 266

Author(s):

Shin-ichiro Takebayashi ◽

Tyrone Ryba ◽

Kelsey Wimbish ◽

Takuya Hayakawa ◽

Morito Sakaue ◽

...

Keyword(s):

Dna Replication ◽

Temporal Order ◽

Expression System ◽

Embryonic Stem ◽

Replication Timing ◽

Dna Methyltransferases ◽

Dna Hypomethylation ◽

Transcriptional Changes ◽

A Cell ◽

Genomic Regions

Multiple epigenetic pathways underlie the temporal order of DNA replication (replication timing) in the contexts of development and disease. DNA methylation by DNA methyltransferases (Dnmts) and downstream chromatin reorganization and transcriptional changes are thought to impact DNA replication, yet this remains to be comprehensively tested. Using cell-based and genome-wide approaches to measure replication timing, we identified a number of genomic regions undergoing subtle but reproducible replication timing changes in various Dnmt-mutant mouse embryonic stem (ES) cell lines that included a cell line with a drug-inducible Dnmt3a2 expression system. Replication timing within pericentromeric heterochromatin (PH) was shown to be correlated with redistribution of H3K27me3 induced by DNA hypomethylation: Later replicating PH coincided with H3K27me3-enriched regions. In contrast, this relationship with H3K27me3 was not evident within chromosomal arm regions undergoing either early-to-late (EtoL) or late-to-early (LtoE) switching of replication timing upon loss of the Dnmts. Interestingly, Dnmt-sensitive transcriptional up- and downregulation frequently coincided with earlier and later shifts in replication timing of the chromosomal arm regions, respectively. Our study revealed the previously unrecognized complex and diverse effects of the Dnmts loss on the mammalian DNA replication landscape.

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Expanding the Structural Diversity of DNA Methyltransferase Inhibitors

Pharmaceuticals ◽

10.3390/ph14010017 ◽

2020 ◽

Vol 14 (1) ◽

pp. 17

Author(s):

K. Eurídice Juárez-Mercado ◽

Fernando D. Prieto-Martínez ◽

Norberto Sánchez-Cruz ◽

Andrea Peña-Castillo ◽

Diego Prada-Gracia ◽

...

Keyword(s):

Histone Deacetylases ◽

Dna Methyltransferase ◽

Structural Diversity ◽

Dna Methyltransferases ◽

Dna Methyltransferase Inhibitors ◽

Epigenetic Drug ◽

Ic50 Values ◽

Dietary Component ◽

Approved Drugs ◽

Epigenetic Processes

Inhibitors of DNA methyltransferases (DNMTs) are attractive compounds for epigenetic drug discovery. They are also chemical tools to understand the biochemistry of epigenetic processes. Herein, we report five distinct inhibitors of DNMT1 characterized in enzymatic inhibition assays that did not show activity with DNMT3B. It was concluded that the dietary component theaflavin is an inhibitor of DNMT1. Two additional novel inhibitors of DNMT1 are the approved drugs glyburide and panobinostat. The DNMT1 enzymatic inhibitory activity of panobinostat, a known pan inhibitor of histone deacetylases, agrees with experimental reports of its ability to reduce DNMT1 activity in liver cancer cell lines. Molecular docking of the active compounds with DNMT1, and re-scoring with the recently developed extended connectivity interaction features approach, led to an excellent agreement between the experimental IC50 values and docking scores.

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Epigenetic mechanisms involved in intrauterine growth restriction and aberrant kidney development and function

Journal of Developmental Origins of Health and Disease ◽

10.1017/s2040174420001257 ◽

2020 ◽

pp. 1-11

Author(s):

Thu N. A. Doan ◽

Jessica F. Briffa ◽

Aaron L. Phillips ◽

Shalem Y. Leemaqz ◽

Rachel A. Burton ◽

...

Keyword(s):

Kidney Disease ◽

Intrauterine Growth Restriction ◽

Kidney Development ◽

Dna Methyltransferases ◽

Growth Restriction ◽

Epigenetic Mechanisms ◽

Specific Expression ◽

Maternal Line ◽

Intrauterine Growth ◽

Increased Risk

Abstract Intrauterine growth restriction (IUGR) due to uteroplacental insufficiency results in a placenta that is unable to provide adequate nutrients and oxygen to the fetus. These growth-restricted babies have an increased risk of hypertension and chronic kidney disease later in life. In rats, both male and female growth-restricted offspring have nephron deficits but only males develop kidney dysfunction and high blood pressure. In addition, there is transgenerational transmission of nephron deficits and hypertension risk. Therefore, epigenetic mechanisms may explain the sex-specific programming and multigenerational transmission of IUGR-related phenotypes. Expression of DNA methyltransferases (Dnmt1and Dnmt3a) and imprinted genes (Peg3, Snrpn, Kcnq1, and Cdkn1c) were investigated in kidney tissues of sham and IUGR rats in F1 (embryonic day 20 (E20) and postnatal day 1 (PN1)) and F2 (6 and 12 months of age, paternal and maternal lines) generations (n = 6–13/group). In comparison to sham offspring, F1 IUGR rats had a 19% decrease in Dnmt3a expression at E20 (P < 0.05), with decreased Cdkn1c (19%, P < 0.05) and increased Kcnq1 (1.6-fold, P < 0.01) at PN1. There was a sex-specific difference in Cdkn1c and Snrpn expression at E20, with 29% and 34% higher expression in IUGR males compared to females, respectively (P < 0.05). Peg3 sex-specific expression was lost in the F2 IUGR offspring, only in the maternal line. These findings suggest that epigenetic mechanisms may be altered in renal embryonic and/or fetal development in growth-restricted offspring, which could alter kidney function, predisposing these offspring to kidney disease later in life.

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Interplay between Histone and DNA Methylation Seen through Comparative Methylomes in Rare Mendelian Disorders

International Journal of Molecular Sciences ◽

10.3390/ijms22073735 ◽

2021 ◽

Vol 22 (7) ◽

pp. 3735

Author(s):

Guillaume Velasco ◽

Damien Ulveling ◽

Sophie Rondeau ◽

Pauline Marzin ◽

Motoko Unoki ◽

...

Keyword(s):

Dna Methylation ◽

Catalytic Domain ◽

De Novo ◽

Mutation Screening ◽

Histone H3 ◽

Regulatory Elements ◽

Germline Mutations ◽

Dna Methyltransferases ◽

Mendelian Disorders ◽

Epigenetic Machinery

DNA methylation (DNAme) profiling is used to establish specific biomarkers to improve the diagnosis of patients with inherited neurodevelopmental disorders and to guide mutation screening. In the specific case of mendelian disorders of the epigenetic machinery, it also provides the basis to infer mechanistic aspects with regard to DNAme determinants and interplay between histone and DNAme that apply to humans. Here, we present comparative methylomes from patients with mutations in the de novo DNA methyltransferases DNMT3A and DNMT3B, in their catalytic domain or their N-terminal parts involved in reading histone methylation, or in histone H3 lysine (K) methylases NSD1 or SETD2 (H3 K36) or KMT2D/MLL2 (H3 K4). We provide disease-specific DNAme signatures and document the distinct consequences of mutations in enzymes with very similar or intertwined functions, including at repeated sequences and imprinted loci. We found that KMT2D and SETD2 germline mutations have little impact on DNAme profiles. In contrast, the overlapping DNAme alterations downstream of NSD1 or DNMT3 mutations underlines functional links, more specifically between NSD1 and DNMT3B at heterochromatin regions or DNMT3A at regulatory elements. Together, these data indicate certain discrepancy with the mechanisms described in animal models or the existence of redundant or complementary functions unforeseen in humans.

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DNA Methyltransferase 3A Is Involved in the Sustained Effects of Chronic Stress on Synaptic Functions and Behaviors

Cerebral Cortex ◽

10.1093/cercor/bhaa337 ◽

2020 ◽

Author(s):

Jing Wei ◽

Jia Cheng ◽

Nicholas J Waddell ◽

Zi-Jun Wang ◽

Xiaodong Pang ◽

...

Keyword(s):

Dna Methylation ◽

Dna Methyltransferase ◽

Epigenetic Modification ◽

Substantial Reduction ◽

Dna Methyltransferases ◽

Male Rats ◽

Neural Pathways ◽

Dnmt Inhibitors ◽

Synaptic Functions

Abstract Emerging evidence suggests that epigenetic mechanisms regulate aberrant gene transcription in stress-associated mental disorders. However, it remains to be elucidated about the role of DNA methylation and its catalyzing enzymes, DNA methyltransferases (DNMTs), in this process. Here, we found that male rats exposed to chronic (2-week) unpredictable stress exhibited a substantial reduction of Dnmt3a after stress cessation in the prefrontal cortex (PFC), a key target region of stress. Treatment of unstressed control rats with DNMT inhibitors recapitulated the effect of chronic unpredictable stress on decreased AMPAR expression and function in PFC. In contrast, overexpression of Dnmt3a in PFC of stressed animals prevented the loss of glutamatergic responses. Moreover, the stress-induced behavioral abnormalities, including the impaired recognition memory, heightened aggression, and hyperlocomotion, were partially attenuated by Dnmt3a expression in PFC of stressed animals. Finally, we found that there were genome-wide DNA methylation changes and transcriptome alterations in PFC of stressed rats, both of which were enriched at several neural pathways, including glutamatergic synapse and microtubule-associated protein kinase signaling. These results have therefore recognized the potential role of DNA epigenetic modification in stress-induced disturbance of synaptic functions and cognitive and emotional processes.

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