Intact Cluster Research Articles

S78. EXAMINING SEMANTIC AND EPISODIC MEMORY IN SCHIZOPHRENIA USING THE HOPKINS VERBAL LEARNING TASK

BackgroundSchizophrenia is associated with deficits in both episodic and semantic memory however, our understanding of how the deficits in each system independently contribute to overall memory performance is poorly understood.The Hopkins Verbal Learning Task (HVLT) is a memory task using a single word list. To perform the task successfully, participants need to use both episodic and semantic abilities. Both episodic and semantic clustering scores can be calculated which provide nuanced information about the memory encoding and retrieval techniques used by those performing the task.MethodsSixty schizophrenia patients and sixty healthy controls were compared in their performance on the HVLT. In addition to analysing immediate recall, learning slope, delayed recall and recognition, semantic and episodic clustering were also compared. Further, given the link between thought disorder and semantic function, this symptom was correlated with memory performance measures.ResultsThe schizophrenia group demonstrated worse performance across learning trials, delayed recall, and recognition indicating a generalised memory problem. Clustering scores were used to probe into semantic and episodic function specifically. The schizophrenia group demonstrated normal episodic clustering in the face of significantly impaired semantic clustering. Further, semantic clustering performance positively correlated with all general memory measures whilst episodic clustering did not. Finally, thought disorder did not correlate with any HVLT performance measure apart from semantic clustering.DiscussionIt is difficult to tease apart the contributions of semantic and episodic memory impairments to poor overall memory function in schizophrenia. In this study, we have first demonstrated intact episodic clustering in the face of impaired semantic clustering. Then, by correlating semantic and episodic clustering scores with general memory performance measures, we were able to demonstrate that semantic memory performance is more significantly related to overall memory performance than episodic performance. Finally, this result supports the specificity of the relationship between thought disorder and semantic memory impairment.

The outer-membrane protein TolC of Vibrio cholerae serves as a second cell-surface receptor for the VP3 phage

Receptor recognition is a key step in the initiation of phage infection. Previously, we found that VP3, the T7 family phage of the Vibrio cholerae serogroup O1 biotype El Tor, can adsorb the core oligosaccharide (OS) of lipopolysaccharides of V. cholerae However, some wildtype strains of V. cholerae possessing the intact OS gene cluster still have VP3 binding but are resistant to VP3 infection. Moreover, an OS gene-deletion mutant still exhibits weak VP3 binding, suggesting multiple factors are possibly involved in VP3 binding to V. cholerae Here, we report that the outer-membrane protein TolC of V. cholerae is involved in the host adsorption of VP3. We observed that TolC directly interacts with the VP3 tail fiber protein gp44 and its C-terminal domains, and we also found that three amino acid residues in the outside loops of TolC, at positions 78, 290, and 291, are critical for binding to gp44. Among the VP3-resistant wildtype V. cholerae strains, frequent amino acid residue mutations were observed in the loops around the sites 78, 290, and 291, which were predicted to be exposed to the cell surface. These findings reveal a co-receptor-binding mechanism for VP3 infection of V. cholerae and that both outer-membrane TolC and OS are necessary for successful VP3 infection of V. cholerae We conclude that mutations on the outside loops of the receptor may confer V. cholerae strains with VP3 phage resistance, enabling these strains to survive in environments containing VP3 or related phages.

The radical SAM protein HemW is a heme chaperone

Radical S-adenosylmethionine (SAM) enzymes exist in organisms from all kingdoms of life, and all of these proteins generate an adenosyl radical via the homolytic cleavage of the S-C(5') bond of SAM. Of particular interest are radical SAM enzymes, such as heme chaperones, that insert heme into respiratory enzymes. For example, heme chaperones insert heme into target proteins but have been studied only for the formation of cytochrome c-type hemoproteins. Here, we report that a radical SAM protein, the heme chaperone HemW from bacteria, is required for the insertion of heme b into respiratory chain enzymes. As other radical SAM proteins, HemW contains three cysteines and one SAM coordinating an [4Fe-4S] cluster, and we observed one heme per subunit of HemW. We found that an intact iron-sulfur cluster was required for HemW dimerization and HemW-catalyzed heme transfer but not for stable heme binding. A bacterial two-hybrid system screen identified bacterioferritins and the heme-containing subunit NarI of the respiratory nitrate reductase NarGHI as proteins that interact with HemW. We also noted that the bacterioferritins potentially serve as heme donors for HemW. Of note, heme that was covalently bound to HemW was actively transferred to a heme-depleted, catalytically inactive nitrate reductase, restoring its nitrate-reducing enzyme activity. Finally, the human HemW orthologue radical SAM domain-containing 1 (RSAD1) stably bound heme. In conclusion, our findings indicate that the radical SAM protein family HemW/RSAD1 is a heme chaperone catalyzing the insertion of heme into hemoproteins.

The Crown-of-Thorns Starfish: From Coral Reef Plague to Model System.

Acanthaster planci, commonly known as the "crown-of-thorns starfish" (COTS) are famous for decimating coral reefs, yet the unique features of the COTS genome make the starfish a useful system for genomic and evolutionary developmental research. The COTS genome assembly is an order of magnitude more highly contiguous than other recently sequenced echinoderm genome assemblies. The high resolution of the COTS assembly is likely related to low heterozygosity resulting from historical population dynamics and possibly a recent population expansion. The high-resolution genome assembly is biologically meaningful, as confirmed by the discovery of several intact gene clusters. Therefore, the COTS genome is an ideal testing ground for new genomic technologies and bioinformatics tools.

Tellurite resistance profiles and performance of different chromogenic agars for detection of non-O157 Shiga toxin-producing Escherichia coli

Shiga toxin-producing Escherichia coli (STEC) are globally important food-borne pathogens. The isolation of non-O157 STEC is a significant public health challenge due to the dramatic diversity of their phenotypes and genotypes. In the present study, 476 non-O157 STEC strains representing 95 different O-serogroups were used to evaluate tellurite resistance and the performance of 12 different chromogenic agars. Of 476 strains, only 108 (22.7%) strains showed the minimal inhibitory concentration (MIC) values for potassium tellurite being higher than 4μg/ml, and 96 (20.2%) strains harbored intact ter genes cluster. The presence of ter genes was significantly correlated with tellurite resistance. Six commercial chromogenic agars (TBX, MAC, SMAC, Rainbow® Agar O157, CHROMagar™ ECC, and Fluorocult O157) supported the growth of all strains. However, CT-SMAC, CHROMagar™ O157, and CHROMagar™ STEC agars exhibited 12.2%, 31.1%, and 38.0% of growth inhibition, respectively. Furthermore, 4.6%, 33.2%, and 45.0% of strains were inhibited on RBA-USDA, RBA-NT, and BCM O157 agar media. Variations in tellurite resistance and colony appearance might result in discrepant performance of non-O157 STEC recovery from different chromogenic agars. Using inclusive agars or less selective agar in combination with highly selective agar should be suggested to recover most non-O157 STEC strains, which would increase the probability of recovering STECs from complex background microflora.

Postharvest dehydration induces variable changes in the primary metabolism of grape berries

Postharvest dehydration causes changes in texture, color, taste and nutritional value of food due to the high temperatures and long drying times required. In grape berries, a gradual dehydration process is normally utilized for raisin production and for making special wines. Here we applied a raisin industry-mimicking dehydration process for eleven days at 50°C to intact berry clusters from cv. Sémillon plants, and a set of molecular, cellular and biochemical analyses were performed to study the impact of postharvest dehydration in the primary metabolism. Transcriptional analyses by real time qPCR showed that several aquaporins (VvTIP1;2 and VvSIP1) and sugar transporters (VvHT1, VvSWEET11, VvSWEET15, VvTMT1, VvSUC12) genes were strongly upregulated. Moreover, the study of key enzymes of osmolytes metabolism, including mannitol dehydrogenase (VvMTD) and sorbitol dehydrogenase (VvSDH), at gene expression and protein activity level, together with the transcriptional analysis of the polyol transporter gene VvPLT1, showed an enhanced polyol biosynthesis capacity, which was supported by the detection of sorbitol in dehydrated grapes only. The metabolism of organic acids was also modulated, by the induction of transcriptional and biochemical activity modifications in malate dehydrogenases and malic enzymes that led to organic acid degradation, as demonstrated by HPLC analysis. Taken together, this study showed that primary metabolism of harvested berries was severely influenced in response to dehydration treatments towards lower organic acid and higher sorbitol concentrations, while sugar transporter and aquaporin genes were significantly upregulated.

Laminin-111-derived peptide conjugated fibrin hydrogel restores salivary gland function.

Hyposalivation reduces the patient quality of life, as saliva is important for maintaining oral health. Current treatments for hyposalivation are limited to medications such as the muscarinic receptor agonists, pilocarpine and cevimeline. However, these therapies only provide temporary relief. Therefore, alternative therapies are essential to restore salivary gland function. An option is to use bioengineered scaffolds to promote functional salivary gland regeneration. Previous studies demonstrated that the laminin-111 protein is critical for intact salivary gland cell cluster formation and organization. However, laminin-111 protein as a whole is not suitable for clinical applications as some protein domains may contribute to unwanted side effects such as degradation, tumorigenesis and immune responses. Conversely, the use of synthetic laminin-111 peptides makes it possible to minimize the immune reactivity or pathogen transfer. In addition, it is relatively simple and inexpensive as compared to animal-derived proteins. Therefore, the goal of this study was to demonstrate whether a 20 day treatment with laminin-111-derived peptide conjugated fibrin hydrogel promotes tissue regeneration in submandibular glands of a wound healing mouse model. In this study, laminin-111-derived peptide conjugated fibrin hydrogel significantly accelerated formation of salivary gland tissue. The regenerated gland tissues displayed not only structural but also functional restoration.

New facet of non-O1/non-O139 Vibrio cholerae hemolysin A: a competitive factor in the ecological niche.

Different serogroups of Vibrio cholerae may inhabit the same ecological niche. However, serogroup O1/O139 strains are rarely isolated from their ecological sources. Quite plausibly, the non-O1/non-O139 vibrios and other bacterial species suppress growth of O1/O139 strains that share the same niche. Our bacterial inhibition assay data indicated that certain non-O1/non-O139 strains used a contact-dependent type VI secretion system (T6SS) to suppress growth of the O1 El Tor, N16961 pandemic strain. Comparative proteomics of the O1 and the suppressive non-O1/non-O139 strains co-cultured in a simulated natural aquatic microcosm showed that SecB and HlyD were upregulated in the latter. The HlyD-related effective factor was subsequently found to be hemolysin A (HlyA). However, not all hlyA-positive non-O1/non-O139 strains mediated growth suppression of the N16961 V. cholerae; only strains harboring intact cluster I HlyA could exert this activity. The key feature of the HlyA is located in the ricin-like lectin domain (β-trefoil) that plays an important role in target cell binding. In conclusion, the results of this study indicated that non-O1/non-O139 V. cholerae suppressed the growth of the O1 pandemic strain by using contact-dependent T6SS as well as by secreting the O1-detrimental hemolysin A during their co-persistence in the aquatic habitat.

Uranyl peroxide nanoclusters at high-pressure

Abstract

The cost-effectiveness of a patient centred pressure ulcer prevention care bundle: Findings from the INTACT cluster randomised trial

BackgroundPressure ulcers are serious, avoidable, costly and common adverse outcomes of healthcare. ObjectivesTo evaluate the cost-effectiveness of a patient-centred pressure ulcer prevention care bundle compared to standard care. DesignCost-effectiveness and cost-benefit analyses of pressure ulcer prevention performed from the health system perspective using data collected alongside a cluster-randomised trial. SettingsEight tertiary hospitals in Australia. ParticipantsAdult patients receiving either a patient-centred pressure ulcer prevention care bundle (n=799) or standard care (n=799). MethodsDirect costs related to the intervention and preventative strategies were collected from trial data and supplemented by micro-costing data on patient turning and skin care from a 4-week substudy (n=317). The time horizon for the economic evaluation matched the trial duration, with the endpoint being diagnosis of a new pressure ulcer, hospital discharge/transfer or 28days; whichever occurred first. For the cost-effectiveness analysis, the primary outcome was the incremental costs of prevention per additional hospital acquired pressure ulcer case avoided, estimated using a two-stage cluster-adjusted non-parametric bootstrap method. The cost-benefit analysis estimated net monetary benefit, which considered both the costs of prevention and any difference in length of stay. All costs are reported in AU$(2015). ResultsThe care bundle cost AU$144.91 (95%CI: $74.96 to $246.08) more per patient than standard care. The largest contributors to cost were clinical nurse time for repositioning and skin inspection. In the cost-effectiveness analysis, the care bundle was estimated to cost an additional $3296 (95%CI: dominant to $144,525) per pressure ulcer avoided. This estimate is highly uncertain. Length of stay was unexpectedly higher in the care bundle group. In a cost-benefit analysis which considered length of stay, the net monetary benefit for the care bundle was estimated to be −$2320 (95%CI −$3900, −$1175) per patient, suggesting the care bundle was not a cost-effective use of resources. ConclusionsA pressure ulcer prevention care bundle consisting of multicomponent nurse training and patient education may promote best practice nursing care but may not be cost-effective in preventing hospital acquired pressure ulcer.

Neurocognitive subtypes in patients with bipolar disorder and their unaffected siblings.

Our previous work revealed substantial heterogeneity in the cognitive profile of bipolar disorder (BD) due to the presence of three underlying cognitive subgroups characterized as: globally impaired, selectively impaired, or cognitively intact. In an effort to determine whether these subgroups are differentially related to genetic risk for the illness, we investigated whether cognitive deficits were more pronounced in unaffected siblings (UAS) of BD probands within identified clusters. Cluster analysis was used to identify cognitive clusters in BD (N = 60). UAS (N = 49) were classified into groups according to their proband sibling's cluster assignment; comparisons were made across all clusters and healthy controls (HCs; N = 71). Three cognitive clusters in BD emerged: a globally impaired (36.7%), a selectively impaired (30%), and a cognitively intact cluster (33.3%). UAS showed a qualitatively similar pattern to their BD siblings; UAS of the globally impaired BD cluster showed verbal memory and general cognitive impairments relative to HCs. In contrast, UAS of the other two clusters did not differ from HCs. This study corroborates findings from prior work regarding the presence of cognitive heterogeneity in BD. UAS of subjects in the globally impaired BD cluster presented with a qualitatively similar cognitive profile to their siblings and performed worse than all other BD clusters and UAS groups. This suggests that inherited risk factors may be contributing to cognitive deficits more notably in one subgroup of patients with BD, pointing toward differential causes of cognitive deficits in discrete subgroups of patients with the disorder.

Crosstalk between Lactobacillus reuteri and mammalian intestinal epithelium

Lactobacillus reuteri ATCC PTA 6475, probiotic bacterium, with an intact hdc gene cluster is known to synthesize and secrete histamine, and can suppress inflammation in mammalian systems via specific activation of the type 2 histamine receptor (H2R). However, it is unclear how L. reuteri‐derived histamine avoids activation of inflammatory H1 receptors in the intestinal epithelium. In this work, we identified a soluble secreted isoform of diacylglycerol kinase (Dgk) from L. reuteri 6475. DGKs belong to a distinct, conserved family of intracellular lipid kinases that phosphorylate diacylglycerol (DAG), catalyzing its conversion into phosphatidic acid (PA). This reaction attenuates DAG levels in the cell membrane, modifying host intracellular signaling proteins that have evolved the ability to bind this lipid for activation. Histamine binding to H1R can cause phosphorylation of PKC via lipid DAG activation. We found that L. reuteri 6475 suppressed basal levels of the pro‐inflammatory cytokines IL‐6, IL‐1, Eotaxin (eosinophilic chemoattractant proteins) and G‐CSF in the luminal mucosa and in blood plasma. L. reuteri lacking dgkA expression could not suppress the aforementioned pro‐inflammatory biomarkers. In addition, we demonstrate for the first time that histamine synthesized by L. reuteri 6475 activates both H1 and H2 receptors, but Dgk synthesized by the bacterium terminates H1R downstream signaling. Inhibition of signaling downstream of H1R was shown by decreased levels of PKC phosphorylation of intestinal epithelium in wild‐type (WT) L. reuteri treated but not in ΔdgkA mutant treated germ‐free (GF) mice. In addition, we also recorded suppression of CD11b+Gr1+ Ly6Ghi immature myeloid cells (IMCs) after WT L. reuteri treatment. In line with mouse experiment PKC phosphorylation was reduced in human epithelial cells after treating the cells with L. reuteri derived conditioned media in vitro. L. reuteri suppress immune response by direct effects of the metabolite histamine and an enzyme diacylglycerol kinase that removes a pro‐inflammatory signal. As a next step we started looking at the effect of L. reuteri and the corresponding mutants in a DSS induced colitis mouse model with conventional microbiome.Support or Funding InformationThis work was supported by the National Institutes of Health (R01 AT004326, UH3 DK083990, and U01 CA170930), the NIH (National Institute for Diabetes and Digestive and Kidney Diseases)‐funded Texas Medical Center Digestive Diseases Center (DK56338), and unrestricted research support from BioGaia AB (Stockholm, Sweden) (J.V.).

Assessing the Precision of Multisite Trials for Estimating the Parameters of a Cross-Site Population Distribution of Program Effects

ABSTRACTMultisite trials, which are being used with increasing frequency in education and evaluation research, provide an exciting opportunity for learning about how the effects of interventions or programs are distributed across sites. In particular, these studies can produce rigorous estimates of a cross-site mean effect of program assignment (intent-to-treat), a cross-site standard deviation of the effects of program assignment, and a difference between the cross-site mean effects of program assignment for two subpopulations of sites. However, to capitalize on this opportunity will require adequately powering future trials to estimate these parameters. To help researchers do so, we present a simple approach for computing the minimum detectable values of these parameters for different sample designs. The article then uses this approach to illustrate for each parameter, the precision trade-off between increasing the number of study sites and increasing site sample size. Findings are presented for multisite trials that randomize individual sample members and for multisite trials that randomize intact groups or clusters of sample members.

Characterizing cognitive heterogeneity on the schizophrenia-bipolar disorder spectrum.

Current group-average analysis suggests quantitative but not qualitative cognitive differences between schizophrenia (SZ) and bipolar disorder (BD). There is increasing recognition that cognitive within-group heterogeneity exists in both disorders, but it remains unclear as to whether between-group comparisons of performance in cognitive subgroups emerging from within each of these nosological categories uphold group-average findings. We addressed this by identifying cognitive subgroups in large samples of SZ and BD patients independently, and comparing their cognitive profiles. The utility of a cross-diagnostic clustering approach to understanding cognitive heterogeneity in these patients was also explored. Hierarchical clustering analyses were conducted using cognitive data from 1541 participants (SZ n = 564, BD n = 402, healthy control n = 575). Three qualitatively and quantitatively similar clusters emerged within each clinical group: a severely impaired cluster, a mild-moderately impaired cluster and a relatively intact cognitive cluster. A cross-diagnostic clustering solution also resulted in three subgroups and was superior in reducing cognitive heterogeneity compared with disorder clustering independently. Quantitative SZ-BD cognitive differences commonly seen using group averages did not hold when cognitive heterogeneity was factored into our sample. Members of each corresponding subgroup, irrespective of diagnosis, might be manifesting the outcome of differences in shared cognitive risk factors.

L1 Peptide–Conjugated Fibrin Hydrogels Promote Salivary Gland Regeneration

Hyposalivation contributes to dental caries, periodontitis, and microbial infections. Additionally, it impairs activities of daily living (e.g., speaking, chewing, and swallowing). Treatments for hyposalivation are currently limited to medications (e.g., the muscarinic receptor agonists pilocarpine and cevimeline) that induce saliva secretion from residual acinar cells and the use of saliva substitutes. However, given that these therapies provide only temporary relief, the development of alternative treatments to restore gland function is essential. Previous studies demonstrated that laminin 1 (L1) is critical for intact salivary cell cluster formation and organization. However, the full L1 sequence is not suitable for clinical applications, as each protein domain may contribute to unwanted effects, such as degradation, tumorigenesis, and immune responses that, when compounded, outweigh the potential benefits provided by their sum. Although the L1 peptides YIGSR and A99 linked to fibrin hydrogels (FHs) promote intact salivary epithelial formation in vitro, little is known about their role during salivary gland regeneration in vivo. Therefore, the goal of this study was to demonstrate whether L1 peptides conjugated to FHs promote tissue regeneration in a wound-healing model of mouse submandibular glands (mSMGs). Our results suggest that YIGSR-A99 peptides, chemically conjugated to FHs and applied to wounded mSMGs in vivo, formed new organized salivary tissue. In contrast, wounded mSMGs treated with FHs alone or in the absence of a scaffold showed disorganized collagen formation and poor tissue healing. Together these studies indicate that damaged salivary gland tissue can grow and differentiate when treated with FHs containing L1 peptides.

Ligand-Exchange-Induced Growth of an Atomically Precise Cu29 Nanocluster from a Smaller Cluster

The copper hydride nanocluster (NC) [Cu29Cl4H22(Ph2phen)12]Cl (2; Ph2phen = 4,7-diphenyl-1,10-phenanthroline) was isolated cleanly, and in good yields, by controlled growth from the smaller NC, [Cu25H22(PPh3)12]Cl (1), in the presence of Ph2phen and a chloride source at room temperature. Complex 2 was fully characterized by single-crystal X-ray diffraction, XANES, and XPS, and represents a rare example of an N* = 2 superatom. Its formation from 1 demonstrates that atomically precise copper clusters can be used as templates to generate larger NCs that retain the fundamental electronic and bonding properties of the original cluster. A time-resolved kinetic evaluation of the formation of 2 reveals that the mechanism of cluster growth is initiated by rapid ligand exchange. The slower extrusion of CuCl monomer, its transport, and subsequent capture by intact clusters resemble elementary steps in the reactant-assisted Ostwald ripening of metal nanoparticles.

Redox Behavior of the S-Adenosylmethionine (SAM)-Binding Fe-S Cluster in Methylthiotransferase RimO, toward Understanding Dual SAM Activity.

RimO, a radical-S-adenosylmethionine (SAM) enzyme, catalyzes the specific C3 methylthiolation of the D89 residue in the ribosomal S12 protein. Two intact iron-sulfur clusters and two SAM cofactors both are required for catalysis. By using electron paramagnetic resonance, Mössbauer spectroscopies, and site-directed mutagenesis, we show how two SAM molecules sequentially bind to the unique iron site of the radical-SAM cluster for two distinct chemical reactions in RimO. Our data establish that the two SAM molecules bind the radical-SAM cluster to the unique iron site, and spectroscopic evidence obtained under strongly reducing conditions supports a mechanism in which the first molecule of SAM causes the reoxidation of the reduced radical-SAM cluster, impeding reductive cleavage of SAM to occur and allowing SAM to methylate a HS- ligand bound to the additional cluster. Furthermore, by using density functional theory-based methods, we provide a description of the reaction mechanism that predicts the attack of the carbon radical substrate on the methylthio group attached to the additional [4Fe-4S] cluster.

Studying mass spectrometric behaviors of {Au6 Ag2 (C)[PPh2 (4-CH3 -Py)]6 }(BF4 )4 and {Au8 [(PPh3 )2 O]3 (PPh3 )2 }(NO3 )2 by electrospray time-of-flight mass spectrometry and electrospray ion trap mass spectrometry.

Mass spectrometry (MS) has been recognized as a powerful technique to detect accurate chemical information about metal clusters. Maintaining metal clusters intact, which is a great challenge in MS analysis, was achieved in this work by choosing a suitable mass analyzer and carefully optimizing analysis parameters. Electrospray ionization time-of-flight mass spectrometry (ESI-TOF-MS) and electrospray ion trap mass spectrometry (ESI-IT-MS) were applied to characterize the synthesized ligand-protected metal clusters [Au6 Ag2 (C)(L(1) )6 ](BF4 )4 (L(1) =2-diphenylphosphanyl-4-methylpyridine) (1) and [Au8 (L(2) )3 (L(3) )2 ](NO3 )2 (L(2) =bis(2-diphenylphosphinophenyl)ether, L(3) =triphenyl-phosphane) (2). Three kinds of buffer gas (helium, mass: 2; nitrogen, mass: 28; argon, mass: 40) and various radiofrequency (RF) amplitudes (from 70 to 330) were chosen to study the fragmentation rate during the "collision cooling" process in the ion trap analyzer. In the ESI-TOF-MS analysis, metal clusters 1 and 2 were mainly observed as intact clusters, which were Au6 Ag2 (C)(L(1) )6 (BF4 )2 (2+) , Au6 Ag2 (C)(L(1) )6 (BF4 )(3+) , Au6 Ag2 (C)(L(1) )6 (4+) for 1 and Au8 (L(2) )3 (L(3) )2 (2+) for 2. While, in the ESI-IT-MS analysis, only fragments could be found, such as Au6 Ag(C)(L(1) )6 (BF4 )(2+) , Au6 (C)(L(1) )6 (2+) , Au5 Ag(C)(L(1) )4 (2+) , Au6 Ag(C)(L(1) )6 (3+) , Au(L(1) )(+) for 1 and Au8 (L(2) )3 (L(3) )(2+) , Au8 (L(2) )3 (2+) , Au6 (L(2) )3 (2+) for 2. It is obvious that the two kinds of mass analyzers caused different MS behaviors of metal clusters. In the ion trap (IT) mass analyzer, particularly, "collision cooling" was contributing to further dissociation of fragile compounds, in which a higher RF amplitude and a larger mass buffer gas led to more fragmentation. In this work, intact metal clusters were obtained in ESI-TOF-MS, instead of ESI-IT-MS, in which the "collision cooling" process caused more cluster dissociation. It was concluded that the analyzer in ESI-TOF-MS is "softer" than that in ESI-IT-MS for metal clusters. Copyright © 2016 John Wiley & Sons, Ltd.

The Epipolythiodiketopiperazine Gene Cluster in Claviceps purpurea: Dysfunctional Cytochrome P450 Enzyme Prevents Formation of the Previously Unknown Clapurines.

Claviceps purpurea is an important food contaminant and well known for the production of the toxic ergot alkaloids. Apart from that, little is known about its secondary metabolism and not all toxic substances going along with the food contamination with Claviceps are known yet. We explored the metabolite profile of a gene cluster in C. purpurea with a high homology to gene clusters, which are responsible for the formation of epipolythiodiketopiperazine (ETP) toxins in other fungi. By overexpressing the transcription factor, we were able to activate the cluster in the standard C. purpurea strain 20.1. Although all necessary genes for the formation of the characteristic disulfide bridge were expressed in the overexpression mutants, the fungus did not produce any ETPs. Isolation of pathway intermediates showed that the common biosynthetic pathway stops after the first steps. Our results demonstrate that hydroxylation of the diketopiperazine backbone is the critical step during the ETP biosynthesis. Due to a dysfunctional enzyme, the fungus is not able to produce toxic ETPs. Instead, the pathway end-products are new unusual metabolites with a unique nitrogen-sulfur bond. By heterologous expression of the Leptosphaeria maculans cytochrome P450 encoding gene sirC, we were able to identify the end-products of the ETP cluster in C. purpurea. The thioclapurines are so far unknown ETPs, which might contribute to the toxicity of other C. purpurea strains with a potentially intact ETP cluster.

A new type of cancer cluster

Cancer Setting out to isolate circulating tumor cells (CTCs) in the blood of cancer patients, Cima et al. instead noticed that clusters, rather than single cells, abounded. But these clusters were not like CTCs: The cells did not express the epithelial cell marker EpCAM and they did not have the same mutations as the primary tumor. Instead, the clusters matched the genotype and phenotype of endothelial cells and were shed from tumors in mice and in humans as intact clusters, rather than forming by coagulation. In patients, the presence of endothelial cell clusters correlated with early-stage disease and might be a unique indicator of cancer before treatment starts. Sci. Transl. Med. 8 , 345ra89 (2016).