A bidirectional fluorescent two-hybrid system for monitoring protein–protein interactions

Two-hybrid systems have been the cornerstone of research into protein–protein interactions, but these systems typically rely on life/death reporters that put additional selective pressure on the host organism, and potentially lead to false positives. Here we report a bidirectional fluorescence-based bacterial two- hybrid system that enables both the association and dissociation of a given protein–protein interaction to be monitored. The functionality of this system and its compatibility with FACS screening are demon- strated in the forward and reverse direction using known interacting protein-partners and their cyclic peptide inhibitors. The reported fluorescent two-hybrid system may be used in the forward direction for the identification of interacting protein partners, or as a reverse two-hybrid system for the high- throughput identification of protein–protein interaction inhibitors

A cyclic peptide inhibitor of HIF-1 heterodimerization that inhibits hypoxia signaling in cancer cells

Hypoxia inducible factor-1 (HIF-1) is a heterodimeric transcription factor that acts as the master regulator of cellular response to reduced oxygen levels, thus playing a key role in the adaptation, survival and progression of tumors. Here we report cyclo-CLLFVY, identified from a library of 3.2 million cyclic hexapeptides using a genetically encoded high-throughput screening platform, as an inhibitor of the HIF-1α/HIF-1β protein-protein interaction in vitro and in cells. The identified compound inhibits HIF-1 dimerization and transcription activity by binding to the PAS-B domain of HIF-1α, reducing HIF-1-mediated hypoxia response signaling in a variety of cell lines, without affecting the function of the closely related HIF-2 isoform. The reported cyclic peptide demonstrates the utility of our high-throughput screening platform for the identification of protein-protein interaction inhibitors, and forms the starting point for the development of HIF-1 targeted cancer therapeutics

Retired A Stars and Their Companions. III. Comparing the Mass-Period Distributions of Planets Around A-Type Stars and Sun-Like Stars

We present an analysis of ~5 years of Lick Observatory radial velocity measurements targeting a uniform sample of 31 intermediate-mass subgiants (1.5 < M*/Msun < 2.0) with the goal of measuring the occurrence rate of Jovian planets around (evolved) A-type stars and comparing the distributions of their orbital and physical characteristics to those of planets around Sun-like stars. We provide updated orbital solutions incorporating new radial velocity measurements for five known planet-hosting stars in our sample; uncertainties in the fitted parameters are assessed using a Markov Chain Monte Carlo method. The frequency of Jovian planets interior to 3 AU is 26 (+9,-8)%, which is significantly higher than the ~5-10% frequency observed around solar-mass stars. The median detection threshold for our sample includes minimum masses down to {0.2, 0.3, 0.5, 0.6, 1.3} MJup within {0.1, 0.3, 0.6, 1.0, 3.0} AU. To compare the properties of planets around intermediate-mass stars to those around solar-mass stars we synthesize a population of planets based on the parametric relationship dN ~ M^{alpha}P^{beta} dlnM dlnP, the observed planet frequency, and the detection limits we derived. We find that the values of alpha and beta for planets around solar-type stars from Cumming et al. fail to reproduce the observed properties of planets in our sample at the 4 sigma level, even when accounting for the different planet occurrence rates. Thus, the properties of planets around A stars are markedly different than those around Sun-like stars, suggesting that only a small (~ 50%) increase in stellar mass has a large influence on the formation and orbital evolution of planets.Comment: Accepted by the Astrophysical Journal; 15 pages, 15 figure

Evaluation of Critical Quality Attributes of a Pentavalent (A, C, Y, W, X) Meningococcal Conjugate Vaccine for Global Use

Towards achieving the goal of eliminating epidemic outbreaks of meningococcal disease in the African meningitis belt, a pentavalent glycoconjugate vaccine (NmCV-5) has been developed to protect against Neisseria meningitidis serogroups A, C, Y, W and X. MenA and X polysaccharides are conjugated to tetanus toxoid (TT) while MenC, Y and W polysaccharides are conjugated to recombinant cross reactive material 197 (rCRM197), a non-toxic genetic variant of diphtheria toxin. This study describes quality control testing performed by the manufacturer, Serum Institute of India Private Limited (SIIPL), and the independent control laboratory of the U.K. (NIBSC) on seven clinical lots of the vaccine to ensure its potency, purity, safety and consistency of its manufacturing. In addition to monitoring upstream-manufactured components, samples of drug substance, final drug product and stability samples were evaluated. This paper focuses on the comparison of the vaccine’s critical quality attributes and reviews key indicators of its stability and immunogenicity. Comparable results were obtained by the two laboratories demonstrating sufficient levels of polysaccharide O-acetylation, consistency in size of the bulk conjugate molecules, integrity of the conjugated saccharides in the drug substance and drug product, and acceptable endotoxin content in the final drug product. The freeze-dried vaccine in 5-dose vials was stable based on molecular sizing and free saccharide assays. Lot-to-lot manufacturing consistency was also demonstrated in preclinical studies for polysaccharide-specific IgG and complement-dependent serum bactericidal activity for each serogroup. This study demonstrates the high quality and stability of NmCV-5, which is now undergoing Phase 3 clinical trials in Africa and India

A MT-TL1 variant identified by whole exome sequencing in an individual with intellectual disability, epilepsy, and spastic tetraparesis

Funder: The Solve-RD project has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 779257Abstract: The genetic etiology of intellectual disability remains elusive in almost half of all affected individuals. Within the Solve-RD consortium, systematic re-analysis of whole exome sequencing (WES) data from unresolved cases with (syndromic) intellectual disability (n = 1,472 probands) was performed. This re-analysis included variant calling of mitochondrial DNA (mtDNA) variants, although mtDNA is not specifically targeted in WES. We identified a functionally relevant mtDNA variant in MT-TL1 (NC_012920.1:m.3291T > C; NC_012920.1:n.62T > C), at a heteroplasmy level of 22% in whole blood, in a 23-year-old male with severe intellectual disability, epilepsy, episodic headaches with emesis, spastic tetraparesis, brain abnormalities, and feeding difficulties. Targeted validation in blood and urine supported pathogenicity, with heteroplasmy levels of 23% and 58% in index, and 4% and 17% in mother, respectively. Interestingly, not all phenotypic features observed in the index have been previously linked to this MT-TL1 variant, suggesting either broadening of the m.3291T > C-associated phenotype, or presence of a co-occurring disorder. Hence, our case highlights the importance of underappreciated mtDNA variants identifiable from WES data, especially for cases with atypical mitochondrial phenotypes and their relatives in the maternal line

Solving patients with rare diseases through programmatic reanalysis of genome-phenome data.

Funder: EC | EC Seventh Framework Programm | FP7 Health (FP7-HEALTH - Specific Programme "Cooperation": Health); doi: https://doi.org/10.13039/100011272; Grant(s): 305444, 305444Funder: Ministerio de Economía y Competitividad (Ministry of Economy and Competitiveness); doi: https://doi.org/10.13039/501100003329Funder: Generalitat de Catalunya (Government of Catalonia); doi: https://doi.org/10.13039/501100002809Funder: EC | European Regional Development Fund (Europski Fond za Regionalni Razvoj); doi: https://doi.org/10.13039/501100008530Funder: Instituto Nacional de Bioinformática ELIXIR Implementation Studies Centro de Excelencia Severo OchoaFunder: EC | EC Seventh Framework Programm | FP7 Health (FP7-HEALTH - Specific Programme "Cooperation": Health)Reanalysis of inconclusive exome/genome sequencing data increases the diagnosis yield of patients with rare diseases. However, the cost and efforts required for reanalysis prevent its routine implementation in research and clinical environments. The Solve-RD project aims to reveal the molecular causes underlying undiagnosed rare diseases. One of the goals is to implement innovative approaches to reanalyse the exomes and genomes from thousands of well-studied undiagnosed cases. The raw genomic data is submitted to Solve-RD through the RD-Connect Genome-Phenome Analysis Platform (GPAP) together with standardised phenotypic and pedigree data. We have developed a programmatic workflow to reanalyse genome-phenome data. It uses the RD-Connect GPAP's Application Programming Interface (API) and relies on the big-data technologies upon which the system is built. We have applied the workflow to prioritise rare known pathogenic variants from 4411 undiagnosed cases. The queries returned an average of 1.45 variants per case, which first were evaluated in bulk by a panel of disease experts and afterwards specifically by the submitter of each case. A total of 120 index cases (21.2% of prioritised cases, 2.7% of all exome/genome-negative samples) have already been solved, with others being under investigation. The implementation of solutions as the one described here provide the technical framework to enable periodic case-level data re-evaluation in clinical settings, as recommended by the American College of Medical Genetics

Selection of cyclic peptide inhibitors of hypoxia-inducible factor

Hypoxia-inducible factor (HIF) is a heterodimer that is formed from the association of HIF-? and HIF-1? subunits. HIF acts as a potent inducer of genes involved in angiogenesis and neovascularisation. Three different variants of HIF have been identified to date; HIF-1 and HIF-2 are thought to play key roles in tumour blood vessel formation while with HIF-3 is thought to act as a negative regulator of the hypoxic response.In this study a genetic selection methodology is employed that combines a bacterial reverse two-hybrid system with SICLOPPS (split-intein circular ligation of peptides and proteins), a protocol for the construction of libraries of around a hundred million cyclic peptides using split-inteins in vivo. After construction of the appropriate reverse twohybrid systems, several SICLOPPS libraries were screened for inhibitors of HIF-1 and HIF-2 heterodimerisation. After a number of rounds of secondary screening, the activity of the remaining compounds was ranked by using the HIF reverse two-hybrid systems. The most potent compounds were synthesised as Tat-tagged derivatives for further characterisation in a luciferase based mammalian cell assay. The most potent cyclic peptide exhibited IC50s ranging from 26 to 106 ?M in U2OS and MCF7 cell lines

AI readiness in the food industry : An analysis of Nordic food companies

Livsmedelsindustrin står inför stora utmaningar i framtiden, varav en är att erbjuda och producera livsmedel till en växande population på ett hållbart sätt. Ett sätt att övervinna denna utmaning är att implementera AI-teknik som kan användas för att effektivisera och främja hållbar utveckling inom livsmedelsindustrin. Syftet med studien är därför att kartlägga nivån av AI-readiness hos livsmedelsföretag i Norden, det vill säga undersöka hur redo dessa verksamheter är att implementera AI-teknik. Frågeställningen har besvarats genom en digital enkätundersökning baserat på det teoretiska ramverket TOE - Technology, Organization and Environment. Enkäten besvarades av 59 anonyma respondenter. Svaren kategoriserades baserat på det huvudsakliga verksamhetslandet för respondentens företag, den del av industrin företaget främst är verksamt inom och företagets storlek. Det insamlade datat har analyserats med hjälp av Chi2-test som resulterade i insikten att större företag har högre nivå av AI-readiness än små företag. Resultatet indikerade även på tre förbättringsområden för att öka nivån av AI-readiness. Företag bör i större utsträckning utveckla en strategisk plan för implementering av AI-teknik, säkerställa tillräckligt med mänskliga resurser med rätt kompetens och öka kunskapen om externa regleringar bland sina anställda. Den kunskap som studien genererar kan vara relevant för företag som arbetar med implementering av AI-teknik, för företag verksamma inom livsmedelsindustrin som vill öka sin AI-readiness samt för framtida forskning inom liknande områden. The food industry faces major challenges in the future, one of which is to provide and produce food for a growing population in a sustainable way. One way to overcome this challenge is to implement AI-technologies that can be used to promote sustainable development in the food industry. The purpose of this study is therefore to map the level of AI-readiness among food companies in the Nordic region, specifically to investigate how ready these businesses are to implement AI-technology. The question has been answered through a digital survey based on the theoretical framework TOE - Technology, Organization and Environment. The survey was answered by 59 anonymous respondents. The answers were categorized based on the main country of operation of the respondent's company, the part of the industry the company is mainly active in and the size of the company. The collected data has been analyzed using Chi2-tests which resulted in the insight that larger companies have a higher level of AI-readiness than smaller companies. The result also indicated three areas of improvement to increase the level of AI-readiness. Companies should to a greater extent develop a strategic plan for the implementation of AI-technology, ensure sufficient human resources with the right skills and increase the knowledge of external regulations among their employees. The knowledge generated by the study can be relevant for companies working on the implementation of AI-technology, for companies operating in the food industry that want to increase their AI-readiness, and for future research in similar areas

Riskbeteende utifrån prospektteorin : En kvantitativ studie av den svenska skogsindustrins risk- och avkastningssamband

Antagandet om individers rationella beteende har länge varit ett vedertaget koncept inom den ekonomiska utvecklingen. Att anomalier, så som finansiella kriser, har observerats på marknaden har bidragit till att den traditionella ekonomiska synen blivit ifrågasatt. Genom dessa anomalier har forskare erhållit ett starkare intresse för de psykologiska faktorernas inverkan på marknadsaktörer, vilket resulterat i forskningsgrenen beteendeekonomi. Det stora genombrottet för beteendeekonomin kom år 1979 i samband med att Kahneman och Tversky (1979) presenterade prospektteorin. Genom prospektteorins uppkomst ifrågasattes det traditionella ekonomiska synsättet att se på individers beslutstagande under risk. Ett antal studier som sökt stöd för prospektteorin har flyttat fokus från undersökningar av individer till företag, genom att studera risk- och avkastningssambandet, med utgångspunkt i prospektteorins värdefunktion.   Studiens teoretiska grund bygger huvudsakligen på prospektteorin samt den förväntade nyttoteorin. Den förväntade nyttoteorin utgör en del av den traditionella ekonomiska synen, vilken antar att en individ agerar nyttomaximerande och antingen är riskaversiv, risksökande eller riskneutral. Detta i kontrast till prospektteorin, vilken är en del av det beteendeekonomiska området som menar att individer inte alltid handlar nyttomaximerande samt handlar utefter subjektiva preferenser. Studiens syfte uppfylldes genom studerandet av risk- och avkastningssambandet utifrån prospektteorins värdefunktion, för företag inom den svenska skogsindustrin, för att se om stöd för prospektteorin som en förklarande modell för beslutstagande under risk förelåg.   De utformade hypoteserna prövades utifrån data som insamlats genom en kvantitativ datainsamlingsmetod. Avkastningsmåtten som hämtats är den årliga avkastningen på eget samt totalt kapital, för tidsperioden 2006-2013. Datat analyserades genom korstabeller samt rangkorrelationsmatriser.   Resultatet visade, med en signifikansnivå på 1%, att prospektteorin är en fullgod alternativ förklaringsmodell för företagens beslutstagande under risk inom den svenska skogsindustrin. Genom att resultatet genererades med utgångspunkt i prospektteorin, kunde studien bevisa att företagen inom den svenska skogsindustrin fattar beslut under risk olika beroende på var i förhållande till branschens genomsnittsprestation de ligger.  Detta talar för att företagen inom den svenska skogsindustrin som, i förhållande till branschgenomsnittet, överpresterat agerar riskaversivt medan företagen som underpresterat agerar risksökande. Resultatet talar därför mot den förväntade nyttoteorins antaganden

Being a newly graduated nurse during the COVID-19 pandemic : The transition from student to registered nurse - a qualitative literature study

Bakgrund: Nyutexaminerade sjuksköterskor under 2020 och 2021 gjorde entré i sjuksköterskeyrket under en eskalerande pandemi. International Council of Nursing (2021a) flaggade för sjuksköterskornas risk för utbrändhet, posttraumatiskt stressyndrom och andra stressrelaterade sjukdomar. Denna litteraturstudie kan bidra med kunskap om hur situationen upplevdes för nyutexaminerade sjuksköterskor under pandemins exceptionella belastning på sjukvården.  Syfte: Syftet med studien var att sammanställa forskning om nyutexaminerade sjuksköterskors erfarenheter av övergångsfasen från student till sjuksköterska under COVID-19 pandemin.  Metod: Litteraturstudien är baserad på åtta kvalitativa artiklar som hämtats från databaserna CINAHL och PubMed. Samtliga artiklar kvalitetsgranskades enligt Olsson och Sörensens (2011) bedömningsmall. Artiklarna analyserades sedan utifrån Fribergs (2022) femstegsanalys.  Resultat: I resultatet framkommer tre kategorier: ”inte tillåtas vara ny som sjuksköterska under pandemin”, ”en verklighetsfrämmande arbetsmiljö” och ”leva igenom övergångsfasen under exceptionella förhållanden”.  Konklusion: COVID-19 pandemin har genererat i huvudsak känslor av ångest, stress och rädslor på grund av brist på erfarenhet, stöd och dåliga arbetsplatsförhållanden. En god introduktion i form av stöd från kollegor och verksamheten var bidragande faktorer till en underlättad övergång.