Identification of a Second Type of AHL-lactonase from Rhodococcus sp. BH4, belonging to the α/ß Hydrolase Superfamily.

N-acyl-homoserine lactone (AHL)-mediated quorum sensing (QS) plays a major role in development of biofilms, which contribute to rise in infections and biofouling in water-related industries. Interference in QS, called quorum quenching (QQ), has recieved a lot of attention in recent years. Rhodococcus spp. are known to have prominent quorum quenching activity and in previous reports it was suggested that this genus possesses multiple QQ enzymes, but only one gene, qsdA, which encodes an AHL-lactonase belonging to phosphotriesterase family, has been identified. Therefore, we conducted a whole genome sequencing and analysis of Rhodococcus sp. BH4 isolated from a wastewater treatment plant. The sequencing revealed another gene encoding a QQ enzyme (named jydB) that exhibited a high AHL degrading activity. This QQ enzyme had a 46% amino acid sequence similarity with the AHL-lactonase (AidH) of Ochrobactrum sp. T63. HPLC analysis and AHL restoration experiments by acidification revealed that the jydB gene encodes an AHL-lactonase which shares the known characteristics of the α/ß hydrolase family. Purified recombinant JydB demonstrated a high hydrolytic activity against various AHLs. Kinetic analysis of JydB revealed a high catalytic efficiency (kcat/KM) against C4-HSL and 3-oxo-C6 HSL, ranging from 1.88 × 106 to 1.45 × 106 M-1 s-1, with distinctly low KM values (0.16 - 0.24 mM). This study affirms that the AHL degrading activity and biofilm inhibition ability of Rhodococcus sp. BH4 may be due to the presence of multiple quorum quenching enzymes, including two types of AHL-lactonases, in addition to AHL-acylase and oxidoreductase, for which the genes have yet to be described.

Stopping Autoinducer-2 Chatter by Means of an Indigenous Bacterium ( Acinetobacter sp. DKY-1): A New Antibiofouling Strategy in a Membrane Bioreactor for Wastewater Treatment.

Bacterial quorum quenching (QQ) by means of degrading signaling molecules has been applied to antibiofouling strategies in a membrane bioreactor (MBR) for wastewater treatment. However, the target signaling molecules have been limited to N-acyl homoserine lactones participating in intraspecies quorum sensing. Here, an approach to disrupting autoinducer-2 (AI-2) signaling molecules participating in interspecies quorum sensing was pursued as a next-generation antibiofouling strategy in an MBR for wastewater treatment. We isolated an indigenous QQ bacterium ( Acinetobacter sp. DKY-1) that can attenuate the expression of the quorum-sensing (QS) response through the inactivation of an autoinducer-2 signaling molecule, 4,5-dihydroxy-2,3-pentanedione (DPD), among four kinds of autoinducer-2 QS bacteria. DKY-1 released AI-2 QQ compounds, which were verified to be hydrophilic with a molecular weight of <400 Da. The addition of DKY-1 entrapping beads into an MBR significantly decreased DPD concentration and remarkably reduced membrane biofouling. This new approach, combining molecular biology with wastewater engineering, could enlarge the range of QQ-MBR for antibiofouling and energy savings in the field of wastewater treatment.

Mutation of the cyclic di-GMP phosphodiesterase gene in Burkholderia lata SK875 attenuates virulence and enhances biofilm formation.

Burkholderia sp. is a gram-negative bacterium that commonly exists in the environment, and can cause diseases in plants, animals, and humans. Here, a transposon mutant library of a Burkholderia lata isolate from a pig with swine respiratory disease in Korea was screened for strains showing attenuated virulence in Caenorhabditis elegans. One such mutant was obtained, and the Tn5 insertion junction was mapped to rpfR, a gene encoding a cyclic di-GMP phosphodiesterase that functions as a receptor. Mutation of rpfR caused a reduction in growth on CPG agar and swimming motility as well as a rough colony morphology on Congo red agar. TLC analysis showed reduced AHL secretion, which was in agreement with the results from plate-based and bioluminescence assays. The mutant strain produced significantly more biofilm detected by crystal violet staining than the parent strain. SEM of the mutant strain clearly showed that the overproduced biofilm contained a filamentous structure. These results suggest that the cyclic di-GMP phosphodiesterase RpfR plays an important role in quorum sensing modulation of the bacterial virulence and biofilm formation.

Mitigation of Membrane Biofouling in MBR Using a Cellulolytic Bacterium, Undibacterium sp. DM-1, Isolated from Activated Sludge.

Biofilm formation on the membrane surface results in the loss of permeability in membrane bioreactors (MBRs) for wastewater treatment. Studies have revealed that cellulose is not only produced by a number of bacterial species but also plays a key role during formation of their biofilm. Hence, in this study, cellulase was introduced to a MBR as a cellulose-induced biofilm control strategy. For practical application of cellulase to MBR, a cellulolytic (i.e., cellulase-producing) bacterium, Undibacterium sp. DM-1, was isolated from a lab-scale MBR for wastewater treatment. Prior to its application to MBR, it was confirmed that the cell-free supernatant of DM-1 was capable of inhibiting biofilm formation and of detaching the mature biofilm of activated sludge and cellulose-producing bacteria. This suggested that cellulase could be an effective anti-biofouling agent for MBRs used in wastewater treatment. Undibacterium sp. DM-1-entrapping beads (i.e., cellulolytic-beads) were applied to a continuous MBR to mitigate membrane biofouling 2.2-fold, compared with an MBR with vacant-beads as a control. Subsequent analysis of the cellulose content in the biofilm formed on the membrane surface revealed that this mitigation was associated with an approximately 30% reduction in cellulose by cellulolytic-beads in MBR.

Enhancing the Physical Properties and Lifespan of Bacterial Quorum Quenching Media through Combination of Ionic Cross-Linking and Dehydration.

Quorum quenching (QQ) bacteria entrapped in a polymeric composite hydrogel (QQ medium) have been successfully applied in membrane bioreactors (MBRs) for effective biofouling control. However, in order to bring QQ technology closer to practice, the physical strength and lifetime of QQ media should be improved. In this study, enforcement of physical strength, as well as an extension of the lifetime of a previously reported QQ bacteria entrapping hollow cylinder (QQ-HC), was sought by adding a dehydration procedure following the cross-linking of the polymeric hydrogel by inorganic compounds like Ca2+ and boric acid. Such prepared medium demonstrated enhanced physical strength possibly through an increased degree of physical cross-linking. As a result, a longer lifetime of QQ-HCs was confirmed, which led to improved biofouling mitigation performance of QQ-HC in an MBR. Furthermore, QQ-HCs stored under dehydrated condition showed higher QQ activity when the storage time lasted more than 90 days owing to enhanced cell viability. In addition, the dormant QQ activity after the dehydration step could be easily restored through reactivation with real wastewater, and the reduced weight of the dehydrated media is expected to make handling and transportation of QQ media highly convenient and economical in practice.

Fungal Quorum Quenching: A Paradigm Shift for Energy Savings in Membrane Bioreactor (MBR) for Wastewater Treatment.

In the last 30 years, the use of membrane bioreactors (MBRs) for advanced wastewater treatment and reuse have been expanded continuously, but they still suffer from excessive energy consumption resulting from the intrinsic problem of membrane biofouling. One of the major causes of biofouling in MBRs is bacterial quorum sensing (QS) via N-acylhomoserine lactones (AHLs) and/or autoinducer-2 (AI-2), enabling intra- and interspecies communications, respectively. In this study, we demonstrate that farnesol can substantially mitigate membrane biofouling in a MBR due to its quorum quenching (QQ) activity. When Candida albicans (a farnesol producing fungus) entrapping polymer beads (AEBs) were placed in the MBR, the rate of transmembrane pressure (TMP) rise-up was substantially decreased, even for lower aeration intensities. This finding corresponds to a specific aeration energy savings of approximately 40% (25% through the physical washing effect and a further 15% through the biological QQ effect of AEBs) compared to conventional MBRs without AEBs. A real-time RT-qPCR analysis revealed that farnesol secreted from C. albicans mitigated the biofilm formation in MBRs via the suppression of AI-2 QS. Successful control of biofouling and energy savings through fungal-to-bacterial QQ could be expanded to the plant scale for MBRs in wastewater treatment with economic feasibility.

Spatial reproducibility of complex fractionated atrial electrogram depending on the direction and configuration of bipolar electrodes: an in-silico modeling study.

Although 3D-complex fractionated atrial electrogram (CFAE) mapping is useful in radiofrequency catheter ablation for persistent atrial fibrillation (AF), the directions and configuration of the bipolar electrodes may affect the electrogram. This study aimed to compare the spatial reproducibility of CFAE by changing the catheter orientations and electrode distance in an in-silico left atrium (LA). We conducted this study by importing the heart CT image of a patient with AF into a 3D-homogeneous human LA model. Electrogram morphology, CFAE-cycle lengths (CLs) were compared for 16 different orientations of a virtual bipolar conventional catheter (conv-cath: size 3.5 mm, inter-electrode distance 4.75 mm). Additionally, the spatial correlations of CFAE-CLs and the percentage of consistent sites with CFAE-CL<120 ms were analyzed. The results from the conv-cath were compared with that obtained using a mini catheter (mini-cath: size 1 mm, inter-electrode distance 2.5 mm). Depending on the catheter orientation, the electrogram morphology and CFAE-CLs varied (conv-cath: 11.5±0.7% variation, mini-cath: 7.1±1.2% variation), however the mini-cath produced less variation of CFAE-CL than conv-cath (p<0.001). There were moderate spatial correlations among CFAE-CL measured at 16 orientations (conv-cath: r=0.3055±0.2194 vs. mini-cath: 0.6074±0.0733, p<0.001). Additionally, the ratio of consistent CFAE sites was higher for mini catheter than conventional one (38.3±4.6% vs. 22.3±1.4%, p<0.05). Electrograms and CFAE distribution are affected by catheter orientation and electrode configuration in the in-silico LA model. However, there was moderate spatial consistency of CFAE areas, and narrowly spaced bipolar catheters were less influenced by catheter direction than conventional catheters.

Effect of the Shape and Size of Quorum-Quenching Media on Biofouling Control in Membrane Bioreactors for Wastewater Treatment.

Recently, spherical beads entrapping quorum quenching (QQ) bacteria have been reported as effective moving QQ-media for biofouling control in MBRs for wastewater treatment owing to their combined effects of biological (i.e., quorum quenching) and physical washing. Taking into account both the mass transfer of signal molecules through the QQ-medium and collision efficiencies of the QQ-medium against the filtration membranes in a bioreactor, a cylindrical medium (QQ-cylinder) was developed as a new shape of moving QQ-medium. The QQ-cylinders were compared with previous QQ-beads in terms of the QQ activity and the physical washing effect under identical loading volumes of each medium in batch tests. It was found that the QQ activity of a QQ-medium was highly dependent on its specific surface area, regardless of the shape of the medium. In contrast, the physical washing effect of a QQ-medium was greatly affected by its geometric structure. The enhanced anti-biofouling property of the QQ-cylinders relative to QQ-beads was confirmed in a continuous laboratory-scale MBR with a flat-sheet membrane module.

Identification of the Regulators Binding to the Upstream Region of glxR in Corynebacterium glutamicum.

GlxR is considered as a global transcriptional regulator controlling a large number of genes having broad physiological aspects in Corynebacterium glutamicum. However, the expression profile revealing the transcriptional control of glxR has not yet been studied in detail. DNA affinity chromatography experiments revealed the binding of transcriptional regulators SucR, RamB, GlxR, and a GntR-type protein (hereafter denoted as GntR3) to the upstream region of glxR. The binding of different regulators to the glxR promoter was confirmed by EMSA experiments. The expression of glxR was analyzed in detail under various carbon sources in the wild-type and different mutant strains. The sucR and gntR3 deletion mutants showed decreased glxR promoter activities, when compared with the wild type, irrespective of the carbon sources. The promoter activity of glxR was derepressed in the ramB deletion mutant under all the tested carbon sources. These results indicate that SucR and GntR3 are acting as activators of GlxR, while RamB plays a repressor. As expected, the expression of glxR in the cyaB and glxR deletion mutants was derepressed under different media conditions, indicating that GlxR is autoregulated.

Quorum quenching bacteria isolated from the sludge of a wastewater treatment plant and their application for controlling biofilm formation.

Bacteria recognize changes in their population density by sensing the concentration of signal molecules, N-acyl-homoserine lactones (AHLs). AHL-mediated quorum sensing (QS) plays a key role in biofilm formation, so the interference of QS, referred to as quorum quenching (QQ), has received a great deal of attention. A QQ strategy can be applied to membrane bioreactors (MBRs) for advanced wastewater treatment to control biofouling. To isolate QQ bacteria that can inhibit biofilm formation, we isolated diverse AHL-degrading bacteria from a laboratory-scale MBR and sludge from real wastewater treatment plants. A total of 225 AHLdegrading bacteria were isolated from the sludge sample by enrichment culture. To identify the enzyme responsible for AHL degradation in QQ bacteria, AHL-degrading activities were analyzed using cell-free lysate, culture supernatant, and whole cells. Afipia sp. and Acinetobacter sp. strains produced the intracellular QQ enzyme, whereas Pseudomonas sp. and Micrococcus sp. produced the extracellular QQ enzyme that was most likely to produce AHLacylase. AHL-degrading activity was observed in whole-cell assay with the Microbacterium sp. and Rhodococcus sp. strains. There has been no report for AHL-degrading capability in the case of Streptococcus sp. and Afipia sp. strains. Finally, inhibition of biofilm formation by isolated QQ bacteria or enzymes was observed on glass slides and 96-well microtiter plates using crystal violet staining. QQ strains or enzymes not only inhibited initial biofilm development but also reduced established biofilms.

Design of quorum quenching microbial vessel to enhance cell viability for biofouling control in membrane bioreactor.

Quorum quenching (QQ) with a microbial vessel has recently been reported as an economically feasible biofouling control platform in a membrane bioreactor (MBR) for wastewater treatment. In this study, a quorum quenching MBR with a ceramic microbial vessel (CMV) was designed to overcome the extremely low F/M ratio inside a microbial vessel. The CMV was prepared with a monolithic ceramic microporous membrane and AHLdegrading QQ bacteria, Pseudomonas sp. 1A1. The "inner flow feeding mode" was introduced, under which fresh feed was supplied to the MBR only through the center lumen in the CMV. The inner flow feeding mode facilitated nutrient transport to QQ bacteria in the CMV and thus enabled relatively long-term maintenance of cell viability. The quorum quenching effect of the CMV on controlling membrane biofouling in the MBR was more pronounced with the inner flow feeding mode, which was identified by the slower increase in the transmembrane pressure as well as by the visual observation of a biocake that formed on the used membrane surface. In the QQ MBR with the CMV, the concentrations of extracellular polymeric substances were substantially decreased in the biocake on the membrane surface compared with those in the conventional MBR. The CMV also showed its potential with effective biofouling control over long-term operation of the QQ MBR.

Elucidation of the regulation of ethanol catabolic genes and ptsG using a glxR and adenylate cyclase gene (cyaB) deletion mutants of Corynebacterium glutamicum ATCC 13032.

The cyclic AMP receptor protein (CRP) homolog, GlxR, controls the expression of several genes involved in the regulation of diverse physiological processes in Corynebacterium glutamicum. In silico analysis has revealed the presence of glxR binding sites upstream of genes adhA, ald, and ptsG, encoding glucose-specific phosphotransferase system protein, alcohol dehydrogenase (ADH), and acetaldehyde dehydrogenase (ALDH), respectively. However, the involvement of the GlxR-cAMP complex on the expression of these genes has been explored only in vitro. In this study, the expressions of ptsG, adhA, and ald were analyzed in detail using an adenylate cyclase gene (cyaB) deletion mutant and glxR deletion mutant. The specific activities of ADH and ALDH were increased in both the mutants in glucose and glucose plus ethanol media, in contrast to the wild type. In accordance, the promoter activities of adhA and ald were derepressed in the cyaB mutant, indicating that glxR acts as a repressor of adhA. Similarly, both the mutants exhibited derepression of ptsG regardless of the carbon source. These results confirm the involvement of GlxR on the expression of important carbon metabolic genes; adhA, ald, and ptsG.

Biofouling inhibition in MBR by Rhodococcus sp. BH4 isolated from real MBR plant.

It has been reported that an indigenous quorum quenching bacterium, Rhodococcus sp. BH4, which was isolated from a real plant of membrane bioreactor (MBR) has promising potential to control biofouling in MBR. However, little is known about quorum quenching mechanisms by the strain BH4. In this study, various characteristics of strain BH4 were investigated to elucidate its behavior in more detail in the mixed liquor of MBR. The N-acyl homoserine lactone hydrolase (AHL-lactonase) gene of strain BH4 showed a high degree of identity to qsdA in Rhodococcus erythropolis W2. The LC-ESI-MS analysis of the degradation product by strain BH4 confirmed that it inactivated AHL activity by hydrolyzing the lactone bond of AHL. It degraded a wide range of N-acyl homoserine lactones (AHLs), but there was a large difference in the degradation rate of each AHL compared to other reported AHL-lactonase-producing strains belonging to Rhodococcus genus. Its quorum quenching activity was confirmed not only in the Luria-Bertani medium, but also in the synthetic wastewater. Furthermore, the amount of strain BH4 encapsulated in the vessel as well as the material of the vessel substantially affected the quorum quenching activity of strain BH4, which provides useful information, particularly for the biofouling control in a real MBR plant from an engineering point of view.

Modulation of Quorum Sensing in Acylhomoserine Lactone-Producing or -Degrading Tobacco Plants Leads to Alteration of Induced Systemic Resistance Elicited by the Rhizobacterium Serratia marcescens 90-166.

Numerous root-associated bacteria (rhizobacteria) are known to elicit induced systemic resistance (ISR) in plants. Bacterial cell-density-dependent quorum sensing (QS) is thought to be important for ISR. Here, we investigated the role of QS in the ISR elicited by the rhizobacterium, Serratia marcescens strain 90-166, in tobacco. Since S. marcescens 90-166 produces at least three QS signals, QS-mediated ISR in strain 90-166 has been difficult to understand. Therefore, we investigated the ISR capacity of two transgenic tobacco (Nicotiana tabacum) plants that contained either bacterial acylhomoserine lactone-producing (AHL) or -degrading (AiiA) genes in conjunction with S. marcescens 90-166 to induce resistance against bacterial and viral pathogens. Root application of S. marcescens 90-166 increased ISR to the bacterial pathogens, Pectobacterium carotovorum subsp. carotovorum and Pseudomonas syringae pv. tabaci, in AHL plants and decreased ISR in AiiA plants. In contrast, ISR to Cucumber mosaic virus was reduced in AHL plants treated with S. marcescens 90-166 but enhanced in AiiA plants. Taken together, these data indicate that QS-dependent ISR is elicited by S. marcescens 90-166 in a pathogen-dependent manner. This study provides insight into QS-dependent ISR in tobacco elicited by S. marcescens 90-166.

Microbial population dynamics and proteomics in membrane bioreactors with enzymatic quorum quenching.

Quorum sensing gives rise to biofilm formation on the membrane surface, which in turn causes a loss of water permeability in membrane bioreactors (MBRs) for wastewater treatment. Enzymatic quorum quenching was reported to successfully inhibit the formation of biofilm in MBRs through the decomposition of signal molecules, N-acyl homoserine lactones (AHLs). The aim of this study was to elucidate the mechanisms of quorum quenching in more detail in terms of microbial population dynamics and proteomics. Microbial communities in MBRs with and without a quorum quenching enzyme (acylase) were analyzed using pyrosequencing and compared with each other. In the quorum quenching MBR, the rate of transmembrane pressure (TMP) rise-up was delayed substantially, and the proportion of quorum sensing bacteria with AHL-like autoinducers (such as Enterobacter, Pseudomonas, and Acinetobacter) also decreased in the entire microbial community of mature biofilm in comparison to that in the control MBR. These factors were attributed to the lower production of extracellular polymeric substances (EPS), which are known to play a key role in the formation of biofilm. Proteomic analysis using the Enterobacter cancerogenus strain ATCC 35316 demonstrates the possible depression of protein expression related to microbial attachments to solid surfaces (outer membrane protein, flagellin) and the agglomeration of microorganisms (ATP synthase beta subunit) with the enzymatic quorum quenching. It has been argued that changes in the microbial population, EPS and proteins via enzymatic quorum quenching could inhibit the formation of biofilm, resulting in less biofouling in the quorum quenching MBR.

Biofouling control with bead-entrapped quorum quenching bacteria in membrane bioreactors: physical and biological effects.

Recently, interspecies quorum quenching by bacterial cells encapsulated in a vessel was described and shown to be efficient and economically feasible for biofouling control in membrane bioreactors (MBRs). In this study, free-moving beads entrapped with quorum quenching bacteria were applied to the inhibition of biofouling in a MBR. Cell entrapping beads (CEBs) with a porous microstructure were prepared by entrapping quorum quenching bacteria ( Rhodococcus sp. BH4) into alginate beads. In MBRs provided with CEBs, the time to reach a transmembrane pressure (TMP) of 70 kPa was 10 times longer than without CEBs. The mitigation of biofouling was attributed to both physical (friction) and biological (quorum quenching) effects of CEBs, the latter being much more important. Because of the quorum quenching effect of CEBs, microbial cells in the biofilm generated fewer extracellular polymeric substances and thus formed a loosely bound biofilm, which enabled it to slough off from the membrane surface more easily. Furthermore, collisions between the moving CEBs and membranes gave rise to frictional forces that facilitated detachment of the biofilm from the membrane surface. CEBs bring bacterial quorum quenching closer to being a practical solution to the problem of biofouling in MBRs.

Control of membrane biofouling in MBR for wastewater treatment by quorum quenching bacteria encapsulated in microporous membrane.

Recently, enzymatic quorum quenching has proven its potential as an innovative approach for biofouling control in the membrane bioreactor (MBR) for advanced wastewater treatment. However, practical issues on the cost and stability of enzymes are yet to be solved, which requires more effective quorum quenching methods. In this study, a novel quorum quenching strategy, interspecies quorum quenching by bacterial cell, was elaborated and proved to be efficient and economically feasible biofouling control in MBR. A recombinant Escherichia coli which producing N-acyl homoserine lactonase or quorum quenching Rhodococcus sp. isolated from a real MBR plant was encapsulated inside the lumen of microporous hollow fiber membrane, respectively. The porous membrane containing these functional bacteria (i.e., "microbial-vessel") was put into the submerged MBR to alleviate biofouling on the surface of filtration membrane. The effect of biofouling inhibition by the microbial-vessel was evaluated over 80 days of MBR operation. Successful control of biofouling in a laboratory scale MBR suggests that the biofouling control through the interspecies quorum quenching could be expanded to the plant scale of MBR and various environmental engineering systems with economic feasibility.

Mitomycin C and doxorubicin elicit conflicting signals by causing accumulation of cyclin E prior to p21WAF1/CIP1 elevation in human hepatocellular carcinoma cells.

Proteins involved in the G1 phase of the cell cycle are aberrantly expressed, sometimes in mutated forms, in human cancers including human hepatocellular carcinoma. Upon attack by a DNA-damaging anticancer drug, a cell arrests at the G1 phase; this is a safety feature prohibiting entry of DNA-damaged cells into S-phase. p21WAF1/CIP1 prevents damaged cells from progressing to the next cell cycle. Here, we show that, in response to mitomycin C and doxorubicin, human hepatocellular carcinoma cells generate conflicting signals, mediated by cyclin E and p21WAF1/CIP1, which respectively accelerates and represses cell cycle transition. Exposure to these anticancer drugs led to rapid accumulation of cyclin E in both p53-proficient HepG2 and p53-deficient Hep3B cells. Such anticancer drug-induced cyclin E accumulation influenced the G1-S-phase transition, but not DNA fragmentation-mediated death. In p53-proficient HepG2 cells, accumulation of cyclin E was followed by an increase in the level of p53-dependent p21WAF1/CIP1, thereby inhibiting further the G1-S-phase transition. Sublethal drug concentrations also induced rapid accumulation of cyclin E, but p21WAF1/CIP1 accumulation was delayed, further facilitating the G1-S-phase transition. Eventually, most cells arrested in G2/M. Thus, mitomycin C- or doxorubicin-induced conflicting signals, mediated by cyclin E and p21WAF1/CIP1, are in play in human hepatocellular carcinoma cells. Damaged G1 cells either immediately enter S-phase, or do not do so at all, depending on the extent of DNA damage.

Paclitaxel stimulates chromosomal fusion and instability in cells with dysfunctional telomeres: implication in multinucleation and chemosensitization.

The anticancer effect of paclitaxel is attributable principally to irreversible promotion of microtubule stabilization and is hampered upon development of chemoresistance by tumor cells. Telomere shortening, and eventual telomere erosion, evoke chromosomal instability, resulting in particular cellular responses. Using telomerase-deficient cells derived from mTREC-/-p53-/- mice, here we show that, upon telomere erosion, paclitaxel propagates chromosomal instability by stimulating chromosomal end-to-end fusions and delaying the development of multinucleation. The end-to-end fusions involve both the p- and q-arms in cells in which telomeres are dysfunctional. Paclitaxel-induced chromosomal fusions were accompanied by prolonged G2/M cell cycle arrest, delayed multinucleation, and apoptosis. Telomere dysfunctional cells with mutlinucleation eventually underwent apoptosis. Thus, as telomere erosion proceeds, paclitaxel stimulates chromosomal fusion and instability, and both apoptosis and chemosensitization eventually develop.

Digestion Pattern of Antihypertensive Angiotensin I-Converting Enzyme Inhibitory Peptides from Saccharomyces cerevisiae in a Successive Simulated Gastricintestinal Bioreactor.

A cell-free extract of Saccharomyces cerevisiae containing the angiotensin I-converting enzyme (ACE) inhibitory peptide was treated in a successive simulated gastric-intestinal bioreactor (step 1: amylase digestion, step 2: gastric fluid digestion, step 3: intestinal fluid digestion) to illustrate the absorption pattern of antihypertensive ACE inhibitory peptide, and the ACE inhibitory activities of each step were determined. Total ACE inhibitory activities of step 1, step 2, and step 3 were 55.96%, 80.09%, and 76.77%, respectively. The peptide sequence of each steps was analyzed by MS/MS spectrophotometry. Eleven kinds of representative peptide sequences were conserved in each step, and representative new peptides including RLPTESVPEPK were identified in step 3.