Escherichia coli HU protein has a role in the repair of abasic sites in DNA

HU is one of the most abundant DNA binding proteins in Escherichia coli. We find that it binds strongly to DNA containing an abasic (AP) site or tetrahydrofuran (THF) (apparent Kd ≈50 nM). It also possesses an AP lyase activity that cleaves at a deoxyribose but not at a THF residue. The binding and cleavage of an AP site was observed only with the HUαβ heterodimer. Site-specific mutations at K3 and R61 residues led to a change in substrate binding and cleavage. Both K3A(α)K3A(β) and R61A(α)R61A(β) mutant HU showed significant reduction in binding to DNA containing AP site; however, only R61A(α)R61A(β) mutant protein exhibited significant loss in AP lyase activity. Both K3A(α)K3A(β) and R61K(α)R61K(β) showed slight reduction in AP lyase activities. The function of HU protein as an AP lyase was confirmed by the ability of hupA or hupB mutations to further reduce the viability of an E. coli dut(Ts) xth mutant, which generates lethal AP sites at 37°C; the hupA and hupB derivatives, respectively, had a 6-fold and a 150-fold lower survival at 37°C than did the parental strain. These data suggest, therefore, that HU protein plays a significant role in the repair of AP sites in E. coli

They Are What You Eat: Can Nutritional Factors during Gestation and Early Infancy Modulate the Neonatal Immune Response?

The ontogeny of the human immune system is sensitive to nutrition even in the very early embryo, with both deficiency and excess of macro- and micronutrients being potentially detrimental. Neonates are particularly vulnerable to infectious disease due to the immaturity of the immune system and modulation of nutritional immunity may play a role in this sensitivity. This review examines whether nutrition around the time of conception, throughout pregnancy, and in early neonatal life may impact on the developing infant immune system

Nutritionally Enhanced Staple Food Crops

Crop biofortification is a sustainable and cost-effective strategy to address malnutrition in developing countries. This review synthesizes the progress toward developing seed micronutrient-dense cereals and legumes cultivars by exploiting natural genetic variation using conventional breeding and/or transgenic technology, and discusses the associated issues to strengthen crop biofortification research and development. Some major QTL for seed iron and zinc, seed phosphorus, and seed phytate in common bean, rice,J;md wheat have been mapped. An iron reductase QTL associated with seed-iron ~QTL is found in common bean where the genes coding for candidate enzymes involved in phytic acid synthesis have also been mapped. Candidate genes for Ipa co segregate with mutant phenotypes identified in rice and soybean. The Gpe-B1 locus in wild emmer wheat accelerates senescence and increases nutrient remobilization from leaves to developing seeds, and another gene named TtNAM-B1 affecting these traits has been cloned. Seed iron-dense common bean and rice in Latin America; seed iron-dense common bean in eastern and southern Africa;....

Glutathione Redox Dynamics and Expression of Glutathione-related Genes in the Developing Embryo

Embryonic development involves dramatic changes in cell proliferation and differentiation that must be highly coordinated and tightly regulated. Cellular redox balance is critical for cell fate decisions, but it is susceptible to disruption by endogenous and exogenous sources of oxidative stress. The most abundant endogenous non-protein antioxidant defense molecule is the tri-peptide glutathione (γ-glutamyl-cysteinylglycine, GSH), but the ontogeny of GSH concentration and redox state during early life stages is poorly understood. Here, we describe the GSH redox dynamics during embryonic and early larval development (0–5 days post-fertilization) in the zebrafish (Danio rerio), a model vertebrate embryo. We measured reduced and oxidized glutathione (GSH, GSSG) using HPLC, and calculated the whole embryo total glutathione (GSHT) concentrations and redox potentials (Eh) over 0–120 hours of zebrafish development (including mature oocytes, fertilization, mid-blastula transition, gastrulation, somitogenesis, pharyngula, prehatch embryos, and hatched eleutheroembryos). GSHT concentration doubled between 12 hours post fertilization (hpf) and hatching. The GSH Eh increased, becoming more oxidizing during the first 12 h, and then oscillated around −190 mV through organogenesis, followed by a rapid change, associated with hatching, to a more negative (more reducing) Eh (−220 mV). After hatching, Eh stabilized and remained steady through 120 hpf. The dynamic changes in GSH redox status and concentration defined discrete windows of development: primary organogenesis, organ differentiation, and larval growth. We identified the set of zebrafish genes involved in the synthesis, utilization, and recycling of GSH, including several novel paralogs, and measured how expression of these genes changes during development. Ontogenic changes in the expression of GSH-related genes support the hypothesis that GSH redox state is tightly regulated early in development. This study provides a foundation for understanding the redox regulation of developmental signaling and investigating the effects of oxidative stress during embryogenesis

Inhaled bacteriophage-loaded polymeric microparticles ameliorate acute lung infections

Lung infections associated with pneumonia, or cystic fibrosis caused by Pseudomonas aeruginosa or other bacteria, result in significant morbidity and mortality, in part owing to the development of multidrug resistance, also against last-resort antibiotics. Lytic bacteriophages (that is, viruses that specifically kill bacteria) can reduce lung-associated infections, yet their clinical use is hindered by difficulties in delivering active phages to the deep lung. Here, we show that phage-loaded polymeric microparticles deposit throughout the lung via dry powder inhalation and that they deliver active phages. Phage-loaded microparticles effectively reduced P. aeruginosa infections and the associated inflammation in wild-type and cystic fibrosis trans-membrane-conductance-regulator knockout mice, and rescued the mice from pneumonia-associated death. These polymeric microparticles might constitute a clinically translatable therapy for eradicating hospital-acquired lung infections and infections associated with cystic fibrosis

Identification and characterization of a human DNA glycosylase for repair of modified bases in oxidatively damaged DNA

8-oxoguanine (8-oxoG), ring-opened purines (formamidopyrimidines or Fapys), and other oxidized DNA base lesions generated by reactive oxygen species are often mutagenic and toxic, and have been implicated in the etiology of many diseases, including cancer, and in aging. Repair of these lesions in all organisms occurs primarily via the DNA base excision repair pathway, initiated with their excision by DNA glycosylase/AP lyases, which are of two classes. One class utilizes an internal Lys residue as the active site nucleophile, and includes Escherichia coli Nth and both known mammalian DNA glycosylase/AP lyases, namely, OGG1 and NTH1. E. coli MutM and its paralog Nei, which comprise the second class, use N-terminal Pro as the active site. Here, we report the presence of two human orthologs of E. coli mutM nei genes in the human genome database, and characterize one of their products. Based on the substrate preference, we have named it NEH1 (Nei homolog). The 44-kDa, wild-type recombinant NEH1, purified to homogeneity from E. coli, excises Fapys from damaged DNA, and oxidized pyrimidines and 8-oxoG from oligodeoxynucleotides. Inactivation of the enzyme because of either deletion of N-terminal Pro or Histag fusion at the N terminus supports the role of N-terminal Pro as its active site. The tissue-specific levels of NEH1 and OGG1 mRNAs are distinct, and S phase-specific increase in NEH1 at both RNA and protein levels suggests that NEH1 is involved in replication-associated repair of oxidized bases

doi:10.1093/nar/gkm567 Escherichia coli HU protein has a role in the repair of abasic sites in DNA

HU is one of the most abundant DNA binding proteins in Escherichia coli. We find that it binds strongly to DNA containing an abasic (AP) site or tetrahydrofuran (THF) (apparent Kd 50 nM). It also possesses an AP lyase activity that cleaves at a deoxyribose but not at a THF residue. The binding and cleavage of an AP site was observed only with the HUab heterodimer. Site-specific mutations at K3 and R61 residues led to a change in substrate binding and cleavage. Both K3A(a)K3A(b) and R61A(a)R61A(b) mutant HU showed significant reduction in binding to DNA containing AP site; however, only R61A(a)R61A(b) mutant protein exhibited significant loss in AP lyase activity. Both K3A(a)K3A(b) and R61K(a)R61K(b) showed slight reduction in AP lyase activities. The function of HU protein as an AP lyase was confirmed by the ability of hupA or hupB mutations to further reduce the viability of an E. coli dut(Ts) xth mutant, which generates lethal AP sites at 378C; the hupA and hupB derivatives, respectively, had a 6-fold and a 150-fold lower survival at 378C than did the parental strain. These data suggest, therefore, that HU protein plays a significant role in the repair of AP sites in E. coli