Methyl Donor Research Articles

S-adenosyl-L-methionine is the unexpected methyl donor for the methylation of mercury by the membrane-associated HgcAB complex

Mercury (Hg) is a heavy metal that exhibits high biological toxicity. Monomethylmercury and dimethylmercury are neurotoxins and a significant environmental concern as they bioaccumulate and biomagnify within the aquatic food web. Microbial Hg methylation involves two proteins, HgcA and HgcB. Here, we show that HgcA and HgcB can be heterologously coexpressed, and the HgcAB complex can be purified. We demonstrated that HgcA is a membrane-associated cobalamin-dependent methyltransferase and HgcB is a ferredoxin-like protein containing two [4Fe-4S] clusters. Further, spectroscopic and kinetic results demonstrate that S-adenosyl-L-methionine (SAM) donates the methyl group to Hg in a two-step reaction involving a methylcob(III)alamin intermediate including Co-thiolate ligation from a conserved Cys residue. Our findings uncover a biological role for SAM in microbial Hg methylation.

Trends in dietary choline and betaine intake among Chinese adults: the China Health and Nutrition Survey 1991-2011.

Choline and its derivative betaine are important methyl donors, components of cell membrane phospholipids, or precursors of the neurotransmitter acetylcholine and the gut microbial metabolite trimethylamine-N-oxide. We aimed to investigate trends in dietary intake and food sources of total choline, individual choline forms, and betaine in Chinese adults using data from the China Health and Nutrition Survey (CHNS) 1991-2011. The sample was drawn from urban and rural communities in 12 provinces or autonomous regions with a multistage, random cluster design. Dietary intake was estimated using three consecutive 24-hour dietary recalls in combination with a household food inventory. Linear mixed-effect models were constructed to calculate adjusted mean intake values and 95% confidence intervals (CIs) using R version 4.2.2. A total of 11,188 men and 12,279 women aged 18 years or older were included. Between 1991 and 2011, total choline intake increased from 219.3 (95% CI: 215.1-223.4) mg/d to 269.0 (95% CI: 265.6-272.5) mg/d in men and from 195.6 (95% CI: 191.8-199.4) mg/d to 240.4 (95% CI: 237.4-243.5) mg/d in women (both P-trends <0.001). Phosphatidylcholine was the major form of dietary choline and its contribution to total choline increased from 46.9% in 1991 to 58.8% in 2011. Cereals were the primary food source of total choline before 2000 while eggs had ranked at the top since 2004. Dietary betaine intake was relatively steady over time with a range of 134.0-151.5 mg/d in men (P-trend <0.001) and 111.7-125.3 mg/d in women (P-trend >0.05). In conclusion, Chinese adults experienced a significant increase in dietary intake of choline, particularly phosphatidylcholine during 1991-2011 and animal-derived foods have replaced plant-based foods as the main food sources of choline. Betaine intake remained relatively stable over time. Future efforts should address the health effects of these changes.

Prenatal choline supplementation enhances metabolic outcomes with differential impact on DNA methylation in Wistar rat offspring and dams.

Choline is an essential nutrient required for proper functioning of organs and serves as a methyl donor. In liver where choline metabolism primarily occurs, glucose homeostasis is regulated through insulin receptor substrates (IRS) 1 and 2. Here, we determined the role of prenatal choline as a modulator of metabolic health and DNA methylation in liver of offspring and dams. Pregnant Wistar rat dams were fed an AIN-93G diet and received drinking water either with supplemented 0.25% choline (w/w) as choline bitartrate or untreated control. All offspring were weaned to a high-fat diet for 12 weeks. Prenatal choline supplementation led to higher insulin sensitivity in female offspring at weaning as well as lower body weight and food intake and higher insulin sensitivity in female and male adult offspring compared to offspring from untreated dams. Higher hepatic betaine concentrations were observed in dams and female offspring of choline-supplemented dams at weaning and higher glycerophosphocholine in female and male offspring at post-weaning compared to the untreated control, suggestive of sustaining different choline pathways. Hepatic gene expression of Irs2 was higher in dams at weaning and female offspring at weaning and post-weaning, whereas Irs1 was lower in male offspring at post-weaning. Gene-specific DNA methylation of Irs2 was lower in female offspring at post-weaning and Irs1 methylation was higher in male offspring at post-weaning that exhibited an inverse relationship between methylation and gene expression. In conclusion, prenatal choline supplementation contributes to improved parameters of insulin signaling but these effects varied across time and offspring sex.

Purification and catalysis of choline dehydrogenase from Escherichia coli

Choline dehydrogenase (CHDH) is a membrane-bound enzyme belonging to the glucose-methanol-choline (GMC) oxidoreductase superfamily, which is characterized by a crucial FAD-binding domain essential for catalytic function. CHDH catalyzes the oxidation of choline to betaine aldehyde, which is further oxidized to betaine, a vital osmoprotectant and methyl donor for cellular physiology and metabolism. However, the detailed catalytic mechanism of CHDH still remains poorly understood. In our investigation, we gained purity E. coli CHDH samples in DDM (n-dodecyl-β-D-maltoside) and SMA (styrene maleic acid) copolymer respectively and examined their structural composition and catalytic activity separately. Our findings demonstrated the effectiveness of SMA, commonly employed for extracting transmembrane proteins and can preserve the natural bio-membrane environment surrounding the enzyme, in extracting peripheral membrane proteins like CHDH here, which lacks transmembrane helices. CHDH exhibited a trimeric conformation in SMA, whereas it existed as monomers in DDM, as determined by our negative staining analysis. Our experiments also revealed that highly pure E. coli CHDH could only oxidize choline to betaine aldehyde but failed to further oxidize betaine aldehyde to betaine as determined by the biochemical and enzymatic reaction kinetic assays. In addition, the enzyme in SMA displayed greater catalytic activity compared to that in DDM. Furthermore, we confirmed the crucial role of His473, which is hypothesized to be a critical site for substrate binding from our structural comparative analysis between CHDH and its highly homologous choline oxidase, in the catalytic activity of the enzyme through gene mutation. Our work also sheds light on CHDH’s contribution to cellular osmotic tolerance through gene knockout. This research enhances our better understanding of CHDH within cellular biochemistry and metabolic pathways.

Methyl Donor Micronutrients Orchestrate Lipid Metabolism: The Role of DNA Methylation Modification

DNA methylation, a key epigenetic modification, plays a crucial role in regulating lipid metabolism. Consistent correlations have been observed between aberrant DNA methylation patterns and lipid metabolic disorders. Emerging evidence indicates that methyl donor micronutrients could influence DNA methylation patterns, consequently exerting an influence on lipid metabolism. Specifically, the deficiency or excesses of methyl donor micronutrients (folate, choline, betaine, B vitamins and methionine) have been associated with altered DNA methylation patterns linked to lipid metabolism. These alteration in DNA methylation levels, occurring globally and within promoter regions, could affect gene expression related to lipid metabolism. However, the mechanisms through which methyl donor micronutrients regulate lipid metabolism via the DNA methylation modification and the role of methyl donor micronutrients supplementation on DNA methylation profiles remain unclear. In this review, we summarized the regulatory role of DNA methylation in lipid metabolism, and highlighted recent findings investigating the impact of methyl donor micronutrients on lipid metabolism, as well as DNA methylation-mediated adipogenesis and adipose deposition. Taken together, this review deepened our understanding of how the complex interplay between methyl donor micronutrients, DNA methylation, and lipid metabolism, and provides valuable information for accurately regulating lipid metabolism of livestock and poultry, thereby improving meat quality, and promoting the development of animal husbandry.

Practical supplements for prevention and management of migraine attacks: a narrative review.

Migraine is one of the most debilitating neurological disorders that causes frequent attacks of headaches and affects approximately 11% of the global population. Deficient or even insufficient levels of vital nutrients would increase the severity and frequency of migraine attacks. Therefore, we aimed to examine the practical supplements for the prevention and management of migraine attacks. This narrative review study was conducted by searching PubMed, ISI web of science, EMBASE, Google Scholar, and Scopus using the keywords of "dietary supplement" and "migraine" plus their MeSH terms. Original articles published in English language from their inception to July 27th, 2024, studies that investigated adult population (aged >18 years), and those assessing the impact of intended nutrient supplementation on clinical symptoms of migraine were included in the study. Oxidative stress and low intake of antioxidants would be risk factors for migraine attacks by inducing inflammation. The secretion of inflammatory cytokines, such as tumor necrosis factor (TNF)-a, would lead to neuroinflammation and migraine episodes by increasing the cellular permeability and interactions. Evidence also indicated a direct association between phases of migraine attacks and calcitonin gene-related peptide (CGRP), mitochondrial disorders, monoaminergic pathway, disruption in brain energy metabolism, and higher serum levels of glutamate and homocysteine. Therefore, supplementation with nutrients involved in mitochondrial function, brain energy metabolism, and even methyl donors would relieve migraine attacks. Evidence indicated that supplementation with riboflavin, omega-3 fatty acids, alpha lipoic acid, magnesium, probiotics, coenzyme Q10, ginger, and caffeine would have favorable effects on migraine patients. However, more prospective studies are required to evaluate the effect of other nutrients on migraine patients.

The epigenetic impact of daily diet food choices on human health and chronic diseases

Nutrition, certain lifestyle behaviors (smoking, drug, alcohol addictions, etc.) and environment all contribute to cancer and other lines development. Similarly, epigenetic pathways are known to occur at the intersection between the generally reversible effects of lifestyle or ecological factors and the irreversible alterations that explain numerous diseases. Chemoprevention is the process of intervening in the epigenome to mitigate the detrimental effects of environmental factors or certain lifestyles before they lead to significant consequences. DNA is permanently exposed to various substances modifying both its genetic and epigenetic configuration. It’s controlled by various agents (methyl donors, sophisticated enzymes, etc.). Pesticides, artificial food additives, drugs and environmental toxins penetrating the placenta, can result in developmental program alterations and epigenetic changes in a fetus. In nutrition context, the research revealed that several foods can alter epigenetic markers. Some vitamins may alter DNA methylation patterns and histone alterations, possibly influencing gene expression and disease risk. Thus, dietary treatments may provide an epigenetic regulation, emphasizing nutrition role in preserving health and preventing illness via epigenetics Keywords: human epigenetics, epigenetic regulation of diseases, daily food ration, healthy food

Discovery of a novel methionine biosynthetic route via O-phospho-l-homoserine.

Methionine (Met), a sulfur-containing amino acid, is essential for the underlying biological processes in living organisms. In addition to its importance as a starting building block for peptide chain elongation in protein biosynthesis, Met is a direct precursor of S-adenosyl-l-methionine, an indispensable methyl donor molecule in primary and secondary metabolism. Streptomyces bacteria are well known to produce diverse secondary metabolites, but many strains lack canonical Met pathway genes for l-homocysteine, a direct precursor of Met in bacteria, plants, and archaea. Here, we report the identification of a novel gene (metM) responsible for the Met biosynthesis in Streptomyces strains and demonstrate the catalytic function of the gene product, MetM. We further identified the metO gene, a downstream gene of metM, and showed that it encodes a sulfur-carrier protein (SCP). In in vitro analysis, MetO was found to play an important role in a sulfur donor by forming a thiocarboxylated SCP. Together with MetO (thiocarboxylate), MetM directly converted O-phospho-l-homoserine to l-homocysteine. O-Phospho-l-homoserine is also known as an intermediate for threonine biosynthesis in bacteria and plants, and MetM shares sequence homology with threonine synthase. Our findings thus revealed that MetM seizes O-phospho-l-homoserine from the threonine biosynthetic pathway and uses it as an intermediate of the Met biosynthesis to generate the sulfur-containing amino acid. Importantly, this MetM/MetO pathway is highly conserved in Streptomyces bacteria and distributed in other bacteria and archaea.IMPORTANCEMethionine (Met) is a sulfur-containing proteinogenic amino acid. Moreover, Met is a direct precursor of S-adenosyl-l-methionine, an indispensable molecule for expanding the structural diversity of natural products. Because Met and its derivatives benefit humans, the knowledge of Met biosynthesis is important as a basis for improving their fermentation. Streptomyces bacteria are well known to produce diverse and valuable natural products, but many strains lack canonical Met pathway genes. Here, we identified a novel l-homocysteine synthase (MetM) in Streptomyces and demonstrated that it converts O-phospho-L-homoserine to l-homocysteine using a thiocarboxylated sulfur-carrier protein as a sulfur donor. Since the metM is distributed in other bacteria and archaea, our pioneering study contributes to understanding Met biosynthesis in these organisms.

Ethionine-induced S-adenosylmethionine deficiency suppressed H3K27me3 and cell differentiation during neural tube development in mice.

S-adenosylmethionine (SAM) as a major methyl donor plays a key role in methylation modification in vivo, and its disorder was closely related to neural tube defects (NTDs). However, the exact mechanism between SAM deficiency and NTDs remained unclearly. Hence, we investigated the association between histone methylation modification and cell differentiation in NTDs mice induced by SAM deficiency. The levels of SAM and SAH (S-adenosylhomocysteine) were determined by enzyme linked immunosorbent assay (ELISA). The level of histone methylation, β-catenin were analyzed by Western blot, reversing transcription and quantitative PCR (RT-qPCR) and immunofluorescence. The results showed that the incidence rate of NTDs induced by ethionine were 46.2%. Post treatment of ethionine combined with SAM, the incidence rate of NTDs was reduced to 26.2%. The level of SAM was significantly decreased (p < 0.05) and a reduction in the SAM/SAH ratio was observed after entionine treatment. The SAM deficiency caused the reduction of H3K27me3 modifications and the elevated UTX activity (p < 0.05), and inhibited the expressions of β-catenin. The differentiations of NSCs into neurons and oligodendrocytes were inhibited under SAM deficiency (p < 0.05). These results indicated that the SAM deficiency led to reduce H3K27me3 modifications, prevented the β-catenin signaling pathway and NSCs differentiation, which provided an understanding of the novel function of epigenetic regulation in NTDs.

Mammalian D-Cysteine controls insulin secretion in the pancreas

D-amino acids are being recognized as important molecules in mammals with function. This is a first identification of endogenous D-cysteine in mammalian pancreas. D-cysteine is synthesized by serine racemase (SR) and SR−/− mice produce 6–10 fold higher levels of insulin in the pancreas and plasma including higher glycogen and ketone bodies in the liver. The excess insulin is stored as amyloid in secretory vesicles and exosomes. In glucose stimulated insulin secretion in mouse and human islets, equimolar amount of D-cysteine showed higher inhibition of insulin secretion compared to D-serine, another closely related stereoisomer synthesized by SR. In mouse models of diabetes (Streptozotocin (STZ) and Non Obese Diabetes (NOD) and human pancreas, the diabetic state showed increased expression of D-cysteine compared to D-serine followed by increased expression of SR. SR−/− mice show decreased cAMP in the pancreas, lower DNA methyltransferase enzymatic and promoter activities followed by reduced phosphorylation of CREB (S133), resulting in decreased methylation of the Ins1 promoter. D-cysteine is efficiently metabolized by D-amino acid oxidase and transported by ASCT2 and Asc1. Dietary supplementation with methyl donors restored the high insulin levels and low DNMT enzymatic activity in SR−/− mice. Our data show that endogenous D-cysteine in the mammalian pancreas is a regulator of insulin secretion.

Methyl-donor supplementation in women with systemic lupus erythematosus with different nutritional status: the protocol for a randomised, double-blind, placebo-controlled trial

IntroductionDNA hypomethylation in patients with systemic lupus erythematosus (SLE) has been recently documented in the literature. Low levels of DNA methylation have been observed globally and in genes associated with...

Regulatory role of microcystin in the response of microcystin-producing cyanobacteria to elevated CO2: Insights from metabolic profiling

The regulatory role of microcystin in the response of microcystin-producing cyanobacteria to elevated CO2 remains poorly understood. To address this gap, this study compared the responses of wild-type toxic Microcystis PCC 7806 and its mcyB-knockout mutant to elevated CO2 using metabolomic profiling under nitrogen (N)-rich and N-poor conditions. Under N-poor conditions, elevated CO2 promoted carbohydrate synthesis and tricarboxylic acid cycle in both strains, without affecting their growth. Under N-rich conditions, both strains exhibited increased biomass with rising CO2 levels, attributed to enhanced carbohydrate synthesis, tricarboxylic acid cycle, glutamate-glutamine cycle, purine synthesis, and arginine synthesis. However, compared to the mutant, the proliferation of wild-type toxic Microcystis was less stimulated by elevated CO2. The difference was associated with its reduced activity in the pentose phosphate pathway, likely linked to microcystin synthesis. Besides, the difference was related to higher utilization of glutamate, arginine, and aspartate due to increased microcystin production, indicating the regulatory role of microcystin in the response of microcystin-producing cyanobacteria to elevated CO2. Importantly, elevated CO2 could enhance microcystin synthesis by promoting the production of carbon backbones (malonyl CoA), amino acids (including arginine, glutamate and aspartate) and methyl donors (S-adenosylmethionine) of the wild-type toxic Microcystis PCC 7806. Notably, sufficient nitrogen sources were required for increased amino acid and methyl donors synthesis at high CO2 concentration. The discovery revealed underlying mechanisms behind the potential for elevated CO2 levels to increase toxicity risk associated with Microcystis blooms.

MAT2B regulates the protein level of MAT2A to preserve RNA N6-methyladenosine

MAT2B works together with MAT2A to synthesize S-Adenosyl methionine (SAM) as the primary methyl donor. MAT2B, despite lacking catalytic activity, exerts regulatory control over the enzymatic activity of MAT2A. In addition to the enzymatic activity regulation, we find that, in an NADP+-dependent manner, MAT2B binds and stabilizes MAT2A. Disruption of the cellular NADP+ remodels the protein level of MAT2A. The pentose phosphatase pathway regulates the level of MAT2A protein through the interaction of NADP+ with MAT2B. Additionally, MAT2B-MAT2A interaction regulates the mRNA m6A modification and stability. In liver tumors, the Mat2a mRNA level is elevated but the protein level is decreased by the restricted NADP+. Blocking the interaction between MAT2B and MAT2A by the keto diet can suppress liver tumor growth. These findings reveal that MAT2B is essential for regulating the protein levels of MAT2A and connecting SAM synthesis to mRNA m6A.

Molecular correlates of social adversity in breast cancer.

48 Background: Neighborhood disadvantage (ND) is associated with shorter breast cancer (BCa) survival independent of individual, tumor, and treatment factors, suggesting more aggressive tumor biology in women from disadvantaged neighborhoods. This study evaluates how differential DNA methylation (DNAm), gene expression, and biologic pathways are wired with each other to delineate the regulatory landscape of ND on BCa. Methods: We leveraged 55 geocoded ER+/HER2- BCa samples from Hispanic White women enrolled in a prospective genomic-epidemiologic cohort study. Objective ND was evaluated using the Area Deprivation Index (ADI) and subjective ND was evaluated using the patient-reported Neighborhood Social Environment Adversity Survey and the Everyday Expanded Discrimination Survey. We analyzed the association between ND and (1) DNAm, (2) RNA transcription and miRNAs, and (3) blood concentration levels of DNAm co-factors (B12 and folate) from the same patient. To evaluate how the DNAm/expressed pathways are wired with each other to delineate the regulatory landscape of ND in these tumors, we performed correlation analyses among CpGs, genes, and miRNAs in the same topologically associated domain (TAD). We used Spearman correlation as our metric and adjusted the resulting p-values to FDR values. Results: The cohort was divided into low ND (ADI&lt;5; 31%) and a high ND (ADI&gt;=5; 69%). There were no significant differences between groups by age, race/ethnicity, genetic ancestry, BMI, smoking/alcohol, subtype, stage, or OncotypeDX scores. We discovered that high ND exhibits epigenetic deregulation of immune pathways, consistent with our findings of lower circulating B12 in high ND, suggesting activation of immune pathways through reduced systemic availability of methyl donors. RNA-seq identified higher tumor infiltration of immune cells in patients from high ND (e.g., Interferon alpha and gamma responses and inflammatory responses) consistent with aggressive biology. Estrogen response genes were downregulated suggesting that patients from high ND may develop estrogen-resistant tumors. Higher subjective ND scores discovered epigenetic changes on cell adhesion and calcium signaling genes, along with gene expression of E2F targets and cell cycle’s G2M checkpoint, indicative of a more proliferative/stemness signature. TAD analysis discovered significant developmental, immune, and signaling pathways when correlating DNAm with gene expression (e.g., estrogen response and EMT). Conclusions: In this multi-omic matched cohort, we discovered differentially methylated/expressed pathways, consistent with aggressive biology, wired with each other to identify a novel regulatory landscape of ND on BCa. These findings can inform interventions such as improved access to healthy nutrition (B12) in ND/food deserts or targeted stress-reduction interventions based on our survey findings to ultimately reverse aggressive tumor biology and reduce BCa disparities by ND.

Closing in on human methylation-the versatile family of seven-β-strand (METTL) methyltransferases.

Methylation is a common biochemical reaction, and a number of methyltransferase (MTase) enzymes mediate the various methylation events occurring in living cells. Almost all MTases use the methyl donor S-adenosylmethionine (AdoMet), and, in humans, the largest group of AdoMet-dependent MTases are the so-called seven-β-strand (7BS) MTases. Collectively, the 7BS MTases target a wide range of biomolecules, i.e. nucleic acids and proteins, as well as severalsmall metabolites and signaling molecules. They play essential roles in key processes such as gene regulation, protein synthesis and metabolism, as well as neurotransmitter synthesis and clearance. A decade ago, roughlyhalf of the human 7BS MTases had been characterized experimentally, whereas the remaining ones merely represented hypothetical enzymes predicted from bioinformatics analysis, many of which were denoted METTLs (METhylTransferase-Like). Since then, considerable progress has been made, and the function of>80% of the human 7BS MTases has been uncovered. In this review, I provide an overview of the (estimated) 120 human 7BS MTases, grouping them according to substrate specificities and sequence similarity. I also elaborate on the challenges faced when studying these enzymes and describe recent major advances in the field.

Peripheral Blood ABCG1 Gene DNA Methylation: Mediating the Relationship between Dietary Intake of Methyl Donor Nutrients and Stroke Risk

Dysregulation of methyl donor nutrients interferes with DNA methylation and is associated with neurological diseases. ABCG1 gene regulates cholesterol to HDL-C, maintains lipid homeostasis, and has been linked to both methyl nutrition and neurological risks. The aim was to investigate whether there is an effect of ABCG1 DNA methylation on the relationship between intake of methyl donor nutrients and the risk of stroke occurrence. We hypothesize that the intake of methyl donor nutrients may influence stroke occurrence by modulating the methylation status of ABCG1. This study utilized a case-control design and selected 52 stroke patients along with 52 healthy controls from Northwest China. Dietary information was collected using a FFQ, and methylation levels were measured at 29 CpG sites of the ABCG1 gene. A significant linear trend was found between dietary intake of the methyl donor nutrient choline and CpG_19.20 methylation of the ABCG1 gene (β = -0.037, P = 0.033). Additionally, a significant association was observed between CpG_19.20 methylation and the risk of stroke (OR 2.325, 95% CI 1.210 - 4.466). Mediation analysis revealed that choline intake indirectly influenced stroke occurrence through its effect on CpG_19.20 methylation levels in the ABCG1 gene (β = -0.015, SE = 0.013, 95% CI = [-0.053, -0.001]). We found that DNA methylation at specific CpG sites of the peripheral blood ABCG1 gene mediates the association between dietary methyl donor nutrient intake and stroke risk in an adult population from Northwest China. New insights are provided on the prevention and treatment of stroke.

Modulation of m6A RNA modification by DAP3 in cancer cells

N6-methyladenosine (m6A) RNA methylation is a prevalent RNA modification that significantly impacts RNA metabolism and cancer development. Maintaining the global m6A levels in cancer cells relies on RNA accessibility to methyltransferases and the availability of the methyl donor S-adenosylmethionine (SAM). Here, we reveal that death associated protein 3 (DAP3) plays a crucial role in preserving m6A levels through two distinct mechanisms. First, although DAP3 is not a component of the m6A writer complex, it directly binds to m6A target regions, thereby facilitating METTL3 binding. Second, DAP3 promotes MAT2A's last intron splicing, increasing MAT2A protein, cellular SAM, and m6A levels. Silencing DAP3 hinders tumorigenesis, which can be rescued by MAT2A overexpression. This evidence suggests DAP3's role in tumorigenesis, partly through m6A regulation. Our findings unveil DAP3's complex role as an RNA-binding protein and tumor promoter, impacting RNA processing, splicing, and m6A modification in cancer transcriptomes.

Paternal undernutrition and overnutrition modify semen composition and preimplantation embryo developmental kinetics in mice

BackgroundThe importance of parental diet in relation to eventual offspring health is increasing in prominence due to the increased frequency of parents of reproductive age consuming poor diets. Whilst maternal health and offspring outcome have been studied in some detail, the paternal impacts are not as well understood. A father’s poor nutritional status has been shown to have negative consequences on foetal growth and development and ultimately impact the long-term adult health of the offspring. In this study, we examined sperm- and seminal vesicle fluid-mediated mechanisms of preimplantation embryo development alterations in response to sub-optimal paternal diets.ResultsMale mice were fed a diet to model either under (low-protein diet (LPD)) or over (high-fat/sugar ‘Western’ diet (WD)) nutrition, LPD or WD supplemented with methyl donors or a control diet (CD) before mating with age-matched females. Male metabolic health was influenced by WD and MD-WD, with significant changes in multiple serum lipid classes and hepatic 1-carbon metabolites. Sperm RNA sequencing revealed significant changes to mRNA profiles in all groups when compared to CD (LPD: 32, MD-LPD: 17, WD: 53, MD-WD: 35 transcripts). Separate analysis of the seminal vesicle fluid proteome revealed a significant number of differentially expressed proteins in all groups (LPD: 13, MD-LPD: 27, WD: 24, MD-WD: 19) when compared to control. Following mating, in vitro time-lapse imaging of preimplantation embryos revealed a significant increase in the timing of development in all experimental groups when compared to CD embryos. Finally, qPCR analysis of uterine tissue at the time of implantation identified perturbed expression of Cd14 and Ptgs1 following mating with WD-fed males.ConclusionsOur current study shows that paternal nutritional status has the potential to influence male metabolic and reproductive health, impacting on embryonic development and the maternal reproductive tract. This study highlights potential direct (sperm-mediated) and indirect (seminal vesicle fluid-mediated) pathways in which a father’s poor diet could shape the long-term health of his offspring.

PSV-13 Betaine supplementation and nutrient restriction during gestation: Influences on growth in fetal sheep offspring

Abstract Restricted nutrition during gestation can reduce offspring productivity. Supplementation of betaine, a methyl donor, promotes fetal growth and liver function. We hypothesized that nutrient restriction during gestation impairs growth, including organ weights and muscle mass, while betaine supplementation would alleviate these negative effects. To determine if maternal betaine supplementation during gestation would improve growth of offspring of restricted-fed dams, multiparous Dorset ewes (n = 47) were estrus synchronized and bred to one of three rams. Ewes confirmed pregnant with twins (n = 32) and singletons (n = 15) were randomly assigned to one of four diets [100% NRC requirements (CON), 60% NRC (RES), CON+betaine (CON+BET; 2 g/d), and RES+BET] from d 30 to 130 of gestation. Among the twins, there were 27 males (CON-8, RES-5, CON+BET-5, and RES+BET-9) and 37 females (CON-8, RES-9, CON+BET-11, and RES+BET-9). Within singletons, there were 7 males (CON-2, RES-2, CON+BET-2, and RES+BET-1) and 8 females (CON-3, RES-1, CON+BET-1, and RES+BET-3). Dam body weight (BW) was measured weekly from d 30 to d 130 of gestation, and diet adjusted accordingly. Dams were euthanized at d 130 of gestation and fetal BW and tissue weights were collected. Data were analyzed using RStudio with P ≤ 0.05 considered significant and 0.05 &amp;lt; P ≤ 0.10 as tendency. At d 130, dams carrying twin fetuses in the CON group were 11.3% and 19.1% heavier than RES and RES+BET dams, respectively (P ≤ 0.05), and CON+BET dams were 18.4% and 10.5% heavier than RES and BET+RES dams, respectively (P ≤ 0.07). However, dams with singletons showed no significant difference in BW (P ≥ 0.37). Regarding fetal BW for singletons, RES fetuses (3.80 ± 0.28kg) were lighter than CON (4.60 ± 0.21kg), CON+BET (4.72 ± 0.27kg), and RES+BET (4.59 ± 0.24kg; P ≤ 0.03). Whereas in twins, RES+BET fetuses (3.19 ± 0.11kg) were lighter than CON (3.68 ± 0.12kg), RES (3.73 ± 0.14kg), and CON+BET (3.60 ± 0.13kg; P ≤ 0.02). In all fetuses, the livers of male RES (28.0 ± 1.49g) and male RES+BET (27.7 ± 1.44g) tended to be heavier than those of female RES+BET (24.5 ± 1.19g; P ≤ 0.07). In terms of muscle mass, female CON+BET (9.33 ± 0.43g) fetuses tended to have greater longissimus muscle (LM) mass compared with female CON (8.30 ± 0.35; P = 0.06), whereas male RES (9.06 ± 0.44g) tended to have heavier LM than male CON+BET (8.02 ± 0.43g; P = 0.09). There were no observed effects of diet, fetal sex, or fetal number on the weights of heart, kidney, perirenal fat, pancreas, semitendinosus muscle, and triceps brachii muscle (P ≥ 0.11). These findings suggest that betaine supplementation may have a significant role in improving fetal BW in singleton pregnancies, with variable effects in twins. Additionally, betaine supplementation may have sex-specific effects on muscle development.

Methyl-Transferase-Like Protein 16 (METTL16): The Intriguing Journey of a Key Epitranscriptomic Player Becoming an Emerging Biological Target.

Key epitranscriptomic players have been increasingly characterized for their structural features and their involvement in several diseases. Accordingly, the design and synthesis of novel epitranscriptomic modulators have started opening a glimmer for drug discovery. m6A is a reversible modification occurring on a specific site and is catalyzed by three sets of proteins responsible for opposite functions. Writers (e.g., methyl-transferase-like protein (METTL) 3/METTL14 complex and METTL16) introduce the methyl group on adenosine N-6, by transferring the methyl group from the methyl donor S-adenosyl-methionine (SAM) to the substrate. Despite the rapidly advancing drug discovery progress on METTL3/METTL14, the METTL16 m6A writer has been marginally explored so far. We herein provide the first comprehensive overview of structural and biological features of METTL16, highlighting the state of the art in the field of its biological and structural characterization. We also showcase initial efforts in the identification of structural templates and preliminary structure-activity relationships for METTL16 modulators.