Single-stranded DNA-binding protein hSSB1 is critical for genomic stability

Single-strand DNA (ssDNA)-binding proteins (SSBs) are ubiquitous and essential for a wide variety of DNA metabolic processes, including DNA replication, recombination, DNA damage detection and repair. SSBs have multiple roles in binding and sequestering ssDNA, detecting DNA damage, stimulating nucleases, helicases and strand-exchange proteins, activating transcription and mediating protein-protein interactions. In eukaryotes, the major SSB, replication protein A (RPA), is a heterotrimer. Here we describe a second human SSB (hSSB1), with a domain organization closer to the archaeal SSB than to RPA. Ataxia telangiectasia mutated (ATM) kinase phosphorylates hSSB1 in response to DNA double-strand breaks (DSBs). This phosphorylation event is required for DNA damage-induced stabilization of hSSB1. Upon induction of DNA damage, hSSB1 accumulates in the nucleus and forms distinct foci independent of cell-cycle phase. These foci co-localize with other known repair proteins. In contrast to RPA, hSSB1 does not localize to replication foci in S-phase cells and hSSB1 deficiency does not influence S-phase progression. Depletion of hSSB1 abrogates the cellular response to DSBs, including activation of ATM and phosphorylation of ATM targets after ionizing radiation. Cells deficient in hSSB1 exhibit increased radiosensitivity, defective checkpoint activation and enhanced genomic instability coupled with a diminished capacity for DNA repair. These findings establish that hSSB1 influences diverse endpoints in the cellular DNA damage response.

Increase in rice grain arsenic for regions of Bangladesh irrigating paddies with elevated arsenic in groundwaters.

Concern has been raised by Bangladeshi and international scientists about elevated levels of arsenic in Bengali food, particularly in rice grain. This is the first inclusive food market-basket survey from Bangladesh, which addresses the speciation and concentration of arsenic in rice, vegetables, pulses, and spices. Three hundred thirty aman and boro rice, 94 vegetables, and 50 pulse and spice samples were analyzed for total arsenic, using inductivity coupled plasma mass spectrometry (ICP-MS). The districts with the highest mean arsenic rice grain levels were all from southwestern Bangladesh:? Faridpur (boro) 0.51 > Satkhira (boro) 0.38 > Satkhira (aman) 0.36 > Chuadanga (boro) 0.32 > Meherpur (boro) 0.29 µg As g-1. The vast majority of food ingested arsenic in Bangladesh diets was found to be inorganic; with the predominant species detected in Bangladesh rice being arsenite (AsIII) or arsenate (AsV) with dimethyl arsinic acid (DMAV) being a minor component. Vegetables, pulses, and spices are less important to total arsenic intake than water and rice. Predicted inorganic arsenic intake from rice is modeled with the equivalent intake from drinking water for a typical Bangladesh diet. Daily consumption of rice with a total arsenic level of 0.08 µg As g-1 would be equivalent to a drinking water arsenic level of 10 µg L-1. Concern has been raised by Bangladeshi and international scientists about elevated levels of arsenic in Bengali food, particularly in rice grain. This is the first inclusive food market-basket survey from Bangladesh, which addresses the speciation and concentration of arsenic in rice, vegetables, pulses, and spices. Three hundred thirty aman and boro rice, 94 vegetables, and 50 pulse and spice samples were analyzed for total arsenic, using inductivity coupled plasma mass spectrometry (ICP-MS). The districts with the highest mean arsenic rice grain levels were all from southwestern Bangladesh:? Faridpur (boro) 0.51 > Satkhira (boro) 0.38 > Satkhira (aman) 0.36 > Chuadanga (boro) 0.32 > Meherpur (boro) 0.29 µg As g-1. The vast majority of food ingested arsenic in Bangladesh diets was found to be inorganic; with the predominant species detected in Bangladesh rice being arsenite (AsIII) or arsenate (AsV) with dimethyl arsinic acid (DMAV) being a minor component. Vegetables, pulses, and spices are less important to total arsenic intake than water and rice. Predicted inorganic arsenic intake from rice is modeled with the equivalent intake from drinking water for a typical Bangladesh diet. Daily consumption of rice with a total arsenic level of 0.08 µg As g-1 would be equivalent to a drinking water arsenic level of 10 µg L-1.

Genomic Analysis of Pseudomonas putida Phage tf with Localized Single-Strand DNA Interruptions

The complete sequence of the 46,267 bp genome of the lytic bacteriophage tf specific to Pseudomonas putida PpG1 has been determined. The phage genome has two sets of convergently transcribed genes and 186 bp long direct terminal repeats. The overall genomic architecture of the tf phage is similar to that of the previously described Pseudomonas aeruginosa phages PaP3, LUZ24 and phiMR299-2, and 39 out of the 72 products of predicted tf open reading frames have orthologs in these phages. Accordingly, tf was classified as belonging to the LUZ24-like bacteriophage group. However, taking into account very low homology levels between tf DNA and that of the other phages, tf should be considered as an evolutionary divergent member of the group. Two distinguishing features not reported for other members of the group were found in the tf genome. Firstly, a unique end structure - a blunt right end and a 4-nucleotide 3'-protruding left end - was observed. Secondly, 14 single-chain interruptions (nicks) were found in the top strand of the tf DNA. All nicks were mapped within a consensus sequence 5'-TACT/RTGMC-3'. Two nicks were analyzed in detail and were shown to be present in more than 90% of the phage population. Although localized nicks were previously found only in the DNA of T5-like and phiKMV-like phages, it seems increasingly likely that this enigmatic structural feature is common to various other bacteriophages.

Arguing with Regions

An analytical survey of how regions have entered into the arguments of the
social sciences serves to highlight the uses and limitations of different understandings of regions and their various theoretical biases. It also provides a way of introducing the articles in the rest of this special issue. It considers how regions have come to be used as a classificatory device across the social sciences, discusses the various meanings given to regions in empirical research, and examines the main philosophical and theoretical controversies that have been sparked by their use. Matching regions to purpose and avoiding a singular conception of ‘region’ that claims to fit all arguments are the main conclusions.

TMAX – a Database System Integrating Tissue Biomarker & Genomic Data

Background: Popular approaches in human tissue-based biomarker discovery include tissue microarrays (TMAs) and DNA Microarrays (DMAs) for protein and gene expression profiling respectively. The data generated by these analytic platforms, together with associated image, clinical and pathological data currently reside on widely different information platforms, making searching and cross-platform analysis difficult. Consequently, there is a strong need to develop a single coherent database capable of correlating all available data types.

Method: This study presents TMAX, a database system to facilitate biomarker discovery tasks. TMAX organises a variety of biomarker discovery-related data into the database. Both TMA and DMA experimental data are integrated in TMAX and connected through common DNA/protein biomarkers. Patient clinical data (including tissue pathological data), computer assisted tissue image and associated analytic data are also included in TMAX to enable the truly high throughput processing of ultra-large digital slides for both TMAs and whole slide tissue digital slides. A comprehensive web front-end was built with embedded XML parser software and predefined SQL queries to enable rapid data exchange in the form of standard XML files.

Results & Conclusion: TMAX represents one of the first attempts to integrate TMA data with public gene expression experiment data. Experiments suggest that TMAX is robust in managing large quantities of data from different sources (clinical, TMA, DMA and image analysis). Its web front-end is user friendly, easy to use, and most importantly allows the rapid and easy data exchange of biomarker discovery related data. In conclusion, TMAX is a robust biomarker discovery data repository and research tool, which opens up the opportunities for biomarker discovery and further integromics research.

Weathering in Arid Regions

Many misconceptions exist regarding weathering in arid regions. Chief among these are assumptions that physical processes dominate and are not very effective because of a perceived lack of moisture. This chapter explores the factors that combine to make weathering in arid regions spatially and temporally complex, reflecting the range of surface microenvironmental conditions. Because of desert landscape complexity, attempts at interpreting weathered features must take into account the long-term history of rock outcrops and debris that mantle them, as most desert landscapes contain legacies of weathering forms and products, which were developed when moisture was more readily available in the past.

Cooperativity of imprinted genes inactivated by acquired chromosome 20q deletions

Large regions of recurrent genomic loss are common in cancers; however, with a few well-characterized exceptions, how they contribute to tumor pathogenesis remains largely obscure. Here we identified primate-restricted imprinting of a gene cluster on chromosome 20 in the region commonly deleted in chronic myeloid malignancies. We showed that a single heterozygous 20q deletion consistently resulted in the complete loss of expression of the imprinted genes L3MBTL1 and SGK2, indicative of a pathogenetic role for loss of the active paternally inherited locus. Concomitant loss of both L3MBTL1 and SGK2 dysregulated erythropoiesis and megakaryopoiesis, 2 lineages commonly affected in chronic myeloid malignancies, with distinct consequences in each lineage. We demonstrated that L3MBTL1 and SGK2 collaborated in the transcriptional regulation of MYC by influencing different aspects of chromatin structure. L3MBTL1 is known to regulate nucleosomal compaction, and we here showed that SGK2 inactivated BRG1, a key ATP-dependent helicase within the SWI/SNF complex that regulates nucleosomal positioning. These results demonstrate a link between an imprinted gene cluster and malignancy, reveal a new pathogenetic mechanism associated with acquired regions of genomic loss, and underline the complex molecular and cellular consequences of "simple" cancer-associated chromosome deletions.

Dual-colour HER2/chromosome 17 chromogenic in situ hybridisation enables accurate assessment of HER2 genomic status in ovarian tumours

Ovarian cancer is a leading cause of gynaecological cancer-related morbidity and mortality. There has been increasing interest in the potential utility of anti-human epidermal growth factor receptor 2 (anti-HER2) agents in the treatment of this disease, with the attendant need to identify suitable predictive biomarkers of response to treatment.

Dual-colour HER2/chromosome 17 chromogenic in situ hybridisation assay enables accurate assessment of HER2 genomic status in gastric cancer and has potential utility in HER2 testing of biopsy samples

Determination of HER2 protein expression by immunohistochemistry (IHC) and genomic status by fluorescent in situ hybridisation (FISH) are important in identifying a subset of high HER2-expressing gastric cancers that might respond to trastuzumab. Although FISH is considered the standard for determination of HER2 genomic status, brightfield ISH is being increasingly recognised as a viable alternative. Also, the impact of HER2 protein expression/genomic heterogeneity on the accuracy of HER2 testing has not been well studied in the context of gastric biopsy samples.

Combined genomic and phenotype screening reveals secretory factor SPINK1 as an invasion and survival factor associated with patient prognosis in breast cancer

Secretory factors that drive cancer progression are attractive immunotherapeutic targets. We used a whole-genome data-mining approach on multiple cohorts of breast tumours annotated for clinical outcomes to discover such factors. We identified Serine protease inhibitor Kazal-type 1 (SPINK1) to be associated with poor survival in estrogen receptor-positive (ER+) cases. Immunohistochemistry showed that SPINK1 was absent in normal breast, present in early and advanced tumours, and its expression correlated with poor survival in ER+ tumours. In ER- cases, the prognostic effect did not reach statistical significance. Forced expression and/or exposure to recombinant SPINK1 induced invasiveness without affecting cell proliferation. However, down-regulation of SPINK1 resulted in cell death. Further, SPINK1 overexpressing cells were resistant to drug-induced apoptosis due to reduced caspase-3 levels and high expression of Bcl2 and phospho-Bcl2 proteins. Intriguingly, these anti-apoptotic effects of SPINK1 were abrogated by mutations of its protease inhibition domain. Thus, SPINK1 affects multiple aggressive properties in breast cancer: survival, invasiveness and chemoresistance. Because SPINK1 effects are abrogated by neutralizing antibodies, we suggest that SPINK1 is a viable potential therapeutic target in breast cancer.

Identification of MET genomic amplification, protein expression and alternative splice isoforms in neuroblastomas

Crizotinib, a dual anaplastic lymphoma kinase (ALK) and mesenchymal-epithelial transition (MET) tyrosine kinase inhibitor, is currently being evaluated for the treatment of neuroblastoma. Its effects are thought to be mediated mainly via its activity against ALK. Although MET genomic/protein expression status might conceivably affect crizotinib efficacy, this issue has hitherto not received attention in neuroblastomas.

Genetic and genomic analyses of musculoskeletal differences between BEH and BEL strains

Berlin high (BEH) and Berlin low (BEL) strains selected for divergent growth differ 3-fold in body weight. We aimed at examining muscle mass, which is a major contributor to body weight, by exploring anatomical characteristics of the soleus muscle, its fiber numbers and their cross sectional area (CSA), by analysing transcriptome of the gastrocnemius and by initiating quantitative trait locus (QTL) mapping. BEH muscles were 4-to-8 times larger compared to BEL strain. In sub-strain BEH+/+, mutant myostatin was replaced with a wild type allele, however, BEH+/+muscles still were 2-to-4 times larger compared to the BEL strain. BEH soleus contained 2-times more (P<0.0001) and 2-times larger in CSA (P<0.0001) fibers compared to BEL strain. In addition, soleus femoral attachment anomaly (SFAA) was observed in all BEL mice. One significant (chromosome 1) and four suggestive (chromosomes 3, 4, 6 and 9) muscle weight QTLs were mapped in 21-day old F2 intercross (n=296) between BEH and BEL strains. The frequency of SFAA incidence in the F2 and in the backcross to BEL strain (BCL) suggested the presence of more than one causative gene. Two suggestive SFAA QTLs were mapped in BCL, however, their peak markers were not associated with the phenotype in F2. RNA-Seq analysis revealed 2,148 differentially expressed (P<0.1) genes and 45,673 SNPs and >2,000 indels between BEH+/+ and BEL males. In conclusion, contrasting muscle traits, genomic and gene expression differences between BEH and BEL strains provide a promising model for the search of genes involved in muscle growth and musculoskeletal morphogenesis.

Neutral Genomic Microevolution of a Recently Emerged Pathogen, Salmonella enterica Serovar Agona

Salmonella enterica serovar Agona has caused multiple food-borne outbreaks of gastroenteritis since it was first isolated in 1952. We analyzed the genomes of 73 isolates from global sources, comparing five distinct outbreaks with sporadic infections as well as food contamination and the environment. Agona consists of three lineages with minimal mutational diversity: only 846 single nucleotide polymorphisms (SNPs) have accumulated in the non-repetitive, core genome since Agona evolved in 1932 and subsequently underwent a major population expansion in the 1960s. Homologous recombination with other serovars of S. enterica imported 42 recombinational tracts (360 kb) in 5/143 nodes within the genealogy, which resulted in 3,164 additional SNPs. In contrast to this paucity of genetic diversity, Agona is highly diverse according to pulsed-field gel electrophoresis (PFGE), which is used to assign isolates to outbreaks. PFGE diversity reflects a highly dynamic accessory genome associated with the gain or loss (indels) of 51 bacteriophages, 10 plasmids, and 6 integrative conjugational elements (ICE/IMEs), but did not correlate uniquely with outbreaks. Unlike the core genome, indels occurred repeatedly in independent nodes (homoplasies), resulting in inaccurate PFGE genealogies. The accessory genome contained only few cargo genes relevant to infection, other than antibiotic resistance. Thus, most of the genetic diversity within this recently emerged pathogen reflects changes in the accessory genome, or is due to recombination, but these changes seemed to reflect neutral processes rather than Darwinian selection. Each outbreak was caused by an independent clade, without universal, outbreak-associated genomic features, and none of the variable genes in the pan-genome seemed to be associated with an ability to cause outbreaks. © 2013 Achtman et al