Single-mode termination of phage transcriptions, disclosing bacterial adaptation for facilitated reinitiations.

Bacterial and bacteriophage RNA polymerases (RNAPs) have divergently evolved and share the RNA hairpin-dependent intrinsic termination of transcription. Here, we examined phage T7, T3 and SP6 RNAP terminations utilizing the single-molecule fluorescence assays we had developed for bacterial terminations. We discovered the phage termination mode or outcome is virtually single with decomposing termination. Therein, RNAP is displaced forward along DNA and departs both RNA and DNA for one-step decomposition, three-dimensional diffusion and reinitiation at any promoter. This phage displacement-mediated decomposing termination is much slower than readthrough and appears homologous with the bacterial one. However, the phage sole mode of termination contrasts with the bacterial dual mode, where both decomposing and recycling terminations occur compatibly at any single hairpin- or Rho-dependent terminator. In the bacterial recycling termination, RNA is sheared from RNA·DNA hybrid, and RNAP remains bound to DNA for one-dimensional diffusion, which enables facilitated recycling for reinitiation at the nearest promoter located downstream or upstream in the sense or antisense orientation. Aligning with proximity of most terminators to adjacent promoters in bacterial genomes, the shearing-mediated recycling termination could be bacterial adaptation for the facilitated reinitiations repeated at a promoter for accelerated expression and coupled at adjoining promoters for coordinated regulation.

PLASMA DYNAMICS OF NEUTROPHIL EXTRACELLULAR TRAPS AND CELL-FREE DNA IN SEPTIC AND NONSEPTIC VASOPLEGIC SHOCK: A PROSPECTIVE COMPARATIVE OBSERVATIONAL COHORT STUDY.

ABSTRACT: Background: The association between neutrophil extracellular traps (NETs) and the requirement for vasopressor and inotropic support in vasoplegic shock is unclear. This study aimed to investigate the dynamics of plasma levels of NETs and cell-free DNA (cfDNA) up to 48 h after the admission to the intensive care unit (ICU) for management of vasoplegic shock of infectious (SEPSIS) or noninfectious (following cardiac surgery, CARDIAC) origin. Methods: This is a prospective, observational study of NETs and cfDNA plasma levels at 0H (admission) and then at 12H, 24H, and 48H in SEPSIS and CARDIAC patients. The vasopressor inotropic score (VIS), the Sequential Organ Failure Assessment (SOFA) score, and time spent with invasive ventilation, in ICU and in hospital, were recorded. Associations between NETs/cfDNA and VIS and SOFA were analyzed by Spearman's correlation (rho), and between NETs/cfDNA and ventilation/ICU/hospitalization times by generalized linear regression. Results: Both NETs and cfDNA remained elevated over 48 h in SEPSIS (n = 46) and CARDIAC (n = 30) patients, with time-weighted average concentrations greatest in SEPSIS (NETs median difference 0.06 [0.02-0.11], P = 0.005; cfDNA median difference 0.48 [0.20-1.02], P < 0.001). The VIS correlated to NETs (rho = 0.3-0.60 in SEPSIS, P < 0.01, rho = 0.36-0.57 in CARDIAC, P ≤ 0.01) and cfDNA (rho = 0.40-0.56 in SEPSIS, P < 0.01, rho = 0.38-0.47 in CARDIAC, P < 0.05). NETs correlated with SOFA. Neither NETs nor cfDNA were independently associated with ventilator/ICU/hospitalization times. Conclusion: Plasma levels of NETs and cfDNA correlated with the dose of vasopressors and inotropes administered over 48 h in patients with vasoplegic shock from sepsis or following cardiac surgery. NETs levels also correlated with organ dysfunction. These findings suggest that similar mechanisms involving release of NETs are involved in the pathophysiology of vasoplegic shock irrespective of an infectious or noninfectious etiology.

Compatibility of termination mechanisms in bacterial transcription with inference on eukaryotic models.

Transcription termination has evolved to proceed through diverse mechanisms. For several classes of terminators, multiple models have been debatably proposed. Recent single-molecule studies on bacterial terminators have resolved several long-standing controversies. First, termination mode or outcome is twofold rather than single. RNA is released alone before DNA or together with DNA from RNA polymerase (RNAP), i.e. with RNA release for termination, RNAP retains on or dissociates off DNA, respectively. The concomitant release, described in textbooks, results in one-step decomposition of transcription complexes, and this 'decomposing termination' prevails at ρ factor-dependent terminators. Contrastingly, the sequential release was recently discovered abundantly from RNA hairpin-dependent intrinsic terminations. RNA-only release allows RNAP to diffuse on DNA in both directions and recycle for reinitiation. This 'recycling termination' enables one-dimensional reinitiation, which would be more expeditious than three-dimensional reinitiation by RNAP dissociated at decomposing termination. Second, while both recycling and decomposing terminations occur at a hairpin-dependent terminator, four termination mechanisms compatibly operate at a ρ-dependent terminator with ρ in alternative modes and even intrinsically without ρ. RNA-bound catch-up ρ mediates recycling termination first and decomposing termination later, while RNAP-prebound stand-by ρ invokes only decomposing termination slowly. Without ρ, decomposing termination occurs slightly and sluggishly. These four mechanisms operate on distinct timescales, providing orderly fail-safes. The stand-by mechanism is benefited by terminational pause prolongation and modulated by accompanying riboswitches more greatly than the catch-up mechanisms. Conclusively, any mechanism alone is insufficient to perfect termination, and multiple mechanisms operate compatibly to achieve maximum possible efficiency under separate controls.

Translocating RNA polymerase generates R-loops at DNA double-strand breaks without any additional factors.

The R-loops forming around DNA double-strand breaks (DSBs) within actively transcribed genes play a critical role in the DSB repair process. However, the mechanisms underlying R-loop formation at DSBs remain poorly understood, with diverse proposed models involving protein factors associated with RNA polymerase (RNAP) loading, pausing/backtracking or preexisting transcript RNA invasion. In this single-molecule study using Escherichia coli RNAP, we discovered that transcribing RNAP alone acts as a highly effective DSB sensor, responsible for generation of R-loops upon encountering downstream DSBs, without requiring any additional factors. The R-loop formation efficiency is greatly influenced by DNA end structures, ranging here from 2.8% to 73%, and notably higher on sticky ends with 3' or 5' single-stranded overhangs compared to blunt ends without any overhangs. The R-loops extend unidirectionally upstream from the DSB sites and can reach the transcription start site, interfering with ongoing-round transcription. Furthermore, the extended R-loops can persist and maintain their structures, effectively preventing the efficient initiation of subsequent transcription rounds. Our results are consistent with the bubble extension model rather than the 5'-end invasion model or the middle insertion model. These discoveries provide valuable insights into the initiation of DSB repair on transcription templates across bacteria, archaea and eukaryotes.

Targeting RNA Polymerase I Transcription Activity in Osteosarcoma: Pre-Clinical Molecular and Animal Treatment Studies.

The survival rate of patients with osteosarcoma (OS) has not improved over the last 30 years. Mutations in the genes TP53, RB1 and c-Myc frequently occur in OS and enhance RNA Polymerase I (Pol I) activity, thus supporting uncontrolled cancer cell proliferation. We therefore hypothesised that Pol I inhibition may be an effective therapeutic strategy for this aggressive cancer. The Pol I inhibitor CX-5461 has demonstrated therapeutic efficacy in different cancers in pre-clinical and phase I clinical trials; thus, the effects were determined on ten human OS cell lines. Following characterisation using genome profiling and Western blotting, RNA Pol I activity, cell proliferation and cell cycle progression were evaluated in vitro, and the growth of TP53 wild-type and mutant tumours was measured in a murine allograft model and in two human xenograft OS models. CX-5461 treatment resulted in reduced ribosomal DNA (rDNA) transcription and Growth 2 (G2)-phase cell cycle arrest in all OS cell lines. Additionally, tumour growth in all allograft and xenograft OS models was effectively suppressed without apparent toxicity. Our study demonstrates the efficacy of Pol I inhibition against OS with varying genetic alterations. This study provides pre-clinical evidence to support this novel therapeutic approach in OS.

Transcriptional pause extension benefits the stand-by rather than catch-up Rho-dependent termination.

Transcriptional pause is essential for all types of termination. In this single-molecule study on bacterial Rho factor-dependent terminators, we confirm that the three Rho-dependent termination routes operate compatibly together in a single terminator, and discover that their termination efficiencies depend on the terminational pauses in unexpected ways. Evidently, the most abundant route is that Rho binds nascent RNA first and catches up with paused RNA polymerase (RNAP) and this catch-up Rho mediates simultaneous releases of transcript RNA and template DNA from RNAP. The fastest route is that the catch-up Rho effects RNA-only release and leads to 1D recycling of RNAP on DNA. The slowest route is that the RNAP-prebound stand-by Rho facilitates only the simultaneous rather than sequential releases. Among the three routes, only the stand-by Rho's termination efficiency positively correlates with pause duration, contrary to a long-standing speculation, invariably in the absence or presence of NusA/NusG factors, competitor RNAs or a crowding agent. Accordingly, the essential terminational pause does not need to be long for the catch-up Rho's terminations, and long pauses benefit only the stand-by Rho's terminations. Furthermore, the Rho-dependent termination of mgtA and ribB riboswitches is controlled mainly by modulation of the stand-by rather than catch-up termination.

Korean Red Ginseng modulates immune function by upregulating CD4+CD8+ T cells and NK cell activities on porcine.

In the present study, we investigated whether treatment with KRG improve the parameters of immune activity such as the cytotoxicity, populations of CD4+ CD8+T cell, CD3-CD172-CD8+ NK cell and CD172+ monocyte as well as natural cytotoxicity receptors such as Nkp46, Nkp44, Nkp30. In results, KRG significantly increased these immune activities. These results indicate that KRG has distinct immune-enhancing effects by increasing the roles of T cells and NK cell in porcine.

Ginsenoside-Rb1 prevents bone cartilage destruction through down-regulation of p-Akt, p-P38, and p-P65 signaling in rabbit.

BACKGROUND: Osteoarthritis (OA) is the most common joint complaint resulting in pain, disability, and loss of quality of life. On the other hand, ginsenoside-Rb1 is a plant product derived from ginseng that possesses immune-regulation and anti-inflammatory activities. However, it has been reported that different rout of administration but hydrogel-based Ginsenoside-Rb1 in an OA rabbit model has not been investigated. PURPOSE: The aim of this study was to investigate the potential effects of ginsenoside-Rb1 such as anti-arthritic activity in a rabbit knee OA model via NF- κB, PI3K/Akt, and P38/(MAPK) pathways. STUDY DESIGN: In the current study, rabbit osteoarthritis was induced by hollow trephine on the femur trochlea and the hydrogel-based Ginsenoside-Rb1 sheets were inserted on the rabbit knee to assess the anti-arthritis activity of ginsenoside-Rb1 which is sustained release. METHODS: After the hydrogel-based Rb1 sheet insert on the rabbit knee, macroscopic and micro CT was performed for investigation of chondroprotective effect. Matrix metalloproteinases (MMPs) and apoptotic expression were assessed through Immunohistochemistry and RT-PCR assay. In addition, the flow cytometry technique was used for the investigation of intracellular reactive oxygen species (ROS) production and histological changes were examined by HE, safranin O, and Masson trichrome staining method. Furthermore, the NF- κB, PI3K/Akt, and P38/(MAPK) pathways were investigated using Western blot analysis. RESULTS: Macroscopic and micro CT investigation of hydrogel-Rb1 treatment showed a dose-dependent chondroprotective effect. Immunohistochemistry and RT-PCR revealed that expression of matrix metalloproteinases (MMPs) and apoptotic markers TNF-α, caspase-3, and bax are down-regulated in a dose-dependent fashion following implantation of hydrogel-Rb. Higher levels of intracellular reactive oxygen species (ROS) were observed in the OA group. In histopathological investigation of hydrogel-Rb1 exhibited larger amounts of chondro cells, glycosaminoglycan's, and collagen compared to the defect group. Furthermore, the NF- κB, PI3K/Akt, and P38/(MAPK) pathways were downregulated by hydrogel-Rb1 while the disease model showed upstream. In the meantime, MMP expression level was considerably down-regulated. CONCLUSIONS: Our results indicate the protective effect of ginsenoside-Rb1 against OA pathogenesis through prevention of apoptosis with suppression of ROS production and activation of NF-κB signaling through downregulation of the MAPK and PI3K/Akt signaling pathways.

Rho-dependent transcription termination proceeds via three routes.

Rho is a general transcription termination factor in bacteria, but many aspects of its mechanism of action are unclear. Diverse models have been proposed for the initial interaction between the RNA polymerase (RNAP) and Rho (catch-up and stand-by pre-terminational models); for the terminational release of the RNA transcript (RNA shearing, RNAP hyper-translocation or displacing, and allosteric models); and for the post-terminational outcome (whether the RNAP dissociates or remains bound to the DNA). Here, we use single-molecule fluorescence assays to study those three steps in transcription termination mediated by E. coli Rho. We find that different mechanisms previously proposed for each step co-exist, but apparently occur on various timescales and tend to lead to specific outcomes. Our results indicate that three kinetically distinct routes take place: (1) the catch-up mode leads first to RNA shearing for RNAP recycling on DNA, and (2) later to RNAP displacement for decomposition of the transcriptional complex; (3) the last termination usually follows the stand-by mode with displacing for decomposing. This three-route model would help reconcile current controversies on the mechanisms.

The therapeutic potential of RNA Polymerase I transcription inhibitor, CX-5461, in uterine leiomyosarcoma.

BACKGROUND: Uterine leiomyosarcoma is a rare aggressive smooth muscle cancer with poor survival rates. RNA Polymerase I (Pol I) activity is elevated in many cancers supporting tumour growth and prior studies in uterine leiomyosarcoma revealed enlarged nucleoli and upregulated Pol I activity-related genes. This study aimed to investigate the anti-tumour potential of CX-5461, a Pol I transcription inhibitor currently being evaluated in clinical trials for several cancers, against the human uterine leiomyosarcoma cell line, SK-UT-1. METHODS: SK-UT-1 was characterised using genome profiling and western blotting. The anti-tumour effects of CX-5461 were investigated using cell proliferation assays, expression analysis using qRT-PCR, and BrdU/PI based cell cycle analysis. RESULTS: Genetic analysis of SK-UT-1 revealed mutations in TP53, RB1, PTEN, APC and TSC1 & 2, all potentially associated with increased Pol I activity. Protein expression analysis showed dysregulated p53, RB1 and c-Myc. CX-5461 treatment resulted in an anti-proliferation response, G2 phase cell-cycle arrest and on-target activity demonstrated by reduced ribosomal DNA transcription. CONCLUSIONS: SK-UT-1 was confirmed as a representative model of uterine leiomyosarcoma and CX-5461 has significant potential as a novel adjuvant for this rare cancer.

IGF-1 Facilitates Cartilage Reconstruction by Regulating PI3K/AKT, MAPK, and NF-kB Signaling in Rabbit Osteoarthritis.

PURPOSE: The pathogenesis of osteoarthritis (OA) is characterized by joint degeneration. The pro-inflammatory cytokine interleukin (IL)-1ß plays a vital role in the pathogenesis of OA by stimulation of specific signaling pathways like NF-κB, PI3K/Akt, and MAPKs pathways. The catabolic role of growth factors in the OA may be inhibited cytokine-activated pathogen. The purpose of this study was to investigate the potential effects of insulin-like growth factor-1 (IGF-1) on IL-1ß-induced apoptosis in rabbit chondrocytes in vitro and in an in vivo rabbit knee OA model. METHODS: In the present study, the OA developed in chondrocyte with the treatment of IL-1ß and articular cartilage ruptures by removal of cartilage from the rabbit knee femoral condyle. After IGF-1 treatment, immunohistochemistry and qRT-PCR were identified OA expression with changes in MMPs (matrix metalloproteinases). The production of ROS (intracellular reactive oxygen species) in the OA was detected by flow cytometry. Further, the disease progression was microscopically investigated and pathophysiological changes were analyzed using histology. The NF-κB, PI3K/Akt and P38 (MAPK) specific pathways that are associated with disease progression were also checked using the Western blot technique. RESULTS: The expression of MMPs and various apoptotic markers are down-regulated following administration of IGF-1 in a dose-dependent fashion while significantly up-regulation of TIMP-1. The results showed that higher levels of ROS were observed upon treatment of chondrocytes and chondral OA with IL-1ß. Collectively, our results indicated that IGF-1 protected NF-κB pathway by suppression of PI3K/Akt and MAPKs specific pathways. Furthermore, the macroscopic and pathological investigation showed that it has a chondroprotective effect by the formation of hyaline cartilage. CONCLUSION: Our results indicate a protective effect of IGF-1 against OA pathogenesis by inhibition of NF-κB signaling via regulation of the MAPK and PI3K/Akt signaling pathways and prevention of apoptosis by suppression of ROS production.

Effect of Korean Red Ginseng through comparative analysis of cardiac gene expression in db/db mice.

Korean Red Ginseng (KRG) is an herbal oriental medicine known to alleviate cardiovascular dysfunction. To analysis the expression of diabetic cardiac complication-associated genes in db/db mice, we studied the cardiac gene expression following KRG treatment. In result, a total of 585 genes were found to be changed in db/db mice. Among the changed expression, 245 genes were found to 2-fold upregulated, and 340 genes were 2-fold downregulated. In addition, the changed gene expressions were ameliorated by KRG. In conclusion, KRG may be possible to normalize cardiac gene expressions in db/db mice.

Ginsenoside Rb1 inhibits monoiodoacetate-induced osteoarthritis in postmenopausal rats through prevention of cartilage degradation.

BACKGROUND: Ginsenoside Rb1 (G-Rb1), one of the major active compounds in Panax ginseng, has already been shown to reduce inflammation in various diseases. Osteoarthritis (OA) has traditionally been considered a degenerative disease with degradation of joint articular cartilage. However, recent studies have shown the association of inflammation with OA. In the present study, we investigated whether Rb1 had an antiinflammatory effect on monoiodoacetate (MIA)-induced OA in ovariectomized rats as a model of postmenopausal arthritis. METHODS: G-Rb1 at a dosage of 3 and 10 µg/kg body weight was administered every 3 days intraarticularly for a period of 4 weeks to observe antiarthritic effects. Diclofenac (10 mg/kg) served as a positive control. RESULTS: The administration of Rb1 significantly ameliorated OA inflammatory symptoms and reduced serum levels of inflammatory cytokines. Furthermore, G-Rb1 administration considerably enhanced the expression of bone morphogenetic protein-2 and collagen 2A and reduced the levels of matrix metalloproteinase-13 genes, indicating a chondroprotective effect of G-Rb1. G-Rb1 also significantly reduced the expression of several inflammatory cytokines/chemokines (interferon gamma (IFN-γ), monocyte chemoattractant protein-1 (MCP-1)/CCL-2, interleukin [IL]-1ß, and IL-6). Histological analysis demonstrated that G-Rb1 significantly attenuated the pathological changes in MIA-induced OA in ovariectomized rats. Safranin O and toluidine blue staining further demonstrated that G-Rb1 effectively prevented the degradation of cartilage and glycosaminoglycans, respectively. CONCLUSION: Overall, our results suggest that G-Rb1 exerts cartilage protective effect on MIA-induced ovariectomized OA rats, by inhibiting inflammatory mediators such as IL-6, IL-1ß, MCP-1/CCL-2, cyclooxygenase-2 (COX-2), and prostaglandin E2 (PGE2). These results shed a light on possible therapeutic application of G-Rb1 in OA.

Hopping and Flipping of RNA Polymerase on DNA during Recycling for Reinitiation after Intrinsic Termination in Bacterial Transcription.

Two different molecular mechanisms, sliding and hopping, are employed by DNA-binding proteins for their one-dimensional facilitated diffusion on nonspecific DNA regions until reaching their specific target sequences. While it has been controversial whether RNA polymerases (RNAPs) use one-dimensional diffusion in targeting their promoters for transcription initiation, two recent single-molecule studies discovered that post-terminational RNAPs use one-dimensional diffusion for their reinitiation on the same DNA molecules. Escherichia coli RNAP, after synthesizing and releasing product RNA at intrinsic termination, mostly remains bound on DNA and diffuses in both forward and backward directions for recycling, which facilitates reinitiation on nearby promoters. However, it has remained unsolved which mechanism of one-dimensional diffusion is employed by recycling RNAP between termination and reinitiation. Single-molecule fluorescence measurements in this study reveal that post-terminational RNAPs undergo hopping diffusion during recycling on DNA, as their one-dimensional diffusion coefficients increase with rising salt concentrations. We additionally find that reinitiation can occur on promoters positioned in sense and antisense orientations with comparable efficiencies, so reinitiation efficiency depends primarily on distance rather than direction of recycling diffusion. This additional finding confirms that orientation change or flipping of RNAP with respect to DNA efficiently occurs as expected from hopping diffusion.

Korean Red Ginseng enhances cardiac hemodynamics on doxorubicin-induced toxicity in rats.

BACKGROUND: Korean Red Ginseng (KRG) has been known to possess many ginsenosides. These ginsenosides are used for curing cardiovascular problems. The present study show the protective potential of KRG against doxorubicin (DOX)-induced myocardial dysfunction, by assessing electrocardiographic, hemodynamic, and biochemical parameters and histopathological findings. METHODS: Animals were fed a standard chow and adjusted to their environment for 3 days before the experiments. Next, the rats were equally divided into five groups (n = 9, each group). The animals were administered with KRG (250 and 500 mg/kg) for 10 days and injected with DOX (20 mg/kg, subcutaneously, twice at a 24-h interval) on the 8th and 9th day. Electrocardiography and echocardiography were performed to study hemodynamics. Plasma levels of superoxide dismutase, catalase, glutathione peroxidase, and malondialdehyde were measured. In addition, the dose of troponin I and activity of myeloperoxidase in serum and cardiac tissue were analyzed, and the histopathological findings were evaluated using light microscopy. RESULTS: Administration of KRG at a dose of 250 and 500 mg/kg recovered electrocardiographic changes, ejection fraction, fractional shortening, left ventricular systolic pressure, the maximal rate of change in left ventricle contraction (+dP/dtmax), and left ventricle relaxation (-dP/dtmax). In addition, KRG treatment significantly normalized the oxidative stress markers in plasma, dose dependently. In addition, the values of troponin I and myeloperoxidase were ameliorated by KRG treatment, dose dependently. And, KRG treatment showed better histopathological findings when compared with the DOX control group. CONCLUSION: These mean that KRG mitigates myocardial damage by modulating the hemodynamics, histopathological abnormality, and oxidative stress related to DOX-induced cardiomyopathy in rats. The results of the present study show protective effects of KRG on cardiac toxicity.

Synergistic activation of NF-κB by TNFAIP3 (A20) reduction and UBE2L3 (UBCH7) augment that synergistically elevate lupus risk.

BACKGROUND: Systemic lupus erythematosus (SLE) is an autoimmune inflammatory rheumatic disease. SLE susceptibility is affected by multiple genetic elements, environmental factors, and their interactions. We aimed in this study to statistically and functionally characterize a gene-gene interaction (epistasis) recently documented to affect SLE risk. METHODS: Two single-nucleotide polymorphisms, rs2230926 in TNFAIP3 (A20) gene and rs131654 in UBE2L3 (UBCH7) gene, were genotyped in all 3525 Korean participants, and their SLE risk association and epistasis were statistically analyzed by calculating odds ratio (OR), 95% confidence interval (CI), and P values in genotype comparisons between 1318 SLE patients and 2207 healthy controls. Furthermore, their effects on gene functions were assessed by comparatively examining separate and combined effects of TNFAIP3 and UBE2L3 knockdowns on NF-κB transcription factor activity in human cells. RESULTS: SLE susceptibility is associated with TNFAIP3 rs2230926 (OR = 1.9, 95% CI 1.6-2.4, P = 8.6 × 10-11) and UBE2L3 rs131654 (OR = 1.2, 95% CI 1.1-1.4, P = 1.1 × 10-4) in a Korean population of this study. Their risk-associated alleles synergistically elevate SLE susceptibility in both multivariate logistic regression analysis (ORinteraction = 1.6, P = 0.0028) and genotype-stratified analysis (ORinteraction = 2.4), confirming the synergistic TNFAIP3-UBE2L3 interaction in SLE risk. Additionally, the SLE-susceptible alleles confer decreased TNFAIP3 expression (P = 1.1 × 10-6, n = 610) and increased UBE2L3 expression (P = 9.5 × 10-11, n = 475), respectively, in B cell analysis of the International HapMap Project individuals with adjustment for ethnicity. Furthermore, when compared with TNFAIP3 non-knockdown and UBE2L3 knockdown in human HeLa cells, TNFAIP3 knockdown and UBE2L3 non-knockdown synergistically increase three cytokines, CCL2, CXCL8 (IL8), and IL6, all regulated by NF-κB in the human TNFR signaling pathway. CONCLUSIONS: A synergistic interaction between TNFAIP3 and UBE2L3 genes is observed in SLE risk, as being evident in comparison of genotype distributions between SLE patients and controls. Additionally, the synergistic gene-gene interaction is functionally validated, as TNFAIP3 reduction and UBE2L3 augment exert synergism in activation of NF-κB and subsequent induction of inflammatory cytokines. Accordingly, SLE inflammation and risk could be synergistically alleviated by TNFAIP3 upregulation and UBE2L3 downregulation.

A gastric cancer cell derived extracellular compounds suppresses CD161+CD3- lymphocytes and aggravates tumor formation in a syngeneic mouse model.

Evasion of the immune system is often associated with malignant tumors. The cancer cell microenvironment plays an important role in tumor progression, but its mechanism is largely unknown. Here we show that an extracellular compound derived from gastric cancer (GC-EC) selectively suppresses CD161+CD3- natural killer (NK) cells. Splenocytes treated with GC-EC showed considerable proliferation and the CD161+CD3- NK cell population was time-dependently suppressed. Intracellular staining of IFN-γ was shown to be down-regulated in concert with granzyme B and perforin. A cytotoxicity assay of splenocytes treated with GC-EC against K-562 cells showed a significant reduction in cytolytic activity. Further, the immune-suppressive effect of GC-EC was more evident in a syngeneic tumor model in C57BL/6 mice. Animals treated with B16 F10 and GC-EC exhibited more aggravated tumor formation than animals treated with B16 F10 only. We demonstrated that inhibition of apoptosis while increasing PI3 K/AKT levels may provoke tumor formation by GC-EC. A cytokine array revealed the presence of several cytokines in GC-EC that negatively regulate immune cytolytic activity and could be potential candidates for immune-suppressive effects.

Korean Red Ginseng attenuates type 2 diabetic cardiovascular dysfunction in Otsuka Long-Evans Tokushima Fatty rats.

Extracts of ginseng species show antihyperglycemic activity. We evaluated the inhibitory effects of diabetic complications for Korean Red Ginseng (KRG), which is enriched in ginsenosides using Otsuka Long-Evans Tokushima Fatty (OLETF) rats. The animals were divided into one of four groups (n = 6∼9): Long-Evans-Tokushima-Otsuka rats (control rats), OLETF rats, rats given 200 mg/kg KRG, and rats given 400 mg/kg KRG. We examined the protective potential of KRG against type 2 diabetic illnesses. The results exhibited that KRG showed significant antihyperglycemic and antioxidative effects in KRG-treated OLETF rats. And, our results proposed the amelioration of cardiac function through normalized ejection fraction, fractional shortening, and vascular reactivity. Furthermore, histopathological abnormalities in the OLETF rats were prevented by KRG treatment.

Transcription reinitiation by recycling RNA polymerase that diffuses on DNA after releasing terminated RNA.

Despite extensive studies on transcription mechanisms, it is unknown how termination complexes are disassembled, especially in what order the essential components dissociate. Our single-molecule fluorescence study unveils that RNA transcript release precedes RNA polymerase (RNAP) dissociation from the DNA template much more often than their concurrent dissociations in intrinsic termination of bacterial transcription. As termination is defined by the release of product RNA from the transcription complex, the subsequent retention of RNAP on DNA constitutes a previously unidentified stage, termed here as recycling. During the recycling stage, post-terminational RNAPs one-dimensionally diffuse on DNA in downward and upward directions, and can initiate transcription again at the original and nearby promoters in the case of retaining a sigma factor. The efficiency of this event, termed here as reinitiation, increases with supplement of a sigma factor. In summary, after releasing RNA product at intrinsic termination, recycling RNAP diffuses on the DNA template for reinitiation most of the time.

Synthesis of Enantiopure ε-Oxapipecolic Acid.

A six-step synthesis of orthogonally protected (S)-ε-oxapipecolic acid is described, starting from a commercially available glutamate diester. The approach features mCPBA-mediated amine oxidation and an intramolecular Mitsunobu reaction to form the tetrahydrooxazine ring. The enantiopure building block was employed in the synthesis of a short model peptide to determine the amide rotamer preference N-terminal to the cyclic residue. In contrast to pipecolic acid, which exhibits a high cis amide population, the ε heteroatom in oxapipecolic acid exerts a strong trans substantiating effect through lone pair repulsion.