Biodeterioration of Mayan buildings at uxmal and tulum,Mexico

Uxmal and Tulum are two important Mayan sites in the Yucatan peninsula. The buildings are mainly composed of limestone and grey/black discoloration is seen on exposed walls and copious greenish biofilms on inner walls. The principal microorganisms detected on interior walls at both Uxmal and Tulum were cyanobacteria; heterotrophic bacteria and filamentous fungi were also present. A dark‐pigmented mitosporic fungus and Bacillus cereus, both isolated from Uxmal, were shown to be acidogenic in laboratory cultures. Cyanobacteria belonging to rock‐degrading genera Synechocystis and Gloeocapsa were identified at both sites. Surface analysis previously showed that calcium ions were present in the biofilms on buildings at Uxmal and Tulum, suggesting the deposition of biosolubilized stone. Apart from their potential to degrade the substrate, the coccoid cyanobacteria supply organic nutrients for bacteria and fungi, which can produce organic acids, further increasing stone degradation.     

Roles of Saccharomyces cerevisiae DNA polymerases Poleta and Polzeta in response to irradiation by simulated sunlight

Sunlight causes lesions in DNA that if unrepaired and inaccurately replicated by DNA polymerases yield mutations that result in skin cancer in humans. Two enzymes involved in translesion synthesis (TLS) of UV-induced photolesions are DNA polymerase η (Polη) and polymerase ζ (Polζ), encoded by the RAD30A and REV3 genes, respectively. Previous studies have investigated the TLS roles of these polymerases in human and yeast cells irradiated with monochromatic, short wavelength UVC radiation (254 nm). However, less is known about cellular responses to solar radiation, which is of higher and mixed wavelengths (310–1100 nm) and produces a different spectrum of DNA lesions, including Dewar photoproducts and oxidative lesions. Here we report on the comparative cytotoxic and mutagenic effects of simulated sunlight (SSL) and UVC radiation on yeast wild-type, rad30Δ, rev3Δ and rev3Δ rad30Δ strains. The results with SSL support several previous interpretations on the roles of these two polymerases in TLS of photodimers and (6–4) photoproducts derived from studies with UVC. They further suggest that Polη participates in the non-mutagenic bypass of SSL-dependent cytosine-containing Dewar photoproducts and 8-oxoguanine, while Polζ is mainly responsible for the mutagenic bypass of all types of Dewar photoproducts. They also suggest that in the absence of Polζ, Polη contributes to UVC- and SSL-induced mutagenesis, possibly by the bypass of photodimers containing deaminated cytosine.     

Hydrodynamic parameters of native C-reactive protein molecule in a solution

The hydrodynamic properties of the C-reactive protein in solution (pH 6.8) were studied using quasi-elastic light scattering and size-exclusion liquid chromatography. It was shown that the solution containing the C-reactive protein represents a polydisperse system. The values of the translation diffusion coefficient and the apparent molecular weight of the C-reactive protein in solution at pH 6.8 were determined. The values of the translation diffusion coefficient, molecular weight and the hydration radius obtained suggest that the native pentameric C-reactive protein is the major form of the protein in solution at pH 6.8.     

Aggregation of C-reactive protein in solutions at acid pH

The hydrodynamic properties of the C-reactive protein (CRP) at different pH were studied using quasi-elastic light scattering, size-exclusion liquid chromatography, and nonreducing gel electrophoresis. It was shown that a CRP solution at pH 5.0-7.2 presents a polydisperse system the major component of which is the native pentameric CRP. At pH 4.0-4.5, CRP exists in two states having different hydrodynamic properties: the native pentameric form with a molecular mass of 120 kDa and with the hydrodynamic radius of 4.03 nm and high-molecular-weight aggregates with a wide range of their molecular weight distribution. The interaction of the C-reactive protein with monoclonal antibodies to it indicates that conformation-dependent surface epitopes of the protein lose the native structure at pH 5.0-5.5. The aggregation of CRP is an irreversible process, which begins in a narrow pH range of pH 5.0-4.5 and is not accompanied by the dissociation into subunits but is determined by intermolecular interactions of its quasi-native pentamers.     

Role for CysJ Flavin Reductase in Molybdenum Cofactor-Dependent Resistance of Escherichia coli to 6-N-Hydroxylaminopurine

We have previously described a novel Escherichia coli detoxification system for the removal of toxic and mutagenic N-hydroxylated nucleobases and related compounds that requires the molybdenum cofactor. Two subpathways (ycbX and yiiM) were identified, each employing a novel molybdo activity capable of inactivating N-hydroxylated compounds by reduction to the corresponding amine. In the present study, we identify the cysJ gene product as one additional component of this system. While the CysJ protein has been identified as the NADPH:flavin oxidoreductase component of the CysJI sulfite reductase complex (CysJ₈I₄), we show that the role of CysJ in base analog detoxification is unique and independent of CysI and sulfite reductase. We further show that CysJ functions as a specific partner of the YcbX molybdoenzyme. We postulate that the function of CysJ in this pathway is to provide, via its NADPH:flavin reductase activity, the reducing equivalents needed for the detoxification reaction at the YcbX molybdocenter. In support of the proposed interaction of the CysJ and YcbX proteins, we show that an apparent CysJ-YcbX “hybrid” protein from two Vibrio species is capable of compensating for a double cysJ ycbX defect in E. coli.Mutagenic base analogs are chemically modified nucleobases that can be incorporated in the cellular metabolism through purine or pyrimidine salvage pathways. Once converted to the deoxynucleoside triphosphate (dNTP) level, they may participate in DNA replication in an error-prone manner because of their ambivalent base-pairing capacity (11). Such synthetic base analogs are often used as a sensitive tool for studying DNA replication fidelity, DNA repair, or the metabolism of nucleic acid precursors. Mutagenic base analogs such as 8-oxoguanine or 3-methyladenine can also be formed in vivo as a consequence of normal cellular metabolism or produced by chemical and physical factors, such as alkylating agents or ionizing radiation.An important group of mutagenic and cytotoxic analogs are the N-hydroxylated nucleobases (or ribosides) such as 6-N-hydroxylaminopurine (HAP), 2-amino-HAP, or N⁴-hydroxycytidine (15). Specifically, HAP was found to be a very strong mutagen in bacteria and fungi, as well as mammalian cells (2, 20, 27). Some data have suggested that HAP may also be formed in vivo under oxidative stress (30) or as a by-product of certain purine salvage/interconversion pathways (5, 22).The genetic control of HAP-induced mutagenesis has been studied in some detail in the yeast Saccharomyces cerevisiae and in the bacterium Escherichia coli. In S. cerevisiae, resistance to HAP depends primarily on genes involved in adjusting and regulating the DNA or RNA precursor pools (HAM1 [ITP/XTPase], AAH1 [adenine aminohydrolase], and ADE genes involved in de novo AMP biosynthesis) (34).In E. coli, the major pathway that protects cells against HAP and related N-hydroxylated compounds is controlled by the moa, moe, and mog genes, which are required for biosynthesis of molybdenum cofactor (MoCo) (18, 19). MoCo is an essential cofactor for a varied group of oxidoreductases that are widely distributed from bacteria to humans. Chemically, MoCo is a pterin derivative (molybdopterin) that coordinates a molybdenum atom that serves as a catalytic redox center (for reviews, see references 23, 28, and 29). Based on catalytic details and sequence homology, molybdopterin-containing enzymes have been divided in four families: the xanthine oxidase family, the sulfite oxidase family, the dimethyl sulfoxide (DMSO) reductase family, and the aldehyde ferredoxin oxidoreductase family (14, 16). However, our previous studies on the MoCo-dependent resistance to HAP showed that none of the known or putative E. coli members of these families are responsible for the major HAP resistance mechanism (19). Instead, we discovered that HAP resistance is dependent on two newly described proteins, YcbX and YiiM, that are characterized by a so-called MOSC domain (molybdenum cofactor sulfurase C-terminal domain) (1, 17). This domain was first described as part of eukaryotic MoCo sulfurases (MOSs) (1), and it most likely represents a novel class of MoCo-binding domain, as indicated by studies on two mammalian MOSC-containing proteins (mARC1 and mARC2) discovered in mitochondria (12, 13).Our studies in E. coli showed that cell-free bacterial extracts were capable of converting HAP to adenine by an N-reductive reaction (17). Importantly, this conversion was entirely dependent on the presence of MoCo and the YcbX or YiiM proteins (17). Consequently, we suggested that this reduction of HAP to adenine forms the basis of the in vivo MoCo-dependent detoxification in E. coli (17). Interestingly, the mammalian MOSC-containing proteins mARC1 and mARC2 were shown to mediate the reduction of the N-hydroxylated prodrug benzamidoxime to its active amino form benzamidine (12, 13). Thus, the reduction of N-hydroxylated compounds may be a defining feature for the broadly distributed MOSC proteins (1).Our previous analyses also revealed that the E. coli ycbX and yiiM genes define two independent subpathways within the MoCo-dependent system (17). This is illustrated in the overall scheme shown in Fig. ​Fig.1.1. MoCo is synthesized in a series of steps from GTP by-products of the moa, moe, and mog operons. MoCo is then used as a cofactor for the YcbX and YiiM proteins, which reduce the N-hydroxylated compound to the corresponding amino form. The ycbX and yiiM pathways are genetically distinct as determined by epistasis experiments (17). They also differ by their substrate specificity patterns: YcbX protects most strongly against HAP, whereas YiiM has its largest effects toward hydroxylamine (NH₂OH) (17).Open in a separate windowFIG. 1.Genetic framework for the major molybdenum cofactor (MoCo)-dependent pathways of detoxification of N-hydroxylated base analogs in E. coli (17). moaA to mogA indicate the series of genes required for MoCo biosynthesis (19, 28), while ycbX and yiiM represent the two independent subpathways identified within the MoCo-dependent pathway (17). Specifically, ycbX and yiiM produce apoenzymes that react with MoCo to form the active YcbX and YiiM proteins. The diagram also indicates the differential specificity of the two subpathways toward the model N-hydroxylated compounds used in our studies: 6-N-hydroxylaminopurine (HAP), 2-amino-HAP (AHAP), and hydroxylamine (NH₂OH). For simplicity, the diagram does not distinguish between the MPT and MGD forms of MoCo (19). As shown elsewhere (19), YcbX and YiiM likely employ the MPT form. One additional, minor pathway for HAP detoxification dependent on biotin sulfoxide reductase (an MGD-requiring enzyme) is observable only in the double ycbX yiiM-deficient background and is likewise not shown here (see reference 17 for details).Prior to the establishment of this scheme of YcbX and YiiM as molybdoproteins, we had entertained certain alternative possibilities for the precise function of the ycbX and yiiM open reading frames (ORFs), including a possible role in MoCo sulfuration (which is a required modification of MoCo in certain molybdoenzymes, such as xanthine oxidase) (23, 29). This sulfuration model was ultimately eliminated (17), but certain experiments related to this hypothesis yielded interesting further clues regarding the detailed mechanisms of HAP resistance. These observations included an unexpected HAP-sensitive phenotype for cysJ mutants as well as a noted sensitization of wild-type strains to HAP by l-cysteine. In the present work, we describe these experiments and show the cysJ gene to be an essential component of the ycbX branch of HAP resistance. In a related mechanism, the observed sensitization of wild-type strains by l-cysteine results from the suppression, by l-cysteine, of the cys regulon. Overall, our experiments suggest that CysJ is a specific protein partner of YcbX and that CysJ mediates the N-reductive reaction through its NADPH:flavin oxidoreductase activity. This activity provides reducing equivalents to its partner YcbX, which ultimately performs the reduction of HAP to nontoxic adenine at its molybdocenter.     

YcbX and yiiM, two novel determinants for resistance of Escherichia coli to N-hydroxylated base analogues

We have shown previously that lack of molybdenum cofactor (MoCo) in Escherichia coli leads to hypersensitivity to the mutagenic and toxic effects of N -hydroxylated base analogues, such as 6- N -hydroxylaminopurine (HAP). However, the nature of the MoCo-dependent mechanism is unknown, as inactivation of all known and putative E. coli molybdoenzymes does not produce any sensitivity. Presently, we report on the isolation and characterization of two novel HAP-hypersensitive mutants carrying defects in the ycbX or yiiM open reading frames. Genetic analysis suggests that the two genes operate within the MoCo-dependent pathway. In the absence of the ycbX - and yiiM -dependent pathways, biotin sulfoxide reductase plays also a role in the detoxification pathway. YcbX and YiiM are hypothetical members of the MOSC protein superfamily, which contain the C-terminal domain (MOSC) of the eukaryotic MoCo sulphurases. However, deletion of ycbX or yiiM did not affect the activity of human xanthine dehydrogenase expressed in E. coli , suggesting that the role of YcbX and YiiM proteins is not related to MoCo sulphuration. Instead, YcbX and YiiM may represent novel MoCo-dependent enzymatic activities. We also demonstrate that the MoCo/YcbX/YiiM-dependent detoxification of HAP proceeds by reduction to adenine.     

Effect of Calcium Ions on Hydrodynamic Properties of Pentameric and Decameric C-Reactive Protein in Solution

The molecular mass and sedimentation coefficient of native C-reactive protein in solution were determined by analytical ultracentrifugation in the presence and absence of calcium ions. Pentameric C-reactive protein was shown to be the major macroscopic form of this protein in solution. The removal of calcium ions from solution caused decompaction of the protein accompanied by changes in its hydrodynamic parameters. The sedimentation coefficient s ⁰ _20,w of pentameric C-reactive protein in solution containing 2 mM Ca²⁺ (6.6S) exceeded that for C-reactive protein in solution containing 2 mM EDTA (6.4S). Analysis of average molecular masses M _w and M _z obtained from sedimentation data demonstrated that the solution of highly purified protein was not homogeneous. As shown by intermolecular crosslinking, the solution also contained the 241-kDa decamer of C-reactive protein (9.5S) as a separate macroscopic form, whose share hardly reached 10% in the presence of 2 mM Ca²⁺ and increased after removal of calcium ions. The decamers were shown to result from intermolecular association of the pentamers.     

Terrigenous sediment-dominated reef platform infilling: an unexpected precursor to reef island formation and a test of the reef platform size–island age model in the Pacific

Low-lying coral reef islands are considered highly vulnerable to climate change, necessitating an improved understanding of when and why they form, and how the timing of formation varies within and among regions. Several testable models have been proposed that explain inter-regional variability as a function of sea-level history and, more recently, a reef platform size model has been proposed from the Maldives (central Indian Ocean) to explain intra-regional (intra-atoll) variability. Here we present chronostratigraphic data from Pipon Island, northern Great Barrier Reef (GBR), enabling us to test the applicability of existing regional island evolution models, and the platform size control hypothesis in a Pacific context. We show that reef platform infilling occurred rapidly (~4–5 mm yr⁻¹) under a “bucket-fill” type scenario. Unusually, this infilling was dominated by terrigenous sedimentation, with platform filling and subsequent reef flat formation complete by ~5000 calibrated years BP (cal BP). Reef flat exposure as sea levels slowly fell post highstand facilitated a shift towards intertidal and subaerial-dominated sedimentation. Our data suggest, however, a lag of ~1500 yr before island initiation (at ~3200 cal BP), i.e. later than that reported from smaller and more evolutionarily mature reef platforms in the region. Our data thus support: (1) the hypothesis that platform size acts to influence the timing of platform filling and subsequent island development at intra-regional scales; and (2) the hypothesis that the low wooded islands of the northern GBR conform to a model of island formation above an elevated reef flat under falling sea levels.

     

Methylated DNA/RNA in Body Fluids as Biomarkers for Lung Cancer

DNA/RNA methylation plays an important role in lung cancer initiation and progression. Liquid biopsy makes use of cells, nucleotides and proteins released from tumor cells into body fluids to help with cancer diagnosis and prognosis. Methylation of circulating tumor DNA (ctDNA) has gained increasing attention as biomarkers for lung cancer. Here we briefly introduce the biological basis and detection method of ctDNA methylation, and review various applications of methylated DNA in body fluids in lung cancer screening, diagnosis, prognosis, monitoring and treatment prediction. We also discuss the emerging role of RNA methylation as biomarkers for cancer.     

Genetic Admixture in the Culturally Unique Peranakan Chinese Population in Southeast Asia

The Peranakan Chinese are culturally unique descendants of immigrants from China who settled in the Malay Archipelago ∼300–500 years ago. Today, among large communities in Southeast Asia, the Peranakans have preserved Chinese traditions with strong influence from the local indigenous Malays. Yet, whether or to what extent genetic admixture co-occurred with the cultural mixture has been a topic of ongoing debate. We performed whole-genome sequencing (WGS) on 177 Singapore (SG) Peranakans and analyzed the data jointly with WGS data of Asian and European populations. We estimated that Peranakan Chinese inherited ∼5.62% (95% confidence interval [CI]: 4.76–6.49%) Malay ancestry, much higher than that in SG Chinese (1.08%, 0.65–1.51%), southern Chinese (0.86%, 0.50–1.23%), and northern Chinese (0.25%, 0.18–0.32%). A sex-biased admixture history, in which the Malay ancestry was contributed primarily by females, was supported by X chromosomal variants, and mitochondrial (MT) and Y haplogroups. Finally, we identified an ancient admixture event shared by Peranakan Chinese and SG Chinese ∼1,612 (95% CI: 1,345–1,923) years ago, coinciding with the settlement history of Han Chinese in southern China, apart from the recent admixture event with Malays unique to Peranakan Chinese ∼190 (159–213) years ago. These findings greatly advance our understanding of the dispersal history of Chinese and their interaction with indigenous populations in Southeast Asia.