Structure-activity relationships of pyrazole-4-carbodithioates as antibacterials against methicillin–resistant Staphylococcus aureus

Methicillin-resistant Staphylococcus aureus (MRSA) is a major cause of serious hospital-acquired infections and is responsible for significant morbidity and mortality in residential care facilities. New agents against MRSA are needed to combat rising resistance to current antibiotics. We recently reported 5-hydroxy-3-methyl-1-phenyl-1H-pyrazole-4-carbodithioate (HMPC) as a new bacteriostatic agent against MRSA that appears to act via a novel mechanism. Here, twenty nine analogs of HMPC were synthesized, their anti-MRSA structure-activity relationships evaluated and selectivity versus human HKC-8 cells determined. Minimum inhibitory concentrations (MIC) ranged from 0.5 to 64?μg/mL and up to 16-fold selectivity was achieved. The 4-carbodithioate function was found to be essential for activity but non-specific reactivity was ruled out as a contributor to antibacterial action. The study supports further work aimed at elucidating the molecular targets of this interesting new class of anti-MRSA agents.     

The incC korB region of RK2 repositions a mini-RK2 replicon in Escherichia coli

Analysis by fluorescence microscopy has established that plasmid RK2 in Escherichia coli and other gram-negative bacteria is present as discrete clusters that are located inside the nucleoid at the mid- or quarter-cell positions. A mini-RK2 replicon containing an array of tetO repeats was visualized in E. coli cells that express a TetR-EYFP fusion protein. Unlike intact RK2, the RK2 mini-replicon (pCV1) was localized as a cluster at the cell poles outside of the nucleoid. Insertion of the O(B1)incC korB partitioning (par) region of RK2 into pCV1 resulted in a shift of the mini-replicon to within the nucleoid region at the mid- and quarter-cell positions. Despite the repositioning of the mini-RK2 replicon to the cellular positions where intact RK2 is normally located, the insertion of the intact O(B1) incC korB region did not significantly stabilize the mini-RK2 plasmid during cell growth. Deletions within the O(B1)incC or the korB region resulted in a failure of this par region to move pCV1 out of its polar position. The insertion of the par system of plasmid F into pCV1 resulted in a similar shift in the location of pCV1 to the nucleoid region. Unlike O(B1)incC korB, the insertion of the RK2 parABC resolvase system into pCV1 did not affect the polar positioning of pCV1. This effect of O(B1)incC korB on the location of pCV1 provides additional evidence for a partitioning role of this region of plasmid RK2. However, the failure of this region to significantly increase the stability of the mini-RK2 plasmid indicates that the localization of the plasmid to the mid- and quarter cell positions in E. coli is not in itself sufficient for the stable maintenance of plasmid RK2.     

Insights from modeling studies on how climate change affects invasive alien species geography

Climate change and biological invasions are threatening biodiversity and ecosystem services worldwide. It has now been widely acknowledged that climate change will affect biological invasions. A large number of studies have investigated predicted shifts and other changes in the geographic ranges of invasive alien species related to climate change using modeling approaches. Yet these studies have provided contradictory evidence, and no consensus has been reached. We conducted a systematic review of 423 modeling case studies included in 71 publications that have examined the predicted effects of climate change on those species. We differentiate the approaches used in these studies and synthesize their main results. Our results reaffirm the major role of climate change as a driver of invasive alien species distribution in the future. We found biases in the literature both regarding the taxa, toward plants and invertebrates, and the areas of the planet investigated. Despite these biases, we found for the plants and vertebrates studied that climate change will more frequently contribute to a decrease in species range size than an increase in the overall area occupied. This is largely due to oceans preventing terrestrial invaders from spreading poleward. In contrast, we found that the ranges of invertebrates and pathogens studied are more likely to increase following climate change. An important caveat to these findings is that researchers have rarely considered the effects of climate change on transport, introduction success, or the resulting impacts. We recommend closing these research gaps, and propose additional avenues for future investigations, as well as opportunities and challenges for managing invasions under climate change.     

Blood-Based Protein Biomarker Panel for the Detection of Colorectal Cancer

Background

The majority of colorectal cancer (CRC) cases are preventable by early detection and removal of precancerous polyps. Even though CRC is the second most common internal cancer in Australia, only 30 per cent of the population considered to have risk factors participate in stool-based test screening programs. Evidence indicates a robust, blood-based, diagnostic assay would increase screening compliance. A number of potential diagnostic blood-based protein biomarkers for CRC have been reported, but all lack sensitivity or specificity for use as a stand-alone diagnostic. The aim of this study was to identify and validate a panel of protein-based biomarkers in independent cohorts that could be translated to a reliable, non-invasive blood-based screening test.

Principal Findings

In two independent cohorts (n = 145 and n = 197), we evaluated seven single biomarkers in serum of CRC patients and age/gender matched controls that showed a significant difference between controls and CRC, but individually lack the sensitivity for diagnostic application. Using logistic regression strategies, we identified a panel of three biomarkers that discriminated between controls and CRC with 73% sensitivity at 95% specificity, when applied to either of the two cohorts. This panel comprised of Insulin like growth factor binding protein 2 (IGFBP2), Dickkopf-3 (DKK3), and Pyruvate kinase M2(PKM2).

Conclusions

Due to the heterogeneous nature of CRC, a single biomarker is unlikely to have sufficient sensitivity or specificity for use as a stand-alone diagnostic screening test and a panel of markers may be more effective. We have identified a 3 biomarker panel that has higher sensitivity and specificity for early stage (Stage I and -II) disease than the faecal occult blood test, raising the possibility for its use as a non-invasive blood diagnostic or screening test.     

Network Analysis of Metabolite GWAS Hits: Implication of CPS1 and the Urea Cycle in Weight Maintenance

Background and Scope

Weight loss success is dependent on the ability to refrain from regaining the lost weight in time. This feature was shown to be largely variable among individuals, and these differences, with their underlying molecular processes, are diverse and not completely elucidated. Altered plasma metabolites concentration could partly explain weight loss maintenance mechanisms. In the present work, a systems biology approach has been applied to investigate the potential mechanisms involved in weight loss maintenance within the Diogenes weight-loss intervention study.

Methods and Results

A genome wide association study identified SNPs associated with plasma glycine levels within the CPS1 (Carbamoyl-Phosphate Synthase 1) gene (rs10206976, p-value = 4.709e-11 and rs12613336, p-value = 1.368e-08). Furthermore, gene expression in the adipose tissue showed that CPS1 expression levels were associated with successful weight maintenance and with several SNPs within CPS1 (cis-eQTL). In order to contextualize these results, a gene-metabolite interaction network of CPS1 and glycine has been built and analyzed, showing functional enrichment in genes involved in lipid metabolism and one carbon pool by folate pathways.

Conclusions

CPS1 is the rate-limiting enzyme for the urea cycle, catalyzing carbamoyl phosphate from ammonia and bicarbonate in the mitochondria. Glycine and CPS1 are connected through the one-carbon pool by the folate pathway and the urea cycle. Furthermore, glycine could be linked to metabolic health and insulin sensitivity through the betaine osmolyte. These considerations, and the results from the present study, highlight a possible role of CPS1 and related pathways in weight loss maintenance, suggesting that it might be partly genetically determined in humans.     

A novel mechanism of G protein-coupled receptor functional selectivity. Muscarinic partial agonist McN-A-343 as a bitopic orthosteric/allosteric ligand

Many G protein-coupled receptors (GPCRs) possess allosteric binding sites distinct from the orthosteric site utilized by their cognate ligands, but most GPCR allosteric modulators reported to date lack signaling efficacy in their own right. McN-A-343 (4-(N-(3-chlorophenyl)carbamoyloxy)-2-butynyltrimethylammonium chloride) is a functionally selective muscarinic acetylcholine receptor (mAChR) partial agonist that can also interact allosterically at the M(2) mAChR. We hypothesized that this molecule simultaneously utilizes both an allosteric and the orthosteric site on the M(2) mAChR to mediate these effects. By synthesizing progressively truncated McN-A-343 derivatives, we identified two, which minimally contain 3-chlorophenylcarbamate, as pure allosteric modulators. These compounds were positive modulators of the orthosteric antagonist N-[(3)H]methylscopolamine, but in functional assays of M(2) mAChR-mediated ERK1/2 phosphorylation and guanosine 5'-3-O-([(35)S]thio)triphosphate binding, they were negative modulators of agonist efficacy. This negative allosteric effect was diminished upon mutation of Y177A in the second extracellular (E2) loop of the M(2) mAChR that is known to reduce prototypical allosteric modulator potency. Our results are consistent with McN-A-343 being a bitopic orthosteric/allosteric ligand with the allosteric moiety engendering partial agonism and functional selectivity. This finding suggests a novel and largely unappreciated mechanism of "directed efficacy" whereby functional selectivity may be engendered in a GPCR by utilizing an allosteric ligand to direct the signaling of an orthosteric ligand encoded within the same molecule.     

Mechanism of GABAB receptor-induced BDNF secretion and promotion of GABAA receptor membrane expression

Recent studies have shown that GABA(B) receptors play more than a classical inhibitory role and can function as an important synaptic maturation signal early in life. In a previous study, we reported that GABA(B) receptor activation triggers secretion of brain-derived neurotrophic factor (BDNF) and promotes the functional maturation of GABAergic synapses in the developing rat hippocampus. To identify the signalling pathway linking GABA(B) receptor activation to BDNF secretion in these cells, we have now used the phosphorylated form of the cAMP response element-binding protein as a biological sensor for endogenous BDNF release. In the present study, we show that GABA(B) receptor-induced secretion of BDNF relies on the activation of phospholipase C, followed by the formation of diacylglycerol, activation of protein kinase C, and the opening of L-type voltage-dependent Ca(2+) channels. We further show that once released by GABA(B) receptor activation, BDNF increases the membrane expression of β(2/3) -containing GABA(A) receptors in neuronal cultures. These results reveal a novel function of GABA(B) receptors in regulating the expression of GABA(A) receptor through BDNF-tropomyosin-related kinase B receptor dependent signalling pathway.