Regulation of DNA Pairing in Homologous Recombination

Homologous recombination (HR) is a major mechanism for eliminating DNA double-strand breaks from chromosomes. In this process, the break termini are resected nucleolytically to form 3′ ssDNA (single-strand DNA) overhangs. A recombinase (i.e., a protein that catalyzes homologous DNA pairing and strand exchange) assembles onto the ssDNA and promotes pairing with a homologous duplex. DNA synthesis then initiates from the 3′ end of the invading strand, and the extended DNA joint is resolved via one of several pathways to restore the integrity of the injured chromosome. It is crucial that HR be carefully orchestrated because spurious events can create cytotoxic intermediates or cause genomic rearrangements and loss of gene heterozygosity, which can lead to cell death or contribute to the development of cancer. In this review, we will discuss how DNA motor proteins regulate HR via a dynamic balance of the recombination-promoting and -attenuating activities that they possess.     

Salicortin suppresses lipopolysaccharide-stimulated inflammatory responses via blockade of NF-κB and JNK activation in RAW 264.7 macrophages

We isolated the phenolic glucoside salicortin from a Populus euramericana bark extract, and examined its ability to suppress inflammatory responses as well as the molecular mechanisms underlying these abilities, using lipopolysaccharide (LPS)-stimulated RAW264.7 cells. Salicortin inhibited iNOS expression and the subsequent production of NO in a dose-dependent manner in the LPS-stimulated RAW 264.7 cells. Salicortin significantly suppressed LPS-induced signal cascades of NF-κB activation, such as IKK activation, IκBα phosphorylation and p65 phosphorylation in RAW 264.7 cells. In addition, salicortin inhibited the LPS-induced activation of JNK, but not ERK or p38 MAPK. Furthermore, salicortin significantly inhibited production of pro-inflammatory cytokines, such as TNF-α, IL-1β and IL-6 in the LPS-stimulated RAW 264.7 cells. These findings suggest that salicortin may show its anti-inflammatory activity by suppressing the LPS-induced expression of pro-inflammatory mediators through inhibition of NF-κB and JNK MAPK signaling cascades in macrophages. [BMB Reports 2014; 47(6): 318-323]     

Chemical constituents isolated from Polygala japonica leaves and their inhibitory effect on nitric oxide production in vitro

A methanolic extract of dried leaves of Polygala japonica Houtt (Polygalaceae) significantly attenuated nitric oxide production in lipopolysaccharide-simulated BV2 microglia. Five anthraquinones chrysophanol (1), emodin (2), aloe-emodin (3), emodin 8-O-β-D-glucopyranoside (4) and trihydroxy anthraquinone (5), and four flavonoids kaempferol (6), chrysoeriol (7), kaempferol 3-gentiobioside (8) and isorhamnetin (9) were isolated from the methanolic extract using bioactivity-guided fractionation. Among them, compounds 1–4, 6 and 7 showed significant inhibitory effect on lipopolysaccharide-induced nitric oxide production in BV2 microglia at the concentrations ranging from 1.0 to 100.0 μM.     

Characterization of highly pathogenic H5N1 avian influenza A viruses isolated from South Korea

An unprecedented outbreak of H5N1 highly pathogenic avian influenza (HPAI) has been reported for poultry in eight different Asian countries, including South Korea, since December 2003. A phylogenetic analysis of the eight viral genes showed that the H5N1 poultry isolates from South Korea were of avian origin and contained the hemagglutinin and neuraminidase genes of the A/goose/Guangdong/1/96 (Gs/Gd) lineage. The current H5N1 strains in Asia, including the Korean isolates, share a gene constellation similar to that of the Penfold Park, Hong Kong, isolates from late 2002 and contain some molecular markers that seem to have been fixed in the Gs/Gd lineage virus since 2001. However, despite genetic similarities among recent H5N1 isolates, the topology of the phylogenetic tree clearly differentiates the Korean isolates from the Vietnamese and Thai isolates which have been reported to infect humans. A representative Korean isolate was inoculated into mice, with no mortality and no virus being isolated from the brain, although high titers of virus were observed in the lungs. The same isolate, however, caused systemic infections in chickens and quail and killed all of the birds within 2 and 4 days of intranasal inoculation, respectively. This isolate also replicated in multiple organs and tissues of ducks and caused some mortality. However, lower virus titers were observed in all corresponding tissues of ducks than in chicken and quail tissues, and the histological lesions were restricted to the respiratory tract. This study characterizes the molecular and biological properties of the H5N1 HPAI viruses from South Korea and emphasizes the need for comparative analyses of the H5N1 isolates from different countries to help elucidate the risk of a human pandemic from the strains of H5N1 HPAI currently circulating in Asia.     

Enhanced biological effects of Phe140Asn, a novel human granulocyte colony-stimulating factor mutant, on HL60 cells

Granulocyte colony-stimulating factor (G-CSF) is a cytokine secreted by stromal cells and plays a role in the differentiation of bone marrow stem cells and proliferation of neutrophils. Therefore, G-CSF is widely used to reduce the risk of serious infection in immunocompromised patients; however, its use in such patients is limited because of its non-persistent biological activity. We created an N-linked glycosylated form of this cytokine, hG-CSF (Phe140Asn), to assess its biological activity in the promyelocyte cell line HL60. Enhanced biological effects were identified by analyzing the JAK2/STAT3/survivin pathway in HL60 cells. In addition, mutant hG-CSF (Phe140Asn) was observed to have enhanced chemoattractant effects and improved differentiation efficiency in HL60 cells. These results suggest that the addition of N-linked glycosylation was successful in improving the biological activity of hG-CSF. Furthermore, the mutated product appears to be a feasible therapy for patients with neutropenia.     

Effect of phosphorus deficiency on aluminium-induced citrate exudation in soybean (Glycine max)

Aluminium (Al) toxicity or phosphorus (P) deficiency can induce exudation of organic acids from the roots of some plants, which is believed to be a tolerance mechanism against Al toxicity or P deficiency. In the present study, the effect of P deficiency on Al-induced citrate exudation was investigated in three soybean varieties differing in low-P tolerance. P starvation alone failed to induce secretion of organic acids from all three soybean varieties. However, P deficiency altered Al-induced citrate exudation over time, showing a complex interaction. Short × term P starvation (4 days) produced up to 50% increase in Al-induced citrate secretion, while longer-term (10 days) starvation reduced Al-induced citrate secretion to trace amounts. However, after a further 1 day in complete nutrient solution for recovery, Al-induced citrate exudation from the recovered roots was approximately 6 times higher than that from the continuously P-starved plants, but still approximately 3.6 times lower than that from the P-sufficient control. With increasing P or Al supply, Al-induced citrate exudation increased, while Al accumulation in soybean roots decreased in parallel with the decrease of P supply. The photosynthetic rate, stomatal conductance and transpiration were decreased by P deficiency, whereas the intracellular CO₂ concentration was increased. These findings indicate that P nutrition has a significant effect on Al-induced citrate exudation and Al accumulation in soybean root apices.     

Comparative proteomic analysis of virulent Korean Mycobacterium tuberculosis K-strain with other mycobacteria strain following infection of U-937 macrophage

In Korea, the Mycobacterium tuberculosis K-strain is the most prevalent clinical isolates and belongs to the Beijing family. In this study, we conducted comparative porteomics of expressed proteins of clinical isolates of the K-strain with H37Rv, H37Ra as well as the vaccine strain of Mycobacterium bovis BCG following phagocytosis by the human monocytic cell line U-937. Proteins were analyzed by 2-D PAGE and MALDITOF-MS. Two proteins, Mb1363 (probable glycogen phosphorylase GlgP) and MT2656 (Haloalkane dehalogenase LinB) were most abundant after phagocytosis of M. tuberculosis K-strain. This approach provides a method to determine specific proteins that may have critical roles in tuberculosis pathogenesis.     

In-vitro pulsatile flow visualization studies in a pulmonary artery model

In-vitro pulsatile flow visualization studies were conducted in an adult-sized pulmonary artery model to observe the effects of valvular pulmonic stenosis on the flow fields of the main, left and right pulmonary arteries. The flow patterns revealed that as the degree of stenosis increased, the jet-type flow created by the valve became narrower, and it impinged on the far (distal) wall of the left pulmonary artery further downstream from the junction of the bifurcation. This in turn led to larger regions of disturbed turbulent flow, as well as helical-type secondary flow motions in the left pulmonary artery, compared to the right pulmonary artery. The flow field in the main pulmonary artery also became more disturbed and turbulent, especially during peak systole and the deceleration phase. The flow visualization observations have been valuable in helping to conduct further quantitative studies such as pressure and velocity field mapping. Such studies are important to understanding the fluid mechanics characteristics of the main pulmonary artery and its two major branches.