Efficacy of an impression disinfectant solution after repeated use: An in vitro study.

Statement of problem: There are very few studies using Benzalkonium Chloride (BAC) as an active disinfection agent for immersion techniques and there are no studies investigating the efficacy of repeated use of a disinfectant solution. Purpose: This study evaluated an impression disinfectant by testing bacterial contamination of disinfectant batches used in a clinical setting after repeated use. Materials and methods: Liquid samples were collected from impression disinfectant solutions used to disinfect dental impressions taken at a university dental clinic. The experimental samples (500 ml from 1 L of solution) were collected from teaching and professional clinics and the in-house commercial processing laboratory and stored at room temperature each day of clinic operation over five weeks. To determine to what extent the disinfectant efficacy of the active product decreased over time, the following tests were carried out: a. Inoculation b. Gram staining technique c. Matrix Assisted Laser Desorption/Ionization Mass spectrometry (MALDI- MS). Microbial growth was monitored and photographed. A culture revival was made from colonies grown on sheep blood agar, to isolate pure colonies incubated for 24 h at 37 °C. Each morphologically distinct type of colony was gram stained and MALDI spectrometry analysis was performed using the VITEK MS (BioMerieux Inc.). Results: Evidence of growth of bacteria was detected in teaching clinics' samples, and no growth from the professional clinic or the commercial laboratory. Conclusions: The study demonstrated that the impression disinfectant solution tested is effective against common oral bacteria, despite some rare species such as Bacillus circulans, Bacillus horneckiae, Bacillus altitudinis/pumilus and Bacillus cereus showing evidence of survival in solutions used for disinfection of impressions. However, in a high use teaching clinic environment its efficacy deteriorated. Though a second level disinfection protocol in the commercial laboratory-maintained impression disinfection.

Immunotherapy for non-small cell lung cancer (NSCLC), as a stand-alone and in combination therapy.

Non-small cell lung cancer (NSCLC) is of major concern for society as it is associated with high mortality and is one of the most commonly occurring of all cancers. Due to the number of mutational variants and general heterogeneity of this type of cancer, treatment using conventional modalities has been challenging. Therefore, it is important to have improved therapeutic treatments like immunotherapy, that can specifically treat the disease while causing minimal damage to healthy tissue and additionally provide systemic immunity. Cancer vaccines are an important element of cancer immunotherapy and have been approved for treatment of a limited number of cancers, including NSCLC. This article highlights scientific evidence for several therapeutic treatment strategies for NSCLC, alone or in combination, which offers new hope for those suffering. Although cancer vaccines have had some success as a monotherapy, their potential in a combination therapy needs to be critically analyzed for future applications.

Conserved bacterial de novo guanine biosynthesis pathway enables microbial survival and colonization in the environmental niche of the urinary tract.

In bacteria, guaA encodes guanosine monophosphate synthetase that confers an ability to biosynthesize guanine nucleotides de novo. This enables bacterial colonization in different environments and, while guaA is widely distributed among Bacteroidetes and Firmicutes, its contribution to the inhabitation of the human microbiome by commensal bacteria is unclear. We studied Streptococcus as a commensal urogenital tract bacterium and opportunistic pathogen, and explored the role of guaA in bacterial survival and colonization of urine. Analysis of guaA-deficient Streptococcus revealed guanine utilization is essential for bacterial colonization of this niche. The genomic location of guaA in other commensals of the human urogenital tract revealed substantial cross-phyla diversity and organizational structures of guaA that are divergent across phyla. Essentiality of guaA for Streptococcus colonization in the urinary tract establishes that purine biosynthesis is a critical element of the ability of this bacterium to survive and colonize in the host as part of the resident human microbiome.

Anti-prion and α-Synuclein Aggregation Inhibitory Sterols from the Sponge Lamellodysidea cf. chlorea.

In a study aimed at identifying new anti-prion compounds we screened a library of 500 Australian marine invertebrate derived extracts using a yeast-based anti-prion assay. This resulted in an extract from the subtropical sponge Lamellodysidea cf. chlorea showing potent anti-prion activity. The bioassay-guided investigation of the sponge extract led to the isolation of three new bioactive polyoxygenated steroids, lamellosterols A-C (1-3). These sterols were all isolated in low yield, and their structures elucidated by extensive NMR and MS data analysis. Lamellosterols A-C displayed potent anti-prion activity against the [PSI+] yeast prion (EC50s of 12.7, 13.8, and 9.8 µM, respectively). Lamellosterol A (1) was further shown to bind to the Parkinson's disease implicated amyloid protein, α-synuclein, and to significantly inhibit its aggregation. Our findings indicate that these polyoxygenated sterol sulfates may be useful compounds to study mechanisms associated with neurodegenerative diseases.

WITHDRAWN: Structural studies of vitrified biological proteins and macromolecules - A review on the microimaging aspects of cryo-electron microscopy.

This article has been withdrawn at the request of the author(s) and/or editor. The Publisher apologizes for any inconvenience this may cause. The full Elsevier Policy on Article Withdrawal can be found at https://www.elsevier.com/about/our-business/policies/article-withdrawal.

Yeast as a Model to Understand Actin-Mediated Cellular Functions in Mammals-Illustrated with Four Actin Cytoskeleton Proteins.

The budding yeast Saccharomyces cerevisiae has an actin cytoskeleton that comprises a set of protein components analogous to those found in the actin cytoskeletons of higher eukaryotes. Furthermore, the actin cytoskeletons of S. cerevisiae and of higher eukaryotes have some similar physiological roles. The genetic tractability of budding yeast and the availability of a stable haploid cell type facilitates the application of molecular genetic approaches to assign functions to the various actin cytoskeleton components. This has provided information that is in general complementary to that provided by studies of the equivalent proteins of higher eukaryotes and hence has enabled a more complete view of the role of these proteins. Several human functional homologues of yeast actin effectors are implicated in diseases. A better understanding of the molecular mechanisms underpinning the functions of these proteins is critical to develop improved therapeutic strategies. In this article we chose as examples four evolutionarily conserved proteins that associate with the actin cytoskeleton: 1) yeast Hof1p/mammalian PSTPIP1, 2) yeast Rvs167p/mammalian BIN1, 3) yeast eEF1A/eEF1A1 and eEF1A2 and 4) yeast Yih1p/mammalian IMPACT. We compare the knowledge on the functions of these actin cytoskeleton-associated proteins that has arisen from studies of their homologues in yeast with information that has been obtained from in vivo studies using live animals or in vitro studies using cultured animal cell lines.

Anti-prion Butenolides and Diphenylpropanones from the Australian Ascidian Polycarpa procera.

A library of 500 Australian marine invertebrate extracts was screened for anti-prion activity using a yeast-based assay, and this resulted in an extract from the ascidian Polycarpa procera showing potent activity. Purification of this extract led to the isolation of six new butenolide metabolites, the procerolides 1-4 and two related diphenylpropanones, the procerones 5 and 6, as the bioactive components. The structures of 1-6 were elucidated from the analysis of 1D/2D NMR and MS data, and their absolute configurations determined from comparison of experimental and computed ECD data. Compounds 1-6 were tested for anti-prion activity in a yeast-based assay, and 1 and 5 displayed potent bioactivity (EC50 of 23 and 29 µM, respectively) comparable to the potently active anti-prion compound guanabenz. The procerolides and procerones are the first anti-prion compounds to be reported from ascidians, indicating that ascidians may be an untapped source of new lead anti-prion compounds.

The regulation of Endosomal Sorting Complex Required for Transport and accessory proteins in multivesicular body sorting and enveloped viral budding - An overview.

ESCRT (Endosomal Sorting Complex Required for Transport) machinery drives different cellular processes such as endosomal sorting, organelle biogenesis, vesicular trafficking, maintenance of plasma membrane integrity, membrane fission during cytokinesis and enveloped virus budding. The normal cycle of assembly and disassembly of some ESCRT complexes at the membrane requires the AAA-ATPase vacuolar protein sorting 4 (Vps4p). A number of ESCRT proteins are hijacked by clinically significant enveloped viruses including Ebola, and Human Immunodeficiency Virus (HIV) to enable enveloped virus budding and Vps4p provides energy for the disassembly/recycling of these ESCRT proteins. Several years ago, the failure of the terminal budding process of HIV following Vps4 protein inhibition was published; although at that time a detailed understanding of the molecular players was missing. However, later it was acknowledged that the ESCRT machinery has a role in enveloped virus budding from cells due to its role in the multivesicular body (MVB) sorting pathway. The MVB sorting pathway facilitates several cellular activities in uninfected cells, such as the down-regulation of signaling through cell surface receptors as well as the process of viral budding from infected host cells. In this review, we focus on summarising the functional organisation of ESCRT proteins at the membrane and the role of ESCRT machinery and Vps4p during MVB sorting and enveloped viral budding.

EGFR and Prion protein promote signaling via FOXO3a-KLF5 resulting in clinical resistance to platinum agents in colorectal cancer.

Epidermal growth factor receptor (EGFR) supports colorectal cancer progression via oncogenic signaling. Anti-EGFR therapy is being investigated as a clinical option for colorectal cancer, and an observed interaction between EGFR and Prion protein has been detected in neuronal cells. We hypothesized that PrPC expression levels may regulate EGFR signaling and that detailed understanding of this signaling pathway may enable identification of resistance mechanisms and new actionable targets in colorectal cancer. We performed molecular pathway analysis following knockdown of PrPC or inhibition of EGFR signaling via gefitinib to identify changes in expression of key signaling proteins that determine cellular sensitivity or resistance to cisplatin. Expression of these proteins was examined in matched primary and metastatic patient samples and was correlated for resistance to therapy and progression of disease. Utilizing three colorectal cancer cell lines, we observed a correlation between high expression of PrPC and resistance to cisplatin. Investigation of molecular signaling in a resistant cell line revealed that PrPC contributed to signaling via colocalization with EGFR, which could be overcome by targeting p38 mitogen-activated protein kinases (p38 MAPK). We revealed that the level of Krüppel-like factor 5 (KLF5), a target downstream of p38 MAPK, was predictive for cell line and patient response to platinum agents. Further, high KLF5 expression was observed in BRAF-mutant colorectal cancer. Our study indicates that the EGFR to KLF5 pathway is predictive of patient progression on platinum-based therapy.

Yeast-based screening of natural product extracts results in the identification of prion inhibitors from a marine sponge.

One of the major medical challenges of the twenty-first century is the treatment of incurable and fatal neurodegenerative disorders caused by misfolded prion proteins. Since the discovery of these diseases a number of studies have been conducted to identify small molecules for their treatment, however to date no curative treatment is available. These studies can be highly expensive and time consuming, but more recent experimental approaches indicate a significant application for yeast prions in these studies. We therefore used yeast prions to optimize previous high-throughput methods for the cheaper, easier and more rapid screening of natural extracts. Through this approach we aimed to identify natural yeast-prion inhibitors that could be useful in the development of novel treatment strategies for neurodegenerative disorders. We screened 500 marine invertebrate extracts from temperate waters in Australia allowing the identification of yeast-prion inhibiting extracts. Through the bioassay-driven chemical investigation of an active Suberites sponge extract, a group of bromotyrosine derivatives were identified as potent yeast-prion inhibitors. This study outlines the importance of natural products and yeast prions as a first-stage screen for the identification of new chemically diverse and bioactive compounds.

Prion protein scrapie and the normal cellular prion protein.

Prions are infectious proteins and over the past few decades, some prions have become renowned for their causative role in several neurodegenerative diseases in animals and humans. Since their discovery, the mechanisms and mode of transmission and molecular structure of prions have begun to be established. There is, however, still much to be elucidated about prion diseases, including the development of potential therapeutic strategies for treatment. The significance of prion disease is discussed here, including the categories of human and animal prion diseases, disease transmission, disease progression and the development of symptoms and potential future strategies for treatment. Furthermore, the structure and function of the normal cellular prion protein (PrP(C)) and its importance in not only in prion disease development, but also in diseases such as cancer and Alzheimer's disease will also be discussed.

Yeast studies reveal moonlighting functions of the ancient actin cytoskeleton.

Classic functions of the actin cytoskeleton include control of cell size and shape and the internal organization of cells. These functions are manifest in cellular processes of fundamental importance throughout biology such as the generation of cell polarity, cell migration, cell adhesion, and cell division. However, studies in the unicellular model eukaryote Saccharomyces cerevisiae (Baker's yeast) are giving insights into other functions in which the actin cytoskeleton plays a critical role. These include endocytosis, control of protein translation, and determination of protein 3-dimensional shape (especially conversion of normal cellular proteins into prions). Here, we present a concise overview of these new "moonlighting" roles for the actin cytoskeleton and how some of these roles might lie at the heart of important molecular switches. This is an exciting time for researchers interested in the actin cytoskeleton. We show here how studies of actin are leading us into many new and exciting realms at the interface of genetics, biochemistry, and cell biology. While many of the pioneering studies have been conducted using yeast, the conservation of the actin cytoskeleton and its component proteins throughout eukaryotes suggests that these new roles for the actin cytoskeleton may not be restricted to yeast cells but rather may reflect new roles for the actin cytoskeleton of all eukaryotes.

Significance of prion and prion-like proteins in cancer development, progression and multi-drug resistance.

Prions are renowned for their role in neurodegenerative diseases in humans and animals. These are manifested as transmissible spongiform encephalopathies (TSEs) that result from the conversion of the normal glycosylphosphatidylinositol (GPI) anchored cellular prion protein (PrP(c)) to a misfolded, aggregated and pathogenic form, prion protein scrapie (PrP(Sc)) via a post-translational process followed by the accumulation of PrP(Sc) within the central nervous system. New research in this area has demonstrated that PrP is over-expressed in a variety of cancers including gastric, pancreatic and breast cancers, affecting the growth and invasiveness of these cancers as well as playing an important role in the acquisition of multi-drug resistant (MDR) gastric cancer. Prion-like doppel protein (Dpl), sharing 25% amino acid sequence homology to PrP and whose function remains elusive, has also been shown to exhibit a high level of expression in a number of cancers including acute myeloid leukemia's, myelodysplastic syndromes, gastric adenocarcinoma, anaplastic meningioma and astrocytomas. Furthermore, the tumour suppressor protein, p53, already known for its involvement in cancer development, has recently been shown to display prion-like tendencies. This review provides an overview of prions and prion-like proteins in mammals discussing their structure, function and role in cell function and disease. Furthermore, current research progress on the role of prion/prion-like proteins in the development, progression, and drug resistance of various cancers will be summarized. Potential implications for future development of new therapeutic treatments targeting prion and prion-like proteins will be discussed.

An ABC pleiotropic drug resistance transporter of Fusarium graminearum with a role in crown and root diseases of wheat.

FgABC1 (FGSG_04580) is predicted to encode a pleiotropic drug resistance class ABC transporter in Fusarium graminearum, a globally important pathogen of wheat. Deletion mutants of FgABC1 showed reduced virulence towards wheat in crown and root infection assays but were unaltered in infectivity on barley. Expression of FgABC1 during head blight and crown rot disease increases during the necrotrophic phases of infection suggestive of a role for FgABC1 in late infection stages in different tissue types. Deletion of FgABC1 also led to increased sensitivity of the fungus to the antifungal compound benalaxyl in culture, but the response to known cereal defence compounds, gramine, 2-benzoxazalinone and tryptamine was unaltered. FgABC1 appears to have a role in protecting the fungus from antifungal compounds and is likely to help combat as yet unidentified wheat defence compounds during disease development.

The F-BAR protein NOSTRIN participates in FGF signal transduction and vascular development.

F-BAR proteins are multivalent adaptors that link plasma membrane and cytoskeleton and coordinate cellular processes such as membrane protrusion and migration. Yet, little is known about the function of F-BAR proteins in vivo. Here we report, that the F-BAR protein NOSTRIN is necessary for proper vascular development in zebrafish and postnatal retinal angiogenesis in mice. The loss of NOSTRIN impacts on the migration of endothelial tip cells and leads to a reduction of tip cell filopodia number and length. NOSTRIN forms a complex with the GTPase Rac1 and its exchange factor Sos1 and overexpression of NOSTRIN in cells induces Rac1 activation. Furthermore, NOSTRIN is required for fibroblast growth factor 2 dependent activation of Rac1 in primary endothelial cells and the angiogenic response to fibroblast growth factor 2 in the in vivo matrigel plug assay. We propose a novel regulatory circuit, in which NOSTRIN assembles a signalling complex containing FGFR1, Rac1 and Sos1 thereby facilitating the activation of Rac1 in endothelial cells during developmental angiogenesis.

Harnessing technology to improve formative assessment of student conceptions in STEM: forging a national network.

Concept inventories, consisting of multiple-choice questions designed around common student misconceptions, are designed to reveal student thinking. However, students often have complex, heterogeneous ideas about scientific concepts. Constructed-response assessments, in which students must create their own answer, may better reveal students' thinking, but are time- and resource-intensive to evaluate. This report describes the initial meeting of a National Science Foundation-funded cross-institutional collaboration of interdisciplinary science, technology, engineering, and mathematics (STEM) education researchers interested in exploring the use of automated text analysis to evaluate constructed-response assessments. Participants at the meeting shared existing work on lexical analysis and concept inventories, participated in technology demonstrations and workshops, and discussed research goals. We are seeking interested collaborators to join our research community.

Verprolin: a cool set of actin-binding sites and some very HOT prolines.

Spatiotemporal organisation of eukaryotic cells is established and maintained by the cytoskeleton, a highly dynamic and complex network of structural and signalling proteins. Many components of the cytoskeleton are functionally and structurally conserved between humans and yeast. Among these are verprolin (Vrp1p) in yeast and its human ortholog Wiskott-Aldrich syndrome protein (WASP)-interacting protein (WIP). Much of our understanding of the function of these proteins has come from genetic analysis in yeast. Verprolin-deficient yeast cells exhibit defects in cytokinesis, endocytosis, and actin cytoskeleton polarisation. Verprolin binds actin, the yeast ortholog of human WASP (Las17p or Bee1p), and the yeast ortholog of human PSTPIP1 (Hof1p or Cyk2p). We propose that verprolin acts as a chaperone that by transient bimolecular interactions maintains the proper function of its partners. Verprolin-related proteins and partners are implicated in cancer, immunodeficiency, and neurodegeneration. Therefore, elucidating how verprolin functions will have major impacts in cell biology and medicine.

Three-dimensional structure of AAA ATPase Vps4: advancing structural insights into the mechanisms of endosomal sorting and enveloped virus budding.

Vps4 is a AAA ATPase that mediates endosomal membrane protein sorting. It is also a host factor hijacked by a diverse set of clinically important viruses, including HIV and Ebola, to facilitate viral budding. Here we present the three-dimensional structure of the hydrolysis-defective Vps4p(E233Q) mutant. Single-particle analysis, multiangle laser light scattering, and the docking of independently determined atomic models of Vps4 monomers reveal a complex with C6 point symmetry, distinguishing between a range of previously suggested oligomeric states (8-14 subunits). The 3D reconstruction also reveals a tail-to-tail subunit organization between the two rings of the complex and identifies the location of domains critical to complex assembly and interaction with partner proteins. Our refined Vps4 structure is better supported by independent lines of evidence than those previously proposed, and provides insights into the mechanism of endosomal membrane protein sorting and viral envelope budding.

Las17p-Vrp1p but not Las17p-Arp2/3 interaction is important for actin patch polarization in yeast.

The actin cytoskeleton plays a central role in many important cellular processes such as cell polarization, cell division and endocytosis. The dynamic changes to the actin cytoskeleton that accompany these processes are regulated by actin-associated proteins Wiskott-Aldrich Syndrome Protein (WASP) (known as Las17p in yeast) and WASP-Interacting Protein (WIP) (known as Vrp1p in yeast). Both yeast and human WASP bind to and stimulate the Arp2/3 complex which in turn nucleates assembly of actin monomers into filaments at polarized sites at the cortex. WASP-WIP interaction in yeast and humans are important for Arp2/3 complex stimulation in vitro. It has been proposed that these interactions are also important for polarized actin assembly in vivo. However, the redundancy of actin-associated proteins has made it difficult to test this hypothesis. We have identified two point mutations (L80T and H94L) in yeast WASP that in combination abolish WASP-WIP interaction in yeast. We also identify an N-terminal fragment of Las17p (N-Las17p1-368) able to interact with Vrp1p but not Arp2/3. Using these mutant and truncated forms of yeast WASP we provide novel evidence that WASP interaction with WIP is more important than interaction with Arp2/3 for polarized actin assembly and endocytosis in yeast.