A Sb(III)-specific efflux transporter from Ensifer adhaerens E-60.

Antimony is pervasive environmental toxic substance, and numerous genes encoding mechanisms to resist, transform and extrude the toxic metalloid antimony have been discovered in various microorganisms. Here we identified a major facilitator superfamily (MFS) transporter, AntB, on the chromosome of the arsenite-oxidizing bacterium Ensifer adhaerens E-60 that confers resistance to Sb(III) and Sb(V). The antB gene is adjacent to gene encoding a LysR family transcriptional regulator termed LysRars, which is an As(III)/Sb(III)-responsive transcriptional repressor that is predicted to control expression of antB. Similar antB and lysRars genes are found in related arsenic-resistant bacteria, especially strains of Ensifer adhaerens, and the lysRars gene adjacent to antB encodes a member of a divergent subgroup of putative LysR-type regulators. Closely related AntB and LysRars orthologs contain three conserved cysteine residues, which are Cys17, Cys99, and Cys350 in AntB and Cys81, Cys289 and Cys294 in LysRars, respectively. Expression of antB is induced by As(III), Sb(III), Sb(V) and Rox(III) (4-hydroxy-3-nitrophenyl arsenite). Heterologous expression of antB in E. coli AW3110 (Δars) conferred resistance to Sb(III) and Sb(V) and reduced the intracellular concentration of Sb(III). The discovery of the Sb(III) efflux transporter AntB enriches our knowledge of the role of the efflux transporter in the antimony biogeochemical cycle.

Microbial diversity, genomics, and phage-host interactions of cyanobacterial harmful algal blooms.

The occurrence of cyanobacterial harmful algal blooms (cyanoHABs) is related to their physical and chemical environment. However, less is known about their associated microbial interactions and processes. In this study, cyanoHABs were analyzed as a microbial ecosystem, using 1 year of 16S rRNA sequencing and 70 metagenomes collected during the bloom season from Lake Okeechobee (Florida, USA). Biogeographical patterns observed in microbial community composition and function reflected ecological zones distinct in their physical and chemical parameters that resulted in bloom "hotspots" near major lake inflows. Changes in relative abundances of taxa within multiple phyla followed increasing bloom severity. Functional pathways that correlated with increasing bloom severity encoded organic nitrogen and phosphorus utilization, storage of nutrients, exchange of genetic material, phage defense, and protection against oxidative stress, suggesting that microbial interactions may promote cyanoHAB resilience. Cyanobacterial communities were highly diverse, with picocyanobacteria ubiquitous and oftentimes most abundant, especially in the absence of blooms. The identification of novel bloom-forming cyanobacteria and genomic comparisons indicated a functionally diverse cyanobacterial community with differences in its capability to store nitrogen using cyanophycin and to defend against phage using CRISPR and restriction-modification systems. Considering blooms in the context of a microbial ecosystem and their interactions in nature, physiologies and interactions supporting the proliferation and stability of cyanoHABs are proposed, including a role for phage infection of picocyanobacteria. This study displayed the power of "-omics" to reveal important biological processes that could support the effective management and prediction of cyanoHABs. IMPORTANCE: Cyanobacterial harmful algal blooms pose a significant threat to aquatic ecosystems and human health. Although physical and chemical conditions in aquatic systems that facilitate bloom development are well studied, there are fundamental gaps in the biological understanding of the microbial ecosystem that makes a cyanobacterial bloom. High-throughput sequencing was used to determine the drivers of cyanobacteria blooms in nature. Multiple functions and interactions important to consider in cyanobacterial bloom ecology were identified. The microbial biodiversity of blooms revealed microbial functions, genomic characteristics, and interactions between cyanobacterial populations that could be involved in bloom stability and more coherently define cyanobacteria blooms. Our results highlight the importance of considering cyanobacterial blooms as a microbial ecosystem to predict, prevent, and mitigate them.

Arsenate reductase of Rufibacter tibetensis is a metallophosphoesterase evolved to catalyze redox reactions.

An arsenate reductase (Car1) from the Bacteroidetes species Rufibacter tibetensis 1351T was isolated from the Tibetan Plateau. The strain exhibits resistance to arsenite [As(III)] and arsenate [As(V)] and reduces As(V) to As(III). Here we shed light on the mechanism of enzymatic reduction by Car1. AlphaFold2 structure prediction, active site energy minimization, and steady-state kinetics of wild-type and mutant enzymes give insight into the catalytic mechanism. Car1 is structurally related to calcineurin-like metallophosphoesterases (MPPs). It functions as a binuclear metal hydrolase with limited phosphatase activity, particularly relying on the divalent metal Ni2+. As an As(V) reductase, it displays metal promiscuity and is coupled to the thioredoxin redox cycle, requiring the participation of two cysteine residues, Cys74 and Cys76. These findings suggest that Car1 evolved from a common ancestor of extant phosphatases by incorporating a redox function into an existing MPP catalytic site. Its proposed mechanism of arsenate reduction involves Cys74 initiating a nucleophilic attack on arsenate, leading to the formation of a covalent intermediate. Next, a nucleophilic attack of Cys76 leads to the release of As(III) and the formation of a surface-exposed Cys74-Cys76 disulfide, ready for reduction by thioredoxin.

Anticancer Effects of the Trivalent Organoarsenical 2-Amino-4-(dihydroxyarsinoyl) Butanoate.

According to the National Cancer Institute, breast cancer is a leading cause of death in women. The lack of progesterone and estrogen receptors in triple-negative breast cancer (TNBC) cells results in a lack of response to hormonal, monoclonal, or tyrosine kinase inhibitor therapies. Despite intensive drug discovery, there is still no approved targeted treatment for TNBC. The metalloid arsenic has been used in herbal medicines, antibiotics, and chemotherapeutic drugs for centuries. This paper demonstrates that a trivalent arsenic-containing, nonproteogenic amino acid, R-AST-OH (2-amino-4-(dihydroxyarsinoyl) butanoate), inhibits kidney-type glutaminase (KGA), the enzyme that controls glutamine metabolism and is correlated with tumor malignancy. Cell-based assays using the TNBC MDA-MB-231 and HCC1569 cell lines showed that R-AST-OH kills TNBC cells and is not cytotoxic to a control cell line. The results of in silico molecular docking predictions indicate that R-AST-OH binds to the glutamine binding site and forms a covalent bond with an active site cysteine residue. We hypothesize that R-AST-OH is a single warhead for KGA that irreversibly binds to KGA through the formation of an As-S bond. We propose that R-AST-OH is a promising lead compound for the design of new drugs for the treatment of TNBC.

Impact of Pre-Diagnostic Risk Factors on Short- and Long-Term Ovarian Cancer Survival Trajectories: A Longitudinal Observational Study.

Tumor- and treatment-related factors are established predictors of ovarian cancer survival. New studies suggest a differential impact of exposures on ovarian cancer survival trajectories (i.e., rapidly fatal to long-term disease). This study examined the impact of pre-diagnostic risk factors on short- and long-term ovarian cancer survival trajectories in the Canadian context. This population-based longitudinal observational study included women diagnosed with invasive epithelial ovarian cancer from 1995 to 2004 in Ontario. Data were obtained from medical records, interviews, and the provincial cancer registry. Extended Cox proportional hazard models estimated the association between risk factors and all-cause and ovarian cancer-specific mortality by survival time intervals (<3 years (i.e., short-term survival), 3 to <6 years, 6 to <10 years, and ≥10 years (i.e., long-term survival)). Among 1421 women, histology, stage, and residual disease were the most important predictors of all-cause mortality in all survival trajectories, particularly for short-term survival. Reproductive and lifestyle factors did not strongly impact short-term overall survival but were associated with long-term overall survival. As such, among long-term survivors, history of breastfeeding significantly decreased the risk of all-cause mortality (HR 0.65; 95% CI 0.46, 0.93; p < 0.05), whereas smoking history (HR 1.75; 95% CI 1.27, 2.40; p < 0.05) and obesity (HR 1.81; 95% CI 1.24, 2.65; p < 0.05) significantly increased the risk of all-cause mortality. The findings were consistent with ovarian cancer-specific mortality. These findings suggest that pre-diagnostic exposures differentially influence survival time following a diagnosis of ovarian cancer.

Cervical cancer management in a low resource setting: A 10-year review in a tertiary care hospital in Kenya.

Background: Cervical cancer is one of the leading causes of cancer mortality among women in Kenya due to late presentations, poor access to health care, and limited resources. Across many low- and middle-income countries infrastructure and human resources for cervical cancer management are currently insufficient to meet the high population needs therefore patients are not able to get appropriate treatment. Objective: This study aimed to describe the clinicopathological characteristics and the treatment profiles of cervical cancer cases seen at Moi Teaching and Referral Hospital (MTRH). Methods: This was a retrospective cross-sectional study conducted at MTRH involving the review of the electronic database and medical charts of 1541 patients with a histologically confirmed diagnosis of cervical cancer between January 2012 and December 2021. Results: Of the 1541 cases analyzed, 91% were squamous cell carcinomas, 8% were adenocarcinomas, and 1% were other histological types. Thirty-eight percent of the patients were HIV infected and less than 30% of the women had health insurance. A majority (75%) of the patients presented with advanced-stage disease (stage IIB-IV). Only 13.9% received chemoradiotherapy with curative intent; of which 33.8% received suboptimal treatment. Of the 13% who received surgical treatment, 45.3% required adjuvant therapy, of which only 27.5% received treatment. Over 40% of the women were lost to follow-up. Conclusion: Most of the patients with cervical cancer in Kenya present at advanced stages with only a third receiving the necessary treatment while the majority receive only palliative treatment or supportive care.

Fundamentals and application in phytoremediation of an efficient arsenate reducing bacterium Pseudomonas putida ARS1.

Arsenic is a ubiquitous environmental pollutant. Microbe-mediated arsenic bio-transformations significantly influence arsenic mobility and toxicity. Arsenic transformations by soil and aquatic organisms have been well documented, while little is known regarding effects due to endophytic bacteria. An endophyte Pseudomonas putida ARS1 was isolated from rice grown in arsenic contaminated soil. P. putida ARS1 shows high tolerance to arsenite (As(III)) and arsenate (As(V)), and exhibits efficient As(V) reduction and As(III) efflux activities. When exposed to 0.6 mg/L As(V), As(V) in the medium was completely converted to As(III) by P. putida ARS1 within 4 hr. Genome sequencing showed that P. putida ARS1 has two chromosomal arsenic resistance gene clusters (arsRCBH) that contribute to efficient As(V) reduction and As(III) efflux, and result in high resistance to arsenicals. Wolffia globosa is a strong arsenic accumulator with high potential for arsenic phytoremediation, which takes up As(III) more efficiently than As(V). Co-culture of P. putida ARS1 and W. globosa enhanced arsenic accumulation in W. globosa by 69%, and resulted in 91% removal of arsenic (at initial concentration of 0.6 mg/L As(V)) from water within 3 days. This study provides a promising strategy for in situ arsenic phytoremediation through the cooperation of plant and endophytic bacterium.

Widespread Distribution of the arsO Gene Confers Bacterial Resistance to Environmental Antimony.

Microbial oxidation of environmental antimonite (Sb(III)) to antimonate (Sb(V)) is an antimony (Sb) detoxification mechanism. Ensifer adhaerens ST2, a bacterial isolate from a Sb-contaminated paddy soil, oxidizes Sb(III) to Sb(V) under oxic conditions by an unknown mechanism. Genomic analysis of ST2 reveals a gene of unknown function in an arsenic resistance (ars) operon that we term arsO. The transcription level of arsO was significantly upregulated by the addition of Sb(III). ArsO is predicted to be a flavoprotein monooxygenase but shows low sequence similarity to other flavoprotein monooxygenases. Expression of arsO in the arsenic-hypersensitive Escherichia coli strain AW3110Δars conferred increased resistance to Sb(III) but not arsenite (As(III)) or methylarsenite (MAs(III)). Purified ArsO catalyzes Sb(III) oxidation to Sb(V) with NADPH or NADH as the electron donor but does not oxidize As(III) or MAs(III). The purified enzyme contains flavin adenine dinucleotide (FAD) at a ratio of 0.62 mol of FAD/mol protein, and enzymatic activity was increased by addition of FAD. Bioinformatic analyses show that arsO genes are widely distributed in metagenomes from different environments and are particularly abundant in environments affected by human activities. This study demonstrates that ArsO is an environmental Sb(III) oxidase that plays a significant role in the detoxification of Sb(III).

Arsinothricin Inhibits Plasmodium falciparum Proliferation in Blood and Blocks Parasite Transmission to Mosquitoes.

Malaria, caused by Plasmodium protozoal parasites, remains a leading cause of morbidity and mortality. The Plasmodium parasite has a complex life cycle, with asexual and sexual forms in humans and Anopheles mosquitoes. Most antimalarials target only the symptomatic asexual blood stage. However, to ensure malaria eradication, new drugs with efficacy at multiple stages of the life cycle are necessary. We previously demonstrated that arsinothricin (AST), a newly discovered organoarsenical natural product, is a potent broad-spectrum antibiotic that inhibits the growth of various prokaryotic pathogens. Here, we report that AST is an effective multi-stage antimalarial. AST is a nonproteinogenic amino acid analog of glutamate that inhibits prokaryotic glutamine synthetase (GS). Phylogenetic analysis shows that Plasmodium GS, which is expressed throughout all stages of the parasite life cycle, is more closely related to prokaryotic GS than eukaryotic GS. AST potently inhibits Plasmodium GS, while it is less effective on human GS. Notably, AST effectively inhibits both Plasmodium erythrocytic proliferation and parasite transmission to mosquitoes. In contrast, AST is relatively nontoxic to a number of human cell lines, suggesting that AST is selective against malaria pathogens, with little negative effect on the human host. We propose that AST is a promising lead compound for developing a new class of multi-stage antimalarials.

Arsenite Methyltransferase Diversity and Optimization of Methylation Efficiency.

Arsenic is methylated by arsenite (As(III)) S-adenosylmethionine (SAM) methyltransferases (ArsMs). ArsM crystal structures show three domains (an N-terminal SAM binding domain (A domain), a central arsenic binding domain (B domain), and a C-terminal domain of unknown function (C domain)). In this study, we performed a comparative analysis of ArsMs and found a broad diversity in structural domains. The differences in the ArsM structure enable ArsMs to have a range of methylation efficiencies and substrate selectivities. Many small ArsMs with 240-300 amino acid residues have only A and B domains, represented by RpArsM from Rhodopseudomonas palustris. These small ArsMs have higher methylation activity than larger ArsMs with 320-400 residues such as Chlamydomonas reinhardtii CrArsM, which has A, B, and C domains. To examine the role of the C domain, the last 102 residues in CrArsM were deleted. This CrArsM truncation exhibited higher As(III) methylation activity than the wild-type enzyme, suggesting that the C-terminal domain has a role in modulating the rate of catalysis. In addition, the relationship of arsenite efflux systems and methylation was examined. Lower rates of efflux led to higher rates of methylation. Thus, the rate of methylation can be modulated in multiple ways.

Anoxygenic phototrophic arsenite oxidation by a Rhodobacter strain.

Microbially mediated arsenic redox transformations are key for arsenic speciation and mobility in rice paddies. Whereas anaerobic anoxygenic photosynthesis coupled to arsenite (As(III)) oxidation has been widely examined in arsenic-replete ecosystems, it remains unknown whether this light-dependent process exists in paddy soils. Here, we isolated a phototrophic purple bacteria, Rhodobacter strain CZR27, from an arsenic-contaminated paddy soil and demonstrated its capacity to oxidize As(III) to arsenate (As(V)) using malate as a carbon source photosynthetically. Genome sequencing revealed an As(III)-oxidizing gene cluster (aioXSRBA) encoding an As(III) oxidase. Functional analyses showed that As(III) oxidation under anoxic phototrophic conditions correlated with transcription of the large subunit of the As(III) oxidase aioA gene. Furthermore, the non-As(III) oxidizer Rhodobacter capsulatus SB1003 heterologously expressing aioBA from strain CZR27 was able to oxidize As(III), indicating that aioBA was responsible for the observed As(III) oxidation in strain CZR27. Our study provides evidence for the presence of anaerobic photosynthesis-coupled As(III) oxidation in paddy soils, highlighting the importance of light-dependent, microbe-mediated arsenic redox changes in paddy arsenic biogeochemistry.

The ArsQ permease and transport of the antibiotic arsinothricin.

The pentavalent organoarsenical arsinothricin (AST) is a natural product synthesized by the rhizosphere bacterium Burkholderia gladioli GSRB05. AST is a broad-spectrum antibiotic effective against human pathogens such as carbapenem-resistant Enterobacter cloacae. It is a non-proteogenic amino acid and glutamate mimetic that inhibits bacterial glutamine synthetase. The AST biosynthetic pathway is composed of a three-gene cluster, arsQML. ArsL catalyzes synthesis of reduced trivalent hydroxyarsinothricin (R-AST-OH), which is methylated by ArsM to the reduced trivalent form of AST (R-AST). In the culture medium of B. gladioli, both trivalent species appear as the corresponding pentavalent arsenicals, likely due to oxidation in air. ArsQ is an efflux permease that is proposed to transport AST or related species out of the cells, but the chemical nature of the actual transport substrate is unclear. In this study, B. gladioli arsQ was expressed in Escherichia coli and shown to confer resistance to AST and its derivatives. Cells of E. coli accumulate R-AST, and exponentially growing cells expressing arsQ take up less R-AST. The cells exhibit little transport of their pentavalent forms. Transport was independent of cellular energy and appears to be equilibrative. A homology model of ArsQ suggests that Ser320 is in the substrate binding site. A S320A mutant exhibits reduced R-AST-OH transport, suggesting that it plays a role in ArsQ function. The ArsQ permease is proposed to be an energy-independent uniporter responsible for downhill transport of the trivalent form of AST out of cells, which is oxidized extracellularly to the active form of the antibiotic.

Arsenic in medicine: past, present and future.

Arsenicals are one of the oldest treatments for a variety of human disorders. Although infamous for its toxicity, arsenic is paradoxically a therapeutic agent that has been used since ancient times for the treatment of multiple diseases. The use of most arsenic-based drugs was abandoned with the discovery of antibiotics in the 1940s, but a few remained in use such as those for the treatment of trypanosomiasis. In the 1970s, arsenic trioxide, the active ingredient in a traditional Chinese medicine, was shown to produce dramatic remission of acute promyelocytic leukemia similar to the effect of all-trans retinoic acid. Since then, there has been a renewed interest in the clinical use of arsenicals. Here the ancient and modern medicinal uses of inorganic and organic arsenicals are reviewed. Included are antimicrobial, antiviral, antiparasitic and anticancer applications. In the face of increasing antibiotic resistance and the emergence of deadly pathogens such as the severe acute respiratory syndrome coronavirus 2, we propose revisiting arsenicals with proven efficacy to combat emerging pathogens. Current advances in science and technology can be employed to design newer arsenical drugs with high therapeutic index. These novel arsenicals can be used in combination with existing drugs or serve as valuable alternatives in the fight against cancer and emerging pathogens. The discovery of the pentavalent arsenic-containing antibiotic arsinothricin, which is effective against multidrug-resistant pathogens, illustrates the future potential of this new class of organoarsenical antibiotics.

Selective Methylation by an ArsM S-Adenosylmethionine Methyltransferase from Burkholderia gladioli GSRB05 Enhances Antibiotic Production.

Arsenic methylation contributes to the formation and diversity of environmental organoarsenicals, an important process in the arsenic biogeochemical cycle. The arsM gene encoding an arsenite (As(III)) S-adenosylmethionine (SAM) methyltransferase is widely distributed in members of every kingdom. A number of ArsM enzymes have been shown to have different patterns of methylation. When incubated with inorganic As(III), Burkholderia gladioli GSRB05 has been shown to synthesize the organoarsenical antibiotic arsinothricin (AST) but does not produce either methylarsenate (MAs(V)) or dimethylarsenate (DMAs(V)). Here, we show that cells of B. gladioli GSRB05 synthesize DMAs(V) when cultured with either MAs(III) or MAs(V). Heterologous expression of the BgarsM gene in Escherichia coli conferred resistance to MAs(III) but not As(III). The cells methylate MAs(III) and the AST precursor, reduced trivalent hydroxyarsinothricin (R-AST-OH) but do not methylate inorganic As(III). Similar results were obtained with purified BgArsM. Compared with ArsM orthologs, BgArsM has an additional 37 amino acid residues in a linker region between domains. Deletion of the additional 37 residues restored As(III) methylation activity. Cells of E. coli co-expressing the BgarsL gene encoding the noncanonical radical SAM enzyme that catalyzes the synthesis of R-AST-OH together with the BgarsM gene produce much more of the antibiotic AST compared with E. coli cells co-expressing BgarsL together with the CrarsM gene from Chlamydomonas reinhardtii, which lacks the sequence for additional 37 residues. We propose that the presence of the insertion reduces the fitness of B. gladioli because it cannot detoxify inorganic arsenic but concomitantly confers an evolutionary advantage by increasing the ability to produce AST.

Clinical Utility of Universal Germline Genetic Testing for Patients With Breast Cancer.

Importance: National Comprehensive Cancer Network guidelines currently recommend germline testing for high-risk genes in selected patients with breast cancer. The clinical utility of recommending testing all patients with breast cancer with multigene panels is currently under consideration. Objective: To examine the implications of universal testing of patients with breast cancer with respect to clinical decision-making. Design, Setting, and Participants: Patients from a previously reported cohort were assessed as in-criteria or out-of-criteria according to the 2017 guidelines and underwent testing with a multigene germline panel between 2017 to 2018. Patients were women and men aged 18 to 90 years, with a new and/or previous diagnosis of breast cancer who had not undergone either single or multigene testing. Clinicians from 20 community and academic sites documented patient clinical information and changes to clinical recommendations made according to test findings. Association between prevalence of pathogenic or likely pathogenic germline variants and previously unreported clinical features, including scores generated by the BRCAPRO statistical model, was determined. Data were analyzed from April 2020 to May 2022. Exposure: New and/or previous diagnosis of breast cancer. Main Outcomes and Measures: Disease management recommendations that were changed as a result of genetic testing results are reported. Results: Clinicians were asked to assess changes to clinical management as a result of germline genetic testing for 952 patients. Informative clinician-reported recommendations were provided for 939 (467 in-criteria and 472 out-of-criteria) of the patients with breast cancer (936 [99.7%] female; 702 [74.8%] White; mean [SD] age at initial diagnosis, 57.6 [11.5] years). One or more changes were reported for 31 of 37 (83.8%) in-criteria patients and 23 of 34 (67.6%) out-of-criteria patients with a pathogenic or likely pathogenic variant. Recommendations were changed as a result of testing results for 14 of 22 (63.6%) out-of-criteria patients who had a variant in a breast cancer predisposition gene. Clinicians considered testing beneficial for two-thirds of patients with pathogenic or likely pathogenic variants and for one-third of patients with either negative results or variants of uncertain significance. There was no difference in variant rate between patients meeting the BRCAPRO threshold (≥10%) and those who did not (P = .86, Fisher exact test). No changes to clinical recommendations were made for most patients with negative results (345 of 349 patients [98.9%]) or variants of uncertain significance (492 of 509 patients [96.7%]). Conclusions and Relevance: In this cohort study, germline genetic testing was used by clinicians to direct treatment for most out-of-criteria patients with breast cancer with pathogenic or likely pathogenic germline variants, including those with moderate-risk variants. Universal germline testing informs clinical decision-making and provides access to targeted treatments and clinical trials for all patients with breast cancer.

A PadR family transcriptional repressor controls transcription of a trivalent metalloid resistance operon of Azospirillum halopraeferens strain Au 4.

Methylarsenite [MAs(III)] is a highly toxic arsenical produced by some microbes as an antibiotic. In this study, we demonstrate that a PadR family transcriptional regulator, PadRars , from Azospirillum halopraeferens strain Au 4 directly binds to the promoter region of the arsenic resistance (ars) operon (consisting of padRars , arsV, and arsW) and represses transcription of arsV and arsW genes involved in MAs(III) resistance. Quantitative reverse transcriptase PCR and transcriptional reporter assays showed that transcription of the ars operon is induced strongly by MAs(III) and less strongly by arsenite and antimonite. Electrophoretic mobility shift assays with recombinant PadRars showed that it represses transcription of the ars operon by binding to two inverted-repeat sequences within the ars promoter. PadRars has two conserved cysteine pairs, Cys56/57 and Cys133/134; mutation of the first pair to serine abolished the transcriptional response of the ars operon to trivalent metalloids, suggesting that Cys56/57 form a binding site for trivalent metalloids. Either C133S or C134S derivative responses to MAs(III) but not As(III) or Sb(III), suggesting that it is a third ligand to trivalent metalloids. PadRars represents a new type of repressor proteins regulating transcription of an ars operon involved in the resistance to trivalent metalloids, especially MAs(III).

Impact of germline mutations in cancer-predisposing genes on long-term survival in patients with epithelial ovarian cancer.

BACKGROUND: Several clinical and tumour factors impact on ovarian cancer survival. It is important to evaluate if germline mutations impact long-term outcomes among patients with epithelial ovarian cancer. METHODS: We followed 1422 Ontario women with ovarian cancer. Clinical information was obtained from medical records and vital status was determined by registry linkage. Germline genetic testing was performed for 12 susceptibility genes. We estimated 20-year cancer-specific survival according to various factors. RESULTS: Twenty-year survival was inferior for women with serous cancers vs. other types (22.3% vs. 68.6%; P < 0.0001). Of the 1422 patients, 248 (17.4%) carried a germline mutation; 119 BRCA1; 75 BRCA2; 7 in a mismatch repair (MMR) gene and 47 in one of seven other genes. Among serous patients, 20-year survival was 28.9% for similar for women with a BRCA1 (28.9%), BRCA2 (21.2%) or no mutation (21.6%). Among endometrioid patients, 20-year survival was poor for women with a BRCA vs. no mutation (47.3% vs. 70.4%; P = 0.004). Six of the seven MMR mutation carriers are currently alive, while all three PALB2 mutation carriers died within 3 years of diagnosis. Among women with Stage III/IV serous cancers, 20-year survival was 9.4% for those with vs. 46.5% for those with no residual disease (HR = 2.91; 95% CI 2.12-4.09, P < 0.0001). CONCLUSIONS: The most important predictor of long-term survival was no residual disease post surgery. BRCA mutation status was not predictive of long-term survival while those with MMR mutations had excellent survival. Larger studies on PALB2 carriers are needed.

The ArsI C-As lyase: Elucidating the catalytic mechanism of degradation of organoarsenicals.

Organoarsenicals such as monosodium methylarsenate (MSMA or MAs(V)) and roxarsone (4-hydroxyl-3-nitrophenylarsenate or Rox(V)) have been extensively used as herbicides and growth enhancers for poultry, respectively. Degradation of organoarsenicals to inorganic arsenite (As(III)) contaminates crops and drinking water. One such process is catalyzed by the bacterial enzyme ArsI, whose gene is found in many soil bacteria. ArsI is a non-heme ferrous iron (Fe(II))-dependent dioxygenase that catalyzes oxygen-dependent cleavage of the carbon­arsenic (C-As) bond in trivalent organoarsenicals, degrading them to inorganic As(III). From previous crystal structures of ArsI, we predicted that a loop-gating mechanism controls the catalytic reaction. Understanding the catalytic mechanism of ArsI requires knowledge of the mechanisms of substrate binding and activation of dioxygen. Here we report new ArsI structures with bound Rox(III) and mutant enzymes with alteration of active site residues. Our results elucidate steps in the catalytic cycle of this novel dioxygenase and enhance understanding of the recycling of environmental organoarsenicals.

An ArsRC fusion protein enhances arsenate sensing and detoxification.

Arsenical resistance (ars) operons encode genes for arsenic resistance and biotransformation. The majority are composed of individual genes, but fusion of ars genes is not uncommon, although it is not clear if the fused gene products are functional. Here we report identification of a four-gene ars operon from Paracoccus sp. SY that has two arsR-arsC gene fusions. ArsRC1 and ArsRC2 are related proteins that consist of an N-terminal ArsR arsenite (As(III))-responsive repressor with a C-terminal ArsC arsenate reductase. The other two genes in the operon are gapdh and arsJ. GAPDH, glyceraldehyde 3-phosphate dehydrogenase, forms 1-arseno-3-phosphoglycerate (1As3PGA) from 3-phosphoglyceraldehyde and arsenate (As(V)), ArsJ is an efflux permease for 1As3PGA that dissociates into extracellular As(V) and 3-phosphoglycerate. The net effect is As(V) extrusion and resistance. ArsRs are usually selective for As(III) and do not respond to As(V). However, the substrates and products of this operon are pentavalent, which would not be inducers of the operon. We propose that ArsRC fusions overcome this limitation by channelling the ArsC product into the ArsR binding site without diffusion through the cytosol, a de facto mechanism for As(V) induction. This novel mechanism for arsenate sensing can confer an evolutionary advantage for detoxification of inorganic arsenate.