DNA replication-associated mutations are repaired by two components: polymerase proofreading and mismatch repair. The mutation consequences of disruption to both repair components in humans are not ...well studied. We sequenced cancer genomes from children with inherited biallelic mismatch repair deficiency (bMMRD). High-grade bMMRD brain tumors exhibited massive numbers of substitution mutations (>250/Mb), which was greater than all childhood and most cancers (>7,000 analyzed). All ultra-hypermutated bMMRD cancers acquired early somatic driver mutations in DNA polymerase ɛ or δ. The ensuing mutation signatures and numbers are unique and diagnostic of childhood germ-line bMMRD (P < 10(-13)). Sequential tumor biopsy analysis revealed that bMMRD/polymerase-mutant cancers rapidly amass an excess of simultaneous mutations (∼600 mutations/cell division), reaching but not exceeding ∼20,000 exonic mutations in <6 months. This implies a threshold compatible with cancer-cell survival. We suggest a new mechanism of cancer progression in which mutations develop in a rapid burst after ablation of replication repair.
Low dietary intake of β-carotene is associated with chronic disease and vitamin A deficiency. β-Carotene is converted to vitamin A in the intestine by the enzyme β-carotene-15,15′-monoxygenase ...(BCMO1) to support vision, reproduction, immune function, and cell differentiation. Considerable variability for this key step in vitamin A metabolism, as reported in the human population, could be related to genetics and individual vitamin A status, but it is unclear how these factors influence β-carotene metabolism and vitamin A homeostasis. Here we show that the intestine-specific transcription factor ISX binds to the Bcmo1 promoter. Moreover, upon induction by the β-carotene derivative retinoic acid, this ISX binding decreased expression of a luciferase reporter gene in human colonic CaCo-2 cells indicating that ISX acts as a transcriptional repressor of BCMO1 expression. Mice deficient for this transcription factor displayed increased intestinal BCMO1 expression and produced significantly higher amounts of vitamin A from supplemental β-carotene. The ISX binding site in the human BCMO1 promoter contains a common single nucleotide polymorphism that is associated with decreased conversion rates and increased fasting blood levels of β-carotene. Thus, our study establishes ISX as a critical regulator of vitamin A production and provides a mechanistic explanation for how both genetics and diet can affect this process.
Background: Dietary β-carotene is the natural precursor of vitamin A.
Results: The retinoic acid-inducible homeobox transcription factor ISX controls intestinal expression of the vitamin A producing enzyme BCMO1.
Conclusion: Vitamin A production is under negative feedback regulation.
Significance: Large individual variability in intestinal β-carotene conversion is associated with this diet-responsive regulatory network.
The synthesis, characterization, and thermogravimetric analysis of tris(N,N′-di-isopropylacetamidinate)molybdenum(III), Mo(iPr-AMD)3, are reported. Mo(iPr-AMD)3 is a rare example of a homoleptic ...mononuclear complex of molybdenum(III) and fills a longstanding gap in the literature of transition metal(III) trisamidinate complexes. Thermogravimetric analysis (TGA) reveals excellent volatilization at elevated temperatures, pointing to potential applications as a vapor phase precursor for higher temperature atomic layer deposition (ALD), or chemical vapor deposition (CVD) growth of Mo-based materials. The measured TGA temperature window was 200–314 °C for samples in the 3–20 mg range. To validate the utility of Mo(iPr-AMD)3, we demonstrate aerosol-assisted CVD growth of MoO3 from benzonitrile solutions of Mo(iPr-AMD)3 at 500 °C using compressed air as the carrier gas. The resulting films are characterized by X-ray photoelectron spectroscopy, X-ray diffraction, and Raman spectroscopy. We further demonstrate the potential for ALD growth at 200 °C with a Mo(iPr-AMD)3/Ar purge/300 W O2 plasma/Ar purge sequence, yielding ultrathin films which retain a nitride/oxynitride component. Our results highlight the broad scope utility and potential of Mo(iPr-AMD)3 as a stable, high-temperature precursor for both CVD and ALD processes.
Ligands of intermediate steric bulk were designed to mimic metalloenzymes with histidine and carboxlyate binding sites. The reaction between tris(3-isopropylpyrazolyl)methane and butyllithium ...followed by SO3NMe3 in THF yielded the new ligand lithium tris(3-isopropylpyrazolyl)methane sulfonate (LiTpmsiPr). Various metal salts reacted with LiTpmsiPr to give the octahedral complexes M(TpmsiPr)2 (M = Zn, Cu, Ni, Co, Fe) in which each ligand has N,N,O binding to the metal. In the reaction between LiTpmsiPr and ZnCl2, in addition to the major product Zn(TpmsiPr)2, LiTpmsiPrZnCl2·2THF was also formed as a minor product with a tetrahedral zinc atom coordinated to either N,N,Cl,Cl in the solid phase or N,N,N,Cl in acetonitrile solution. Although TpmsiPr is coordinatively flexible and can act as a bipodal or tripodal ligand, it appears to favor the formation of octahedral L2M complexes.
Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a disorder of cardiomyocyte intercalated disk proteins causing sudden death. Heterozygous mutations of the desmosomal protein plakophilin‐2 ...(PKP‐2) are the commonest genetic cause of ARVC. Abnormal gap junction connexin43 expression has been reported in autosomal dominant forms of ARVC (Naxos and Carvajal disease) caused by homozygous mutations of desmosomal plakoglobin and desmoplakin. In tissue culture, suppression of PKP‐2 results in decreased expression of connexin43. We sought to characterize the expression and localization of connexin43 in patients with ARVC secondary to heterozygous PKP‐2 mutations. Complete PKP‐2 gene sequencing of 27 ARVC patients was utilized to identify mutant genotypes. Endomyocardial biopsies of identified carriers were then assessed by immunofluorescence to visualize intercalated disk proteins. N‐cadherin was targeted to highlight intercalated disks, followed by counterstaining for PKP‐2 or connexin43 using confocal double immunofluorescence microscopy. Immunofluorescence was quantified using an Adobe® Photoshop protocol, and colocalization coefficients were determined. PKP‐2 siRNA experiments were performed in mouse cardiomyocyte (HL1) cell culture with Western blot analysis to assess connexin43 expression following PKP‐2 suppression. Missense and frameshift mutations of the PKP‐2 gene were found in four patients with biopsy material available for analysis. Immunofluorescent studies showed PKP‐2 localization to the intercalated disk despite mutations, but associated with decreased connexin43 expression and abnormal colocalization. PKP‐2 siRNA in HL1 culture confirmed decreased connexin43 expression. Reduced connexin43 expression and localization to the intercalated disk occurs in heterozygous human PKP‐2 mutations, potentially explaining the delayed conduction and propensity to develop arrhythmias seen in this disease.
Maternal anti-Ro and anti-La antibodies are associated with congenital heart block (CHB). Although endocardial fibroelastosis (EFE) has been described in isolated cases of autoantibody-mediated CHB, ...the natural history and pathogenesis of this disease are poorly understood.
We retrospectively reviewed the clinical history, echocardiography, and pathology of fetuses and children with EFE associated with CHB born to mothers positive for anti-Ro or anti-La antibodies at 5 centers. Thirteen patients were identified, 6 with a prenatal and 7 with a postnatal diagnosis. Six mothers were positive for anti-Ro and anti-La antibodies, and 7 were positive for anti-Ro antibodies only. Only 1 mother had autoimmune disease. Severe ventricular dysfunction was seen in all fetal and postnatal cases. Four fetal and 3 postnatal cases had EFE at initial presentation. However, 2 fetal and 4 postnatal cases developed EFE 6 to 12 weeks and 7 months to 5 years from CHB diagnosis, respectively, even despite ventricular pacing in 6 postnatal cases. Eleven (85%) either died (n=9) or underwent cardiac transplantation (n=2) secondary to the EFE. Pathologic assessment of the explanted heart, available in 10 cases, revealed moderate to severe EFE in 7 and mild EFE in 3 cases, predominantly involving the left ventricle. Immunohistochemistry in 4 cases (including 3 fetuses) demonstrated deposition of IgG in 4 and IgM in 3 and T-cell infiltrates in 3 cases, suggesting an immune response by the affected fetus or child.
EFE occurs in the presence of autoantibody-mediated CHB despite adequate ventricular pacing. Autoantibody-associated EFE has a very high mortality rate, whether developing in fetal or postnatal life.
Indirect modification of animal genomes by interspecific hybridization, cross-breeding, and selection has produced an enormous spectrum of phenotypic diversity over more than 10,000 yr of animal ...domestication. Using these established technologies, the farming community has successfully increased the yield and efficiency of production in most agricultural species while utilizing land resources that are often unsuitable for other agricultural purposes. Moving forward, animal well-being and agricultural sustainability are moral and economic priorities of consumers and producers alike. Therefore, these considerations will be included in any strategy designed to meet the challenges produced by global climate change and an expanding world population. Improvements in the efficiency and precision of genetic technologies will enable a timely response to meet the multifaceted food requirements of a rapidly increasing world population.
Heart disease accounts for a significant proportion of sudden unexpected deaths among children. We describe here demographic features, pathological conditions, and the frequency of premonitory ...symptoms in a retrospective series of cases of sudden unexpected cardiac death (SUCD) attributable to undiagnosed structural heart disease.
A chart review of autopsies involving children 0 to 17 years of age that were performed at the Hospital for Sick Children (Toronto, Ontario, Canada) between 1984 and 2003 was conducted. Cases of sudden unexpected death within 24 hours after clinical presentation with previously undetected fatal heart disease were included. Cases with multiple or thoracic trauma and chronic or multisystem disease were excluded.
During the 20-year study period, 4926 autopsies were performed. A total of 103 cases (2.1%), involving 51 male patients and 52 female patients 1 day to 15 years of age (mean: 2.9 ± 4.2 years), were diagnosed as having SUCD. The most common diagnoses were myocarditis (n = 37 35.9%), hypoplastic left heart syndrome (HLHS) (n = 19 18.4%), dilated cardiomyopathy (DCM) (n = 16 16.5%), coronary artery anomalies (n = 6 5.8%), and aortic stenosis (n = 5 4.9%). There was a significant difference in the mean age of presentation between leading causes of SUCD (6.5 days for HLHS, 1.7 years for DCM, and 5.4 years for myocarditis; P < .0001). Of 103 cases, 27 (26.2%) had premonitory symptoms documented.
SUCD accounted for 2.1% of all autopsies, and HLHS, DCM, and myocarditis were the 3 most common diagnoses, which presented at increasing ages.
This study was designed to document the association of endocardial fibroelastosis (EFE) and maternal autoantibodies.
Neonatal lupus erythematosus is associated with the transplacental passage of ...maternal anti-Ro and anti-La antibodies, leading to complete atrioventricular block (CAVB). In some cases, CAVB is associated with EFE. Isolated EFE may be independently related to maternal anti-Ro and anti-La antibodies.
We identified three cases (one fetus and two infants, all female) of isolated EFE in infants born to autoantibody-positive mothers in the absence of CAVB. Demographics, echocardiograms, and pathology were reviewed. Immunohistochemical analyses for immunoglobulin (Ig)G, IgM, IgA, T-cell, B-cell, and terminal deoxynucleoleotidyl transferase-mediated dUTP-biotin nick end-labeling (TUNEL) (test for cell apoptosis) staining were performed on multiple sections of the heart in each case and compared with negative controls.
Two cases died and one received a cardiac transplant. All three cases had histologically confirmed EFE. All cases demonstrated significant diffuse IgG infiltration. To a lesser extent, myocardial tissue was also positive for IgM, CD43, and Granzyme B. None of the specimens were TUNEL positive.
These are the first reported cases of isolated EFE associated with maternal anti-Ro and anti-La antibodies in the absence of CAVB. The diffuse deposition of IgG and the presence of a T-cell infiltrate throughout the myocardium suggest that the transplacental passage of maternal autoantibodies induces an immune reaction within the myocardium, leading to isolated EFE. Autoantibody-mediated EFE may be an etiologic factor in cases of fetal and neonatal “idiopathic” dilated cardiomyopathy.
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