How the microbiome challenges “our concept of self”

This commentary [attached] is enjoyable to mull over. In just this past decade, the “gut-brain-microbiome” has been exploding in significance and challlenging many concepts that clinical medicine has held for decades. Previously, we humans have always considered ourselves to be individuals, based on our own DNA and chromosomes. And our genomes contained, or catalogued, the many different ways in which humans –– across time and space –– have learned to make sense of what it means to be “an individual self”. The “discrete self” was a philosophical certainty in both the natural and the human sciences.

Today, this philosophical certainty –– and therefore our sense of self –– faces major challenges, which would have seemed so improbable a decade ago. This change has to do with the enormous amounts of bacterial colonies that live in our intestines. It has been known, since invention of the microscope by van Leeuwenhoek in the 17th century, that animals, including humans, are hosts to many microorganisms. However, until recently these microorganisms were generally treated as either pathogens or as insignificant; in fact, the absence of microbes was equated with better health.

This classical understanding of microbes has been called into question, due to low-cost high-throughput gene-sequencing techniques, that has enabled us to study microbial communities without the need to grow them in a Petri dish. There is now overwhelming evidence that normal development, as well as the maintenance of any animal having a gut, depends upon our microbiome. Humans are not a unitary entity, but rather a dynamic and interactive community of human cells and microbial cells. By current estimates, approximately half of the cells in our body are microbial. Studies of the microbiome are leading to a major reassessment of biological processes –– as varied as the physiological function of specific organs, composition of metabolites in body fluids, and management of transmissible diseases.

Evidence now shows that our resident microbes orchestrate the adaptive immune system, influence the brain, and contribute more gene functions than our own genome. Realization that humans are not individual, discrete entities –– but rather the outcome of ever-changing interactions with microorganisms –– has consequences beyond the biological disciplines. In particular, these conseuences call into question the assumption that distinctive human traits set us apart from all other animals. And therefore also “the traditional disciplinary divisions between the arts and the sciences.”

PLoS Biol Feb 2o18; 16: e2005358

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Perfluoroalkyl substances and changes in body weight and resting metabolic rate (RMR) in response to weight-loss diets: A prospective study

This is a “correlation ––> inferred-causation” epidemiological study, and I would appreciate any comments/criticisms about these findings. Perfluoroalkyl substances (PFASs) –– especially perfluorooctanoic acid (PFOA) and perfluorooctanesulfonic acid (PFOS) –– have been in the news a lot lately, and identified as plausible endocrine disruptors with the potential to perturb weight regulation. Evidence from animal studies has suggested that PFASs may be involved in altering energy metabolism and thyroid hormone homeostasis, likely through activation of various transcriptional factors such as the peroxisome proliferator-activated receptors. However, given species-specific toxicokinetics and tissue distribution of PFASs, extrapolation from animals to humans is always difficult.

Authors [see attached] examined associations of PFAS exposure with changes in body weight and resting metabolic rate (RMR) in a diet-induced weight-loss setting. In the 2-year “POUNDS Lost” randomized clinical trial –– based in Boston, Massachusetts, and Baton Rouge, Louisiana –– which examined effects of energy-restricted diets on weight changes, baseline plasma concentrations of major PFASs were measured among 621 overweight and obese participants (aged 30-70 years). Body weight was measured at baseline and 6, 12, 18, and 24 months. Participants lost an average of 6.4 kg of body weight during the first 6 months (weight-loss period) and subsequently regained an average of 2.7 kg of body weight during the period of 6 to 24 months (weight-regain period). [Note that the annoying television ads for “weight-loss programs” describe enthusastically the ‘weight-loss period’, but fail to mention the ‘weight-regain period’.] 🙁

After multivariate adjustment, baseline PFAS concentrations were not significantly associated with concurrent body weight or weight loss during the first 6 months. In contrast, higher baseline levels of PFASs were significantly associated with a greater weight-regain, primarily in women. In women, comparing the highest to the lowest tertiles (statistical divisions of any population into three equal parts, the two extreme groups being divided by the middle group) of PFAS concentrations, the multivariate-adjusted mean weight-regain was: 4.3 vs 2.2 kg for PFOA; 4.0 vs 2.1 kg for PFOS; 4.7 vs 2.5 kg for perfluorononanoic acid (PFNA); 4.9 vs 2.7 kg for perfluorohexanesulfonic acid; and 4.2 vs 2.5 kg for perfluorodecanoic acid.

Higher baseline plasma PFAS concentrations, especially for PFOS and PFNA, were significantly associated with greater decline in RMR during the weight-loss period and less increase in RMR during the weight-regain period in both men and women. Caveats (limitations of the study) include the possibility of unmeasured or residual confounding by socioeconomic and psychosocial factors (i.e. greater obesity among the poor), as well as possible relapse to the usual diet prior to randomization, which could have been rich in foods contaminated by PFASs through food-packaging and also dense in energy.

Authors conclude that, in this diet-induced weight-loss study, higher baseline plasma PFAS concentrations were associated with a greater weight-regain, especially in women, possibly explained by a slower return to their normal RMR levels. These data convincingly illustrate a potential novel pathway through which PFASs interfere with clinical body weight regulation and metabolism. Possible impact of environmental chemicals on the obesity epidemic thus deserves more attention.

PLoS Med Feb 2o18; 15: e1002502

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A STUDY OF EVOLUTION: Different mutational rates in single human cells at pregastrulation, neurogenesis, and during aging and neurodegeneration

Any replacement of a DNA base –– in the single strand of viruses and prokaryotes (bacteria & archaebacteria), or in one of the paired strands of eukaryotes (plants, fungi & animals) –– results in a MUTATION. DNA mutations accumulate at a steady pace across the genome, passing from one generation to the next; this is one of the principal mechanisms by which species evolve and improve their chances of survival (via finding food, avoiding predators & reproduction). Enhanced virulence of a virus, development of bacterial resistance to an antibiotic, adaptation of a tumor to resist ongoing exposure to a chemotherapeutic drug, and evolution of a species –– are ALL examples of this evolutionary process. Because most DNA is not involved in encoding proteins or participating in critical-life regulatory DNA modules, the vast majority of mutations are “silent.” Nevertheless, on the basis of the degree of shared mutations, a genealogical relationship can be reconstructed from ancient and modern individuals –– allowing one to go back hundreds of thousands of years in human evolutionary history.

Instead of comparing individuals, in the two [attached] full articles, authors assessed the rate of DNA mutation in single cells from developing [1st article] and aging [2nd article] human brains –– revealing mutational histories in neurodevelopment, aging, and neurodegeneration. These approaches also have implications for understanding complex diseases that could result from somatic mutations that arise later in life (e.g. cancer). De novo mutations in DNA of the egg or sperm can be associated with devastating disorders affecting young individuals, because all cells in the body inherit these germline mutations.

By contrast, somatic DNA mutations sporadically occur throughout the life of an organism as a result of (environmentally-caused) DNA damage –– as well as errors in DNA replication or repair. [By “environment”, this means ionizing radiation, in utero signals, dietary and lifestyle effects, and exposure to mutagenic chemicals or metals.] When somatic mutations occur early in the life in dividing cells, they are obviously found in a large number of cellular descendants. If mutations occur in dividing cells as humans age, they are found in only a limited number of cells, resulting in tissue mutational mosaicism (see figure in attached editorial). The inheritance pattern of mutations in cells within a tissue can be used to establish a temporal, or genealogical, relationship of mutations to understand better the role of mutational mosaicism in human diseases.

Somatic mosaicism in human brain may alter function of individual neurons. Authors [attached full article 550] analyzed genomes of single cells from three human forebrains (15 to 21 weeks postconception). They detected 200-400 single-nucleotide variations (SNVs) per cell. SNVs with a frequency of more than 2% in brain were also present in spleen, confirming these mutations were pregastrulation in origin. Authors reconstructed cell lineages for the first five postzygotic cleavages and calculated a mutation rate of ~1.3 mutations per division per cell (this is a pretty small number, considering the cell’s haploid genome is more than 1 billion nucleotides). Later in development –– during neurogenesis (formation of neurons and the nervous sytem), the mutation spectrum intriguingly shifted toward oxidative damage, and the mutation rate increased. Both neurogenesis and early embryogenesis exhibited substantially more mutagenesis than adulthood. [Could this finding reflect insufficient defenses against oxidative stress during early embryogenesis?]

Based on a comparison of counts of germline SNVs –– 3,746,847 for subject 275; 3,809,591 for subject 316; 4,316,547 for subject 320; to those derived by The 1000 Genomes Project across different human subpopulations –– authors concluded that subjects 275 and 316 were of non-African origin, whereas subject 320 was of African descent. To explain this quite simply (and in papers of the Great Human Diaspora frequently described in these GEITP pages), humans of African descent have existed on the planet longer than humans of non-African origin, and therefore African DNA has been “exposed to” environmental insults longer than DNA of non-African origin.

Authors [attached full article 555] used single-cell whole-genome sequencing to perform genome-wide somatic SNV identification on DNA from 161 single neurons from prefrontal cortex and hippocampus of 15 normal individuals (aged 4 months to 82 years), as well as nine individuals affected by early-onset neurodegeneration due to genetic disorders of DNA repair (Cockayne syndrome and xeroderma pigmentosum). Somatic SNVs increased approximately linearly with age in both areas of the brain and were greater in number in patients with neurodegenerative disease. Accumulation of somatic mutations with age showed age-related, region-related, and disease-related molecular signatures –– which authors suggest might be important in other human age-associated conditions.

Science 2 Feb 2o18; 359: 550–555 & 555–559 & editorial pp 521–522

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Evolutionary Rewiring of Human Regulatory Networks by Waves of Genome Expansion

Why does a human “look different” than an elephant, which “appears different” than a frog? As these GEITP pages have often described, whereas the genotype is the genetic makeup of an organism, the phenotype (trait) is how genetic and environmental influences come together to create an organism’s physical appearance and behavior. A trait includes such things as: height, body mass index, blood pressure, serum cholesterol level, shape of ears, or manifestation of any Mendelian or complex disease (Marfan syndrome, type-2 diabetes, cancer). Evolution of regulatory networks is believed to be the cause of a substantial fraction of phenotypic divergence among vertebrates (animals having a spinal column).

Genetic events affecting gene regulation can be classified into two classes: [a] exaptation (i.e. a trait that has been co-opted for a use other than the one for which natural selection has built it) of existing DNA sequence (and epigenetic effects) through accumulation of small-scale mutations; and [b] de novo appearance of regulatory DNA through genome expansion driven (for example) by transposable elements (TEs; i.e. “jumping genes” or DNA segments that move around and re-insert elsewhere in chromosomes). Both mechanisms have been shown to be relevant in the evolution of human regulatory DNA.

In particular, information-rich binding sites (BSs) –– such as the one recognized by CTCF (a single protein that is a TF; CCCTC-binding factor) –– are much less likely to arise through accumulation of random point mutations than simpler binding motifs. In fact, it has been shown that the expansion of lineage-specific TEs efficiently remodeled the CTCF regulome. In a study of the activity of TEs in generating transcription-factor-binding sites (TFBSs), it was observed that ~20% of BSs are embedded within TEs, thus revealing the latent regulatory potential of these elements. It was shown that recent enhancer evolution in mammals can be largely explained by exaptation of existing ancestral sequences rather than by the expansion of lineage-specific repeated elements. A systematic investigation of the role of genomic sequence expansion in rewiring regulatory networks is, however, still missing; authors [see attached] have therefore tried to fill this gap –– by attempting to reconstruct a much longer evolutionary history, focusing on regulatory evolution through genome expansion since the evolutionary time of the common ancestor of all vertebrates.

To investigate the role of newly arising sequences in rewiring regulatory networks, authors [see attached] estimated the age of each region of the human genome by applying maximum parsimony (in phylogenetic biology, maximum parsimony is an optimality criterion –– under which the phylogenetic tree that minimizes the total number of character-state changes is the preferred condition). Authors carried out genome-wide alignments with 100 vertebrate species. They then studied the age distribution of several types of functional regions –– with a focus on regulatory elements. The age distribution of regulatory elements reveals the extensive use of newly formed genomic sequence in the evolution of regulatory interactions. Intriguingly, many TFs have expanded their repertoire of targets through waves of genomic expansions that can be traced to specific evolutionary times.

Repeated elements contributed a major part of such expansion: many classes of such elements are enriched in BSs of one or a few specific TFs, whose binding sites are localized in specific portions of DNA modules and characterized by distinctive motif signatures; these features suggest that the BSs were available as soon as the new sequence entered the genome –– rather than being created later by accumulation of point mutations. By comparing the age of regulatory regions to the evolutionary shift in expression of nearby genes, authors show that rewiring through genome expansion played an important role in shaping human regulatory networks. This is a really cool, but very complicated, study to appreciate. 🙂

Am J Hum Genet 1 Feb 218; 102: 207–218

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The GREAT HUMAN DIASPORA: Evidence of an early migration of modern humans leaving Africa

The editorial [attached] is titled “When did modern humans leave Africa?” Which is a bit misleading (maybe sensationalistic?) because it has been known for decades that modern human (Homo sapiens) evolved originally (from other hominid species) in southeastern Africa –– at least 300,000 years ago –– and then migrated in a series of waves “Out of Africa”, first to the Levant, as well as eastward toward present-day China, and more recently through the Caucasus Mountains to Europe and northeastward across Asia toward present-day Mongolia. Many of the early Homo sapiens “waves” died out, and it wasn’t until 70,000-90,000 years that a wave became “successfully fixed” in the eastern Mediterrean, outside Africa.

This information is not to be confused with the Homo neanderthalensis and Homo denisove sublines. Neanderthals and Denisovans emerged as distinct hominids as old as 600,000 years ago and appear to have been successfully entrenched in parts of Europe and Southeast Asia, respectively –– long before Homo sapiens became irreversibly inserted.

The skeletal features of Homo sapiens include a globular brain-case, brow ridges that are divided into central and side portions, a flat and retracted midface, chin on the lower jaw, and narrow pelvis. Fossils showing many of these characteristics have been excavated from Ethiopian sites between 160,000-195,000 years ago. Possibly more primitive members of Homo sapiens have been found in present-day Morocco and South Africa –– dated at ~315,000 and ~259,000 years ago, respectively. Yet, the oldest known H. sapiens fossils outside of Africa were found in Israel and have been dated at 90,000-120,000 years old.

The attached full article describes fossils from a cave in Israel, providing evidence that one of the early Homo sapiens waves outside had occurred ~180,000 years ago. A maxilla and associated teeth –– recently discovered at Misliya Cave, Israel –– was dated to 177,000-194,000 years ago. These anthropological studies are consistent with recent DNA sequencing studies, which have speculated the possibility of at least one of the earliest dispersals of Homo sapiens ~220,000 years ago. The Misliya maxilla fossil is associated with full-fledged Levallois technology (i.e. creating a knife or scraper from a stone or bone) in the Levant, suggesting that emergence of this technology is linked to the appearance of Homo sapiens in the region, as had been previously documented in Africa.

Science 26 Jan 2o18; 359: 456–459 & editorial, pp 389–390

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Chronic Inflammation Suppresses Immune Cells that Fight Liver Cancer

This interesting study was funded, in part, by the NIEHS-funded Superfund Research Program (SRP),

Chronic Inflammation Suppresses Immune Cells that Fight Liver Cancer

Researchers at the University of California, San Diego (UCSD) showed that chronic liver inflammation can promote cancer by suppressing one of the body’s natural mechanisms to fight cancer development. The study helps to explain the success of some types of cancer immunotherapy and suggests novel targets for new therapies.

Inflammation is one way the liver responds to substances people are exposed to through pollution, diet, and other means. Chronic inflammation can drive many cancers, especially liver cancer, and researchers have long thought that this was because the effects of inflammation both stimulate cancer cells to divide and protect them from cell death.

In the new study, scientists found that in the liver, chronic inflammation also suppresses a natural defense mechanism known as immuno-surveillance. This mechanism involves cells in the immune system, known as cytotoxic T cells, that guard against disease agents such as viruses, bacteria, and cancerous and precancerous cells.

According to the authors, the results provide one of the strongest and most direct demonstrations that immunosurveillance actively prevents cancer emergence, while specific immune cells associated with chronic inflammation hamper this process. These findings could help scientists develop new ways to interfere with immunosuppressive lymphocytes to prevent or treat early liver cancer.

Guarding Against Cancer

Cytotoxic T cells surround a cancer cell.The researchers studied a new mouse model of liver cancer that more closely mimics the way human liver cancer develops. In these mice, tumors develop as a natural consequence of nonalcoholic steatohepatitis (NASH), a chronic metabolic disorder that causes liver damage, fibrosis, and cell mutations. Michael Karin, Ph.D., led the study with first author Shabnam Shalapour, Ph.D., an assistant professor in his group.

The researchers found that mutations associated with NASH-induced liver cancer provoke cytotoxic T cells to recognize and attack newly emerging cancer cells. However, chronic liver inflammation also led to the accumulation of another type of immune cell known as immunosuppressive lymphocytes, or IgA+ plasmocytes.

Experiments in the new study showed that immunosuppressive lymphocytes use a molecule known as PD-L1 (the correct nomenclature is CD274) to interfere with cytotoxic T cells, leading to liver tumors in the chronically inflamed mice. By stopping the T cells, liver tumors formed and grew.

In mice lacking tumor-fighting cytotoxic T cells, 27 percent (i.e. four mice) of 15 mice had large liver tumors at six months. Mice of the same age that had retained their cytotoxic T cells had no tumors. Similarly, mice without immunosuppressive lymphocytes had almost no tumors even at 11 months, most likely because cytotoxic T cells could fight the emerging cancer.

When the researchers inhibited CD274 with a drug or by genetic engineering, cytotoxic T cells were re-invigorated and cleared the tumors.

New Targets for Treatment

According to the authors, these findings provide an explanation for the ability of anti-CD274 drugs, which block the receptor for CD274, to induce liver cancer regression in a recent clinical trial.

Moving forward, the research team is investigating how immunosuppressive lymphocytes are recruited to the liver. Finding a way to interfere with the recruitment or generation of these cells may provide a new approach to prevent or treat liver cancer.

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Large genome-wide association study (GWAS) identifies gene variants associated with neuroticism

Neuroticism is a relatively stable personality trait –– characterized by negative emotionality (for example, worry and guilt). Clearly the trait of neuroticism would be regarded as multifactorial, i.e. contributions of hundreds if not thousands of genes, plus epigenetic factors, plus environmental effects. Heritability estimated from twin studies ranges from 30% to 50% “heritable”, and DNA-variant-based heritability ranges from 6% to 15%. Increased neuroticism is also associated with poor mental and physical health, translating to a high economic burden.

The strong genetic correlation between neuroticism and mental health, especially anxiety and major depressive disorder (MDD), means that exploring genetic contribution to differences in neuroticism is one way to understand more about these common and burdensome illnesses. In the largest genome-wide association study (GWAS) of MDD –– 130,664 cases versus 330,470 controls –– 44 independent associated genetic loci were identified, and as many as 11 genetic loci have been associated with neuroticism.

Authors [see attached] report 116 significant independent loci from a GWAS of neuroticism in 329,821 UK Biobank participants; 15 of these loci were replicated at P <0.00045 in an unrelated cohort (N = 122,867). Genetic signals were enriched in neuronal genesis and brain-differentiation pathways, and substantial genetic correlations were found between neuroticism and depressive symptoms (rg = 0.82), MDD (rg = 0.69), and subjective well-being (rg = –0.68) together with other mental health traits. These data should help advance our understanding of neuroticism and its association with MDD. Nat Genet Jan 2o18; 50: 6–10

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Large genome-wide association study (GWAS) identifies protein-altering variants associated with BMI and energy intake, in obesity

Obesity is an inherited multifactorial trait –– which reflects the contribution of genetics (DNA sequence variants), epigenetic factors (DNA-methylation, RNA-interference, histone modifications, and chromatin remodeling), plus environmental effects (diet, lifestyle, in utero environment during the pregnancy, drug-drug interactions, chronic diseases, etc.). Genome-wide association studies (GWAS) for body-mass index (BMI) and obesity risk, over the past decade, have identified more than 250 common variants. Tissue expression and gene set enrichment analyses for genes implicated in BMI-associated loci have shown that the central nervous system (CNS) plays a critical role in body weight regulation. Whereas numerous GWAS-loci have provided insight into broad biological mechanisms underlying body weight regulation, pinpointing the causal gene(s) and variant(s) remains a major challenge, because GWAS-identified variants are typically noncoding and may affect genes at long distances from the identified DNA variant nucleotide.

The association of intronic FTO variants with BMI –– is a great example of the challenges of identifying causal regulatory effects. The proposed causal variant in this locus was found to regulate expression of nearby RPGRIP1L in some studies, whereas others found that it regulated the distant IRX3 and IRX5 genes in specific cell types. Authors [see attached] performed whole-exome sequencing (WES) to search for low-frequency (minor allele frequency; MAF = 1–5%) and rare (MAF <1%) single-nucleotide variants (SNVs) associated with BMI. A meta-analysis of 125 studies (N = 718,734) included single-variant associations between 246,000+ SNVs and BMI. In addition, authors performed gene-based meta-analyses to aggregate rare and low-frequency coding SNVs across 14,541 genes. Using genetic, functional and computational follow-up analyses, authors gained insights into the function of BMI-implicated genes and the biological pathways through which they might influence body weight. Authors identified 14 coding variants in 13 genes, of which eight variants were in genes newly implicated in obesity (ZBTB7B, ACHE, RAPGEF3, RAB21, ZFHX3, ENTPD6, ZFR2 and ZNF169), two variants were in genes (MC4R and KSR2) previously observed to be mutated in extreme obesity, and two variants were in GIPR. (gastric inhibitory polypeptide receptor). The effect-sizes of rare variants were ~10 times larger than those of common variants, with the largest effect observed in carriers (who weighed ~7 kg more than non-carriers) of an MC4R mutation (melanocortin-4 receptor). Pathway analyses –– based on the variants associated with BMI –– confirm enrichment of neuronal genes and provide new evidence for adipocyte (fat cell) and energy expenditure biology, widening the potential of genetically supported therapeutic targets in obesity. Nat Genet Jan 2o18; 50: 26–41

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Clostridium difficile (C diff) prevalence is enhanced by dietary trhalose-containing foods !!

Between 2001 and 2006, epidemic strains of the bacterium Clostridium difficile (C.diff) –– which can persist in some patient’s bowel and cause dangerous diarrhea, unexpectedly emerged in the United States, Canada, and several European countries. Most of these strains originated from a single lineage of C. diff known as ribotype 027 (RT027), which has now spread around the world. Of par­ticular concern has been the correlation between RT027 and dramatic increases in deaths related to C. difficile. The mystery of why this ribotype, and a second one, RT078, became so prevalent –– apparently out of thin air –– has remained largely unsolved. Authors [see attached] raise the possibility that the seemingly harmless addition of a sugar called trehalose to the food supply might have contributed to this disease epidemic.

Whole-genome sequencing (WGS) analysis of C. diff RT027 strains demonstrated that two independent lineages had emerged in North America between 2000 and 2003. Comparison with historic pre-epidemic RT027 strains showed that both epidemic lineages had acquired a mutation in the gyrA gene, leading to increased resistance to fluoroquinolone antibiotics. While the development of fluoroquinolone resistance has almost certainly played a role in the spread of RT027 strains, fluoroquinolone resistance has also been observed in non-epidemic C. diff ribotypes and has been identified in strains dating back to the mid-1980s. Thus, other factors must have contributed to the emergence of epidemic RT027 strains.

The prevalence of a second C. diff ribotype, RT078, increased 10-fold in hospitals and clinics from 1995 to 2007 and was associated with enhanced disease severity. It is noteworthy that the RT027 and RT078 lineages are phylogenetically distant from one another, indicating that the evolutionary changes leading to concurrent increases in epidemics and disease severity might have emerged by independent mechanisms. Authors show that the two epidemic ribotypes (RT027 and RT078) have acquired unique mechanisms to metabolize low concentrations of the disaccharide trehalose –– which reflects a single point-mutation in the trehalose repressor gene (gyrA), which increases sensitivity of this ribotype to trehalose by more than 500-fold. Dietary trehalose was also shown to increase virulence of a RT027 strain in a mouse model of infection.

It so happens that RT078 strains acquired a cluster of four genes involved in trehalose metabolism, including a “PTS permease” (phosphotransferase system, or PTS, is a distinct method used by bacteria for sugar uptake in which the source of energy is from phosphoenolpyruvate (PEP); this multicomponent system always involves enzymes of the plasma membrane and cytoplasm) –– that is both necessary and sufficient for growth on low concentrations of trehalose. Authors propose that implementation of trehalose as a food additive into the human diet, which occurred shortly before the emergence of these two epidemic lineages, might have helped these bacteria to select for their emergence (i.e. “evolutionary survival”) and contributed to the observed hypervirulence.

Nature 18 Jan 2o18; 553: 291–294 & News-N-Views ed pp 285–286

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Characterizing the single-cell DNA methylome of mouse and human preimplantation embryos

As these GEITP pages have often stated, a trait (phenotype) reflects the contribution of genetics (DNA sequence differences) plus epigenetics (chromosomal changes other than DNA sequence) plus adverse environmental insults that constantly affect DNA sequence and epigenetic variations. Epigenetics includes DNA-methylation, RNA-interference, histone modifications, and chromatin remodeling. These two papers [attached] describe DNA-methylation events that happen in pluripotent mouse, and human, pre-implantation embryos. In mammals, early lineage specification in pre-implantation and post-implantation embryonic development generates founder tissues for all subsequent somatic development.

The first lineage specification starts at the morula stage, when the inner cell mass (ICM) and the trophectoderm (TE) begin to segregate. The ICM contains both cells of the epiblast lineage, which give rise to the entire fetus, and cells of the primitive endoderm lineage, which form visceral endoderm and parietal endoderm. Visceral endoderm becomes the chief metabolic component of the visceral yolk sac, and parietal endoderm contributes to the transient parietal yolk sac. The TE contains progenitor cells for trophoblasts, which form the majority of the fetal-origin part of the placenta. In mice, by embryonic day 6.5 (E6.5), the anterior epiblast gives rise to ectoderm, and the posterior proximal epiblast develops into the primitive streak, which then forms mesoderm and endoderm. The resulting three germ layers contain virtually all progenitors for the future body plan (epiderm ––> skin; mesoderm ––> muscle and bone; and endoderm ––> internal organs such as liver).

However, the dynamics of transcriptomes (arising from DNA sequence ––> messenger RNA), and epigenomes (DNA-methylation, RNA-interference, histone modifications, and chromatin remodeling), acting in concert with initial cell fate commitment, remains poorly understood. Authors [first attached paper] report investigation of transcriptomes, and base-resolution methylomes, for early lineages in peri- and post-implantation mouse embryos. They found allele-specific and lineage-specific de novo DNA-methylation that leads to differential methylation between embryonic and extra-embryonic lineages at promoters of lineage regulators, gene bodies, and

DNA-methylation valleys. By defining chromatin architecture across the same developmental period –– authors demonstrated that both global demethylation and remethylation in early development correlate with chromatin compartments. Dynamic local methylation was evident during gastrulation, which enabled identification of putative regulatory elements. Lastly, authors found that de novo methylation patterning does not strictly require implantation. These findings thus reveal dynamic transcriptomes, DNA methylomes, and 3-dimensional chromatin landscapes during the earliest stages of mouse lineage specification.

Authors [second attached paper] performed single-cell DNA-methylome sequencing for human preimplantation embryos and found that tens of thousands of genomic loci exhibit de novo DNA-methylation. This observation indicates that genome-wide DNA-methylation reprogramming during pre-implantation development is in a dynamic balance between strong global demethylation and extremely-focused remethylation. Moreover, demethylation of the paternal genome is much faster and thorough than that of the maternal genome. From the two-cell zygote to the post-implantation stage, methylation of the paternal genome is consistently lower than that of the maternal genome. Authors also found that the genetic lineage of early blastomeres can be traced by DNA-methylation analysis. This breakthrough work paves the way for deciphering the secrets of DNA-methylation reprogramming in early human embryos.

Nat Genet Jan 2o18; 50: 12–19 & pp 96–105

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