Whole genome sequence of Treponema pallidum ssp. pallidum, strain Mexico A, suggests recombination between yaws and syphilis strains

Treponema pallidum ssp. pallidum (TPA), the causative agent of syphilis, and Treponema pallidum ssp. pertenue (TPE), the causative agent of yaws, are closely related spirochetes causing diseases with distinct clinical manifestations. The TPA Mexico A strain was isolated in 1953 from male, with primary syphilis, living in Mexico. Attempts to cultivate TPA Mexico A strain under in vitro conditions have revealed lower growth potential compared to other tested TPA strains.The complete genome sequence of the TPA Mexico A strain was determined using the Illumina sequencing technique. The genome sequence assembly was verified using the whole genome fingerprinting technique and the final sequence was annotated. The genome size of the Mexico A strain was determined to be 1,140,038 bp with 1,035 predicted ORFs. The Mexico A genome sequence was compared to the whole genome sequences of three TPA (Nichols, SS14 and Chicago) and three TPE (CDC-2, Samoa D and Gauthier) strains. No large rearrangements in the Mexico A genome were found and the identified nucleotide changes occurred most frequently in genes encoding putative virulence factors. Nevertheless, the genome of the Mexico A strain, revealed two genes (TPAMA_0326 (tp92) and TPAMA_0488 (mcp2-1)) which combine TPA- and TPE- specific nucleotide sequences. Both genes were found to be under positive selection within TPA strains and also between TPA and TPE strains.The observed mosaic character of the TPAMA_0326 and TPAMA_0488 loci is likely a result of inter-strain recombination between TPA and TPE strains during simultaneous infection of a single host suggesting horizontal gene transfer between treponemal subspecies

Resequencing of Treponema pallidum ssp. pallidum strains Nichols and SS14: Correction of sequencing errors resulted in increased separation of syphilis treponeme subclusters

Article on the resequencing of Treponema pallidum ssp. pallidum strains nichols and SS14 and how the correction of sequencing errors resulted in increased separation of syphilis treponeme subclusters

Effects of oxidation agents and metal ions on binding of p53 to supercoiled DNA

Wild type human full length (f.l.) tumor suppressor p53 protein binds preferentially to supercoiled (sc) DNA in vitro both in the presence and absence of the p53 consensus sequence (p53CON). This binding produces a ladder of retarded bands on the agarose gel. Bands revealed by immunoblotting with antibody DO-1 corresponded to the ethidium stained retarded bands. The intensity and the number of bands of p53-scDNA complex were decreased by physiological concentrations of unchelated zinc ions. Nickel and cobalt ions inhibited binding of p53 to scDNA and to p53CON in linear DNA fragments less efficiently than zinc. Compared to the intrinsic zinc strongly bound to Cys 176, Cys 238, Cys 242 and His 179 in the p53 core domain, binding of additional Zn2+ to p53 was much weaker as shown by an easy removal of the latter ions by low concentrations of EDTA. Oxidation of the protein with diamide resulted in a decrease of the number of the retarded bands. Under the same conditions, no binding of oxidized p53 to p53CON in a linear DNA fragment was observed. In agreement with the literature oxidation of f.l. p53 with diamide was irreversible and was not reverted by an excess of DTT. We showed that in the presence of 0.1 mM zinc ions, oxidation of p53 became reversible. Other divalent cations tested (cadmium, cobalt, nickel) exhibited no such effect. We suggested that the irreversibility of p53 oxidation was due, at least in part, to the removal of intrinsic zinc from its position in the DNA binding domain (after oxidation of the three cysteines to which the zinc ion is coordinated in the reduced protein) accompanied by a change in the p53 conformation. Binding of C-terminal anti-p53 antibody also protected bacterially expressed protein against irreversible loss of activity due to diamide oxidation. Binding the human p53 core domain (segment 94-312) to scDNA greatly differed from that observed with the full-length p53. The core domain did not posses the ability to bind strongly to many sites in scDNA regardless of the presence or absence of p53CON suggesting involvement of some other domain (probably C-terminal) in binding of the full-length p53 to scDNA. Supershift experiments using antibodies against p53 N- or C-terminus suggested that in oxidized p53, scDNA binding through the C-terminus gained importance

Recognition and coacervation of G-quadruplexes by a multifunctional disordered region in RECQ4 helicase

Biomolecular polyelectrolyte complexes can be formed between oppositely charged intrinsically disordered regions (IDRs) of proteins or between IDRs and nucleic acids. Highly charged IDRs are abundant in the nucleus, yet few have been functionally characterized. Here, we show that a positively charged IDR within the human ATP-dependent DNA helicase Q4 (RECQ4) forms coacervates with G-quadruplexes (G4s). We describe a three-step model of charge-driven coacervation by integrating equilibrium and kinetic binding data in a global numerical model. The oppositely charged IDR and G4 molecules form a complex in the solution that follows a rapid nucleation-growth mechanism leading to a dynamic equilibrium between dilute and condensed phases. We also discover a physical interaction with Replication Protein A (RPA) and demonstrate that the IDR can switch between the two extremes of the structural continuum of complexes. The structural, kinetic, and thermodynamic profile of its interactions revealed a dynamic disordered complex with nucleic acids and a static ordered complex with RPA protein. The two mutually exclusive binding modes suggest a regulatory role for the IDR in RECQ4 function by enabling molecular handoffs. Our study extends the functional repertoire of IDRs and demonstrates a role of polyelectrolyte complexes involved in G4 binding

Whole genome sequence of the Treponema Fribourg- Blanc: Unspecified simian isolate is highly similar to the yaws subspecies

BACKGROUND:Unclassified simian strain Treponema Fribourg-Blanc was isolated in 1966 from baboons (Papio cynocephalus) in West Africa. This strain was morphologically indistinguishable from T. pallidum ssp. pallidum or ssp. pertenue strains, and it was shown to cause human infections. METHODOLOGY/PRINCIPAL FINDINGS:To precisely define genetic differences between Treponema Fribourg-Blanc (unclassified simian isolate, FB) and T. pallidum ssp. pertenue strains (TPE), a high quality sequence of the whole Fribourg-Blanc genome was determined with 454-pyrosequencing and Illumina sequencing platforms. Combined average coverage of both methods was greater than 500×. Restriction target sites (n = 1,773), identified in silico, of selected restriction enzymes within the Fribourg-Blanc genome were verified experimentally and no discrepancies were found. When compared to the other three sequenced TPE genomes (Samoa D, CDC-2, Gauthier), no major genome rearrangements were found. The Fribourg-Blanc genome clustered with other TPE strains (especially with the TPE CDC-2 strain), while T. pallidum ssp. pallidum strains clustered separately as well as the genome of T. paraluiscuniculi strain Cuniculi A. Within coding regions, 6 deletions, 5 insertions and 117 substitutions differentiated Fribourg-Blanc from other TPE genomes. CONCLUSIONS/SIGNIFICANCE:The Fribourg-Blanc genome showed similar genetic characteristics as other TPE strains. Therefore, we propose to rename the unclassified simian isolate to Treponema pallidum ssp. pertenue strain Fribourg-Blanc. Since the Fribourg-Blanc strain was shown to cause experimental infection in human hosts, non-human primates could serve as possible reservoirs of TPE strains. This could considerably complicate recent efforts to eradicate yaws. Genetic differences specific for Fribourg-Blanc could then contribute for identification of cases of animal-derived yaws infections

Brown Carbon in Primary and Aged Coal Combustion Emission

Smog chamber experiments were conducted to characterize the light absorption of brown carbon (BrC) from primary and photochemically aged coal combustion emissions. Light absorption was measured by the UV-visible spectrophotometric analysis of water and methanol extracts of filter samples. The single-scattering albedo at 450 nm was 0.73 ± 0.10 for primary emissions and 0.75 ± 0.13 for aged emissions. The light absorption coefficient at 365 nm of methanol extracts was higher than that of water extracts by a factor of 10 for primary emissions and a factor of 7 for aged emissions. This suggests that the majority of BrC is water-insoluble even after aging. The mass absorption efficiency of this BrC (MAE365) for primary OA (POA) was dependent on combustion conditions, with an average of 0.84 ± 0.54 m2 g-1, which was significantly higher than that for aged OA (0.24 ± 0.18 m2 g-1). Secondary OA (SOA) dominated aged OA and the decreased MAE365 after aging indicates that SOA is less light absorbing than POA and/or that BrC is bleached (oxidized) with aging. The estimated MAE365 of SOA (0.14 ± 0.08 m2 g-1) was much lower than that of POA. A comparison of MAE365 of residential coal combustion with other anthropogenic sources suggests that residential coal combustion emissions are among the strongest absorbing BrC organics

Nuclear inclusions of pathogenic ataxin-1 induce oxidative stress and perturb the protein synthesis machinery

Spinocerebellar ataxia type-1 (SCA1) is caused by an abnormally expanded polyglutamine (polyQ) tract in ataxin-1. These expansions are responsible for protein misfolding and self-assembly into intranuclear inclusion bodies (IIBs) that are somehow linked to neuronal death. However, owing to lack of a suitable cellular model, the downstream consequences of IIB formation are yet to be resolved. Here, we describe a nuclear protein aggregation model of pathogenic human ataxin-1 and characterize IIB effects. Using an inducible Sleeping Beauty transposon system, we overexpressed the ATXN1(Q82) gene in human mesenchymal stem cells that are resistant to the early cytotoxic effects caused by the expression of the mutant protein. We characterized the structure and the protein composition of insoluble polyQ IIBs which gradually occupy the nuclei and are responsible for the generation of reactive oxygen species. In response to their formation, our transcriptome analysis reveals a cerebellum-specific perturbed protein interaction network, primarily affecting protein synthesis. We propose that insoluble polyQ IIBs cause oxidative and nucleolar stress and affect the assembly of the ribosome by capturing or down-regulating essential components. The inducible cell system can be utilized to decipher the cellular consequences of polyQ protein aggregation. Our strategy provides a broadly applicable methodology for studying polyQ diseases

Deficiency and haploinsufficiency of histone macroH2A1.1 in mice recapitulate hematopoietic defects of human myelodysplastic syndrome

Background: Epigenetic regulation is important in hematopoiesis, but the involvement of histone variants is poorly understood. Myelodysplastic syndromes (MDS) are heterogeneous clonal hematopoietic stem cell (HSC) disorders characterized by ineffective hematopoiesis. MacroH2A1.1 is a histone H2A variant that negatively correlates with the self-renewal capacity of embryonic, adult, and cancer stem cells. MacroH2A1.1 is a target of the frequent U2AF1 S34F mutation in MDS. The role of macroH2A1.1 in hematopoiesis is unclear. Results: MacroH2A1.1 mRNA levels are significantly decreased in patients with low-risk MDS presenting with chromosomal 5q deletion and myeloid cytopenias and tend to be decreased in MDS patients carrying the U2AF1 S34F mutation. Using an innovative mouse allele lacking the macroH2A1.1 alternatively spliced exon, we investigated whether macroH2A1.1 regulates HSC homeostasis and differentiation. The lack of macroH2A1.1 decreased while macroH2A1.1 haploinsufficiency increased HSC frequency upon irradiation. Moreover, bone marrow transplantation experiments showed that both deficiency and haploinsufficiency of macroH2A1.1 resulted in enhanced HSC differentiation along the myeloid lineage. Finally, RNA-sequencing analysis implicated macroH2A1.1-mediated regulation of ribosomal gene expression in HSC homeostasis. Conclusions: Together, our findings suggest a new epigenetic process contributing to hematopoiesis regulation. By combining clinical data with a discrete mutant mouse model and in vitro studies of human and mouse cells, we identify macroH2A1.1 as a key player in the cellular and molecular features of MDS. These data justify the exploration of macroH2A1.1 and associated proteins as therapeutic targets in hematological malignancies

Highly time-resolved chemical speciation and source apportionment of organic aerosol components in Delhi, India, using extractive electrospray ionization mass spectrometry

In recent years, the Indian capital city of Delhi has been impacted by very high levels of air pollution, especially during winter. Comprehensive knowledge of the composition and sources of the organic aerosol (OA), which constitutes a substantial fraction of total particulate mass (PM) in Delhi, is central to formulating effective public health policies. Previous source apportionment studies in Delhi identified key sources of primary OA (POA) and showed that secondary OA (SOA) played a major role but were unable to resolve specific SOA sources. We address the latter through the first field deployment of an extractive electrospray ionization time-of-flight mass spectrometer (EESI-TOF) in Delhi, together with a high-resolution aerosol mass spectrometer (AMS). Measurements were conducted during the winter of 2018/19, and positive matrix factorization (PMF) was used separately on AMS and EESI-TOF datasets to apportion the sources of OA. AMS PMF analysis yielded three primary and two secondary factors which were attributed to hydrocarbon-like OA (HOA), biomass burning OA (BBOA-1 and BBOA-2), more oxidized oxygenated OA (MO-OOA), and less oxidized oxygenated OA (LO-OOA). On average, 40 % of the total OA mass was apportioned to the secondary factors. The SOA contribution to total OA mass varied greatly between the daytime (76.8 %, 10:00–16:00 local time (LT)) and nighttime (31.0 %, 21:00–04:00 LT). The higher chemical resolution of EESI-TOF data allowed identification of individual SOA sources. The EESI-TOF PMF analysis in total yielded six factors, two of which were primary factors (primary biomass burning and cooking-related OA). The remaining four factors were predominantly of secondary origin: aromatic SOA, biogenic SOA, aged biomass burning SOA, and mixed urban SOA. Due to the uncertainties in the EESI-TOF ion sensitivities, mass concentrations of EESI-TOF SOA-dominated factors were related to the total AMS SOA (i.e. MO-OOA + LO-OOA) by multiple linear regression (MLR). Aromatic SOA was the major SOA component during the daytime, with a 55.2 % contribution to total SOA mass (42.4 % contribution to total OA). Its contribution to total SOA, however, decreased to 25.4 % (7.9 % of total OA) during the nighttime. This factor was attributed to the oxidation of light aromatic compounds emitted mostly from traffic. Biogenic SOA accounted for 18.4 % of total SOA mass (14.2 % of total OA) during the daytime and 36.1 % of total SOA mass (11.2 % of total OA) during the nighttime. Aged biomass burning and mixed urban SOA accounted for 15.2 % and 11.0 % of total SOA mass (11.7 % and 8.5 % of total OA mass), respectively, during the daytime and 15.4 % and 22.9 % of total SOA mass (4.8 % and 7.1 % of total OA mass), respectively, during the nighttime. A simple dilution–partitioning model was applied on all EESI-TOF factors to estimate the fraction of observed daytime concentrations resulting from local photochemical production (SOA) or emissions (POA). Aromatic SOA, aged biomass burning, and mixed urban SOA were all found to be dominated by local photochemical production, likely from the oxidation of locally emitted volatile organic compounds (VOCs). In contrast, biogenic SOA was related to the oxidation of diffuse regional emissions of isoprene and monoterpenes. The findings of this study show that in Delhi, the nighttime high concentrations are caused by POA emissions led by traffic and biomass burning and the daytime OA is dominated by SOA, with aromatic SOA accounting for the largest fraction. Because aromatic SOA is possibly more toxic than biogenic SOA and primary OA, its dominance during the daytime suggests an increased OA toxicity and health-related consequences for the general public.</p

Post-Transcriptional Regulation of BCL2 mRNA by the RNA-Binding Protein ZFP36L1 in Malignant B Cells

The human ZFP36 zinc finger protein family consists of ZFP36, ZFP36L1, and ZFP36L2. These proteins regulate various cellular processes, including cell apoptosis, by binding to adenine uridine rich elements in the 3′ untranslated regions of sets of target mRNAs to promote their degradation. The pro-apoptotic and other functions of ZFP36 family members have been implicated in the pathogenesis of lymphoid malignancies. To identify candidate mRNAs that are targeted in the pro-apoptotic response by ZFP36L1, we reverse-engineered a gene regulatory network for all three ZFP36 family members using the ‘maximum information coefficient’ (MIC) for target gene inference on a large microarray gene expression dataset representing cells of diverse histological origin. Of the three inferred ZFP36L1 mRNA targets that were identified, we focussed on experimental validation of mRNA for the pro-survival protein, BCL2, as a target for ZFP36L1. RNA electrophoretic mobility shift assay experiments revealed that ZFP36L1 interacted with the BCL2 adenine uridine rich element. In murine BCL1 leukemia cells stably transduced with a ZFP36L1 ShRNA lentiviral construct, BCL2 mRNA degradation was significantly delayed compared to control lentiviral expressing cells and ZFP36L1 knockdown in different cell types (BCL1, ACHN, Ramos), resulted in increased levels of BCL2 mRNA levels compared to control cells. 3′ untranslated region luciferase reporter assays in HEK293T cells showed that wild type but not zinc finger mutant ZFP36L1 protein was able to downregulate a BCL2 construct containing the BCL2 adenine uridine rich element and removal of the adenine uridine rich core from the BCL2 3′ untranslated region in the reporter construct significantly reduced the ability of ZFP36L1 to mediate this effect. Taken together, our data are consistent with ZFP36L1 interacting with and mediating degradation of BCL2 mRNA as an important target through which ZFP36L1 mediates its pro-apoptotic effects in malignant B-cells