Mutational spectra of aflatoxin B

Aflatoxin B₁ (AFB₁) and/or hepatitis B and C viruses are risk factors for human hepatocellular carcinoma (HCC). Available evidence supports the interpretation that formation of AFB₁-DNA adducts in hepatocytes seeds a population of mutations, mainly G:C→T:A, and viral processes synergize to accelerate tumorigenesis, perhaps via inflammation. Responding to a need for early-onset evidence predicting disease development, highly accurate duplex sequencing was used to monitor acquisition of high-resolution mutational spectra (HRMS) during the process of hepatocarcinogenesis. Four-day-old male mice were treated with AFB₁ using a regimen that induced HCC within 72 wk. For analysis, livers were separated into tumor and adjacent cellular fractions. HRMS of cells surrounding the tumors revealed predominantly G:C→T:A mutations characteristic of AFB₁ exposure. Importantly, 25% of all mutations were G→T in one trinucleotide context (CGC; the underlined G is the position of the mutation), which is also a hotspot mutation in human liver tumors whose incidence correlates with AFB₁ exposure. The technology proved sufficiently sensitive that the same distinctive spectrum was detected as early as 10 wk after dosing, well before evidence of neoplasia. Additionally, analysis of tumor tissue revealed a more complex pattern than observed in surrounding hepatocytes; tumor HRMS were a composite of the 10-wk spectrum and a more heterogeneous set of mutations that emerged during tumor outgrowth. We propose that the 10-wk HRMS reflects a short-term mutational response to AFB₁, and, as such, is an early detection metric for AFB₁-induced liver cancer in this mouse model that will be a useful tool to reconstruct the molecular etiology of human hepatocarcinogenesis.National Institutes of Health (U.S.) (Grant R01-ES016313)National Institutes of Health (U.S.) (Grant P30-ES002109)National Institutes of Health (U.S.) (Grant T32-ES007020)National Institutes of Health (U.S.) (Grant R01-CA080024

Ancestral Components of Admixed Genomes in a Mexican Cohort

For most of the world, human genome structure at a population level is shaped by interplay between ancient geographic isolation and more recent demographic shifts, factors that are captured by the concepts of biogeographic ancestry and admixture, respectively. The ancestry of non-admixed individuals can often be traced to a specific population in a precise region, but current approaches for studying admixed individuals generally yield coarse information in which genome ancestry proportions are identified according to continent of origin. Here we introduce a new analytic strategy for this problem that allows fine-grained characterization of admixed individuals with respect to both geographic and genomic coordinates. Ancestry segments from different continents, identified with a probabilistic model, are used to construct and study “virtual genomes” of admixed individuals. We apply this approach to a cohort of 492 parent–offspring trios from Mexico City. The relative contributions from the three continental-level ancestral populations—Africa, Europe, and America—vary substantially between individuals, and the distribution of haplotype block length suggests an admixing time of 10–15 generations. The European and Indigenous American virtual genomes of each Mexican individual can be traced to precise regions within each continent, and they reveal a gradient of Amerindian ancestry between indigenous people of southwestern Mexico and Mayans of the Yucatan Peninsula. This contrasts sharply with the African roots of African Americans, which have been characterized by a uniform mixing of multiple West African populations. We also use the virtual European and Indigenous American genomes to search for the signatures of selection in the ancestral populations, and we identify previously known targets of selection in other populations, as well as new candidate loci. The ability to infer precise ancestral components of admixed genomes will facilitate studies of disease-related phenotypes and will allow new insight into the adaptive and demographic history of indigenous people

7,8-Dihydro-8-oxo-1,N6-ethenoadenine: an exclusively Hoogsteen-paired thymine mimic in DNA that induces A→T transversions in Escherichia coli

14 pags., 9 figs.This work investigated the structural and biological properties of DNA containing 7,8-dihydro-8-oxo-1,N6-ethenoadenine (oxo-ϵA), a non-natural synthetic base that combines structural features of two naturally occurring DNA lesions (7,8-dihydro-8-oxoadenine and 1,N6-ethenoadenine). UV-, CD-, NMR spectroscopies and molecular modeling of DNA duplexes revealed that oxo-ϵA adopts the non-canonical syn conformation (χ = 65º) and fits very well among surrounding residues without inducing major distortions in local helical architecture. The adduct remarkably mimics the natural base thymine. When considered as an adenine-derived DNA lesion, oxo-ϵA was >99% mutagenic in living cells, causing predominantly A→T transversion mutations in Escherichia coli. The adduct in a single-stranded vector was not repaired by base excision repair enzymes (MutM and MutY glycosylases) or the AlkB dioxygenase and did not detectably affect the efficacy of DNA replication in vivo. When the biological and structural data are viewed together, it is likely that the nearly exclusive syn conformation and thymine mimicry of oxo-ϵA defines the selectivity of base pairing in vitro and in vivo, resulting in lesion pairing with A during replication. The base pairing properties of oxo-ϵA, its strong fluorescence and its invisibility to enzymatic repair systems in vivo are features that are sought in novel DNA-based probes and modulators of gene expression.MIT Skoltech Next Generation Program Pilot Grant (to J.M.E.); National Institutes of Health (NIH) [R01-CA080024 to J.M.E.]; NIEHS Center Grant [P30-ES002109 (to Center for Environmental Health Sciences, which provided access to NGS facilities)]; Skoltech (to T.S.Z.); MICINN [PID2020-116620GB-I00 to C.G.]; Ministry of Science and Higher Education Russian Federation [07515-2021-1049 to A.V.A. – synthesis and UV/CD studies]. Funding for open access charge: Skoltech.Peer reviewe

Association between cholinesterase activity and critical illness brain dysfunction

Abstract Background Delirium is a frequent manifestation of acute brain dysfunction and is associated with cognitive impairment. The hypothesized mechanism of brain dysfunction during critical illness is centered on neuroinflammation, regulated in part by the cholinergic system. Point-of-care serum cholinesterase enzyme activity measurements serve as a real-time index of cholinergic activity. We hypothesized that cholinesterase activity during critical illness would be associated with delirium in the intensive care unit (ICU) and cognitive impairment after discharge. Methods We enrolled adults with respiratory failure and/or shock and measured plasma acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) activity on days 1, 3, 5, and 7 after enrollment. AChE values were also normalized per gram of hemoglobin (AChE/Hgb). We assessed for coma and delirium twice daily using the Richmond Agitation Sedation Scale and the Confusion Assessment Method for the ICU to evaluate daily mental status (delirium, coma, normal) and days alive without delirium or coma. Cognitive impairment, disability, and health-related quality of life were assessed at up to 6 months post-discharge. We used multivariable regression to determine whether AChE, AChE/Hgb, and BChE activity were associated with outcomes after adjusting for relevant covariates. Results We included 272 critically ill patients who were a median (IQR) age 56 (39–67) years and had a median Sequential Organ Failure Assessment score at enrollment of 8 (5–11). Higher daily AChE levels were associated with increased odds of being delirious versus normal mental status on the same day (Odds Ratio [95% Confidence Interval] 1.64 [1.11, 2.43]; P = 0.045). AChE/Hgb and BChE activity levels were not associated with delirious mental status. Lower enrollment BChE was associated with fewer days alive without delirium or coma (P = 0.048). AChE, AChE/Hgb, and BChE levels were not significantly associated with cognitive impairment, disability, or quality of life after discharge. Conclusion Cholinesterase activity during critical illness is associated with delirium but not with outcomes after discharge, findings that may reflect mechanisms of acute brain organ dysfunction. Trial Registration: NCT03098472. Registered 31 March 2017

Loss of Heterozygosity Is Present in SEC63 Germline Carriers with Polycystic Liver Disease

Contains fulltext : 108556.pdf (publisher's version ) (Open Access)Polycystic liver disease (PCLD) is an autosomal dominant disorder characterised by multiple fluid filled cysts in the liver. This rare disease is caused by heterozygous germline mutations in PRKCSH and SEC63. We previously found that, in patients with a PRKCSH mutation, over 76% of the cysts acquired a somatic 'second-hit' mutation in the wild type PRKCSH allele. We hypothesise that somatic second-hit mutations are a general mechanism of cyst formation in PCLD which also plays a role in PCLD patients carrying a SEC63 germline mutation. We collected cyst epithelial cells from 52 liver cysts from three different SEC63 patients using laser microdissection. DNA samples were sequenced to identify loss of heterozygosity (LOH) mutations and other somatic mutations in cyst epithelial DNA. We discovered somatic SEC63 mutations in patient 3 (1/14 cysts), but not in patient 1 and 2 (38 cysts). Upon review we found that the germline mutation of patient 1 and 2 (SEC63 c.1703_1705delAAG) was present in the same frequency in DNA samples from healthy controls, suggesting that this variant is not causative of PCLD. In conclusion, as somatic second-hit mutations also play a role in cyst formation in patients with a SEC63 germline mutation, this appears to be a general mechanism of cyst formation in PCLD