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Genetically identical cells in the same stressful condition die at different times. The origin of this stochasticity is unclear; it may arise from different initial conditions that affect the time of demise, or from a stochastic damage accumulation mechanism that erases the initial conditions and instead amplifies noise to generate different lifespans. To address this requires measuring damage dynamics in individual cells over the lifespan, but this has rarely been achieved. Here, we used a microfluidic device to measure membrane damage in 648 carbon-starvedE. colicells at high temporal resolution. We find that initial conditions of damage, size or cell-cycle phase do not explain most of the lifespan variation. Instead, the data points to a stochastic mechanism in which noise is amplified by a rising production of damage that saturates its own removal. Surprisingly, the relative variation in damage drops with age: cells become more similar to each other in terms of relative damage, indicating increasing determinism with age. Thus, chance erases initial conditions and then gives way to increasingly deterministic dynamics that dominate the lifespan distribution.

Surveillance screening at scale to identify people infected by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) prior to extensive transmission is key to bringing an end to the coronavirus disease 2019 (COVID-19) pandemic, even though vaccinations have already begun. Here we describe Corona Detective, a sensitive and rapid molecular test to detect the virus, based on loop-mediated isothermal amplification, which could be applied anywhere at low cost. Critically, the method uses freeze-dried reagents, readily shipped without cold-chain dependence. The reaction detects the viral nucleocapsid gene through a sequence-specific quenched-fluorescence readout, which avoids false positives and also allows multiplex detection with an internal control cellular RNA. Corona Detective can be used in 8-tube strips to be read with a simple open-design fluorescence detector. Other methods to use and produce Corona Detective locally in a variety of formats are possible and already openly shared. Detection specificity is ensured through inclusion of positive and negative control reactions to run in parallel with the diagnostic reactions. A simple user protocol, including sample preparation, and a bioinformatics pipeline to ensure that viral variants will still be detectable with SARS-CoV-2 primer sets complete the method. Through rapid production and distribution of Corona Detective reactions, quite inexpensive at scale, daily or weekly surveillance testing of large populations, without waiting for symptoms to develop, is anticipated, in combination with vaccination campaigns, to finally control this pandemic.

Adult acute lymphoblastic leukemia/lymphoma (ALL/LBL) is a rare and heterogeneous malignancy characterized by uncontrolled proliferation of B or T cell precursor cells. Here, we retrospectively analyzed the outcome of early autologous stem cell transplantation in standard-risk patients in first complete remission (n=24) and of allogeneic transplantation in high and highest risk, and relapsed/refractory patients (n=35). The 10-year overall survival after autologous transplantation was 45%. The 10-year overall survival after allogeneic transplantation was 58%. The cumulative incidence of relapse was 29% after allogeneic and 67% after autologous transplantation. The cumulative incidence of non-relapse mortality was 0% after autologous and 12% after allogeneic transplantation. This retrospective single center analysis in a limited number of standard-risk patients clearly demonstrates that early autologous transplantation in first complete remission leads to an acceptable long-term outcome with a short overall treatment duration of less than 6 months compared with more than 2 years with conventional chemotherapy. More sensitive and standardized methods to detect minimal residual disease (MRD) will further help to identify those patients more accurately who are most likely to benefit from such a short and intensive treatment strategy (i.e., MRD negative standard-risk patients) or those who require early targeted therapy (e.g., blinatumomab) in case of MRD positivity. Early allogeneic transplantation results in long-term survival/cure in nearly two-thirds of all high and highest risk, and relapsed/refractory patients.

In this study we analyzed gender differences in the clinical presentation of patients with molecular confirmed influenza A. Additionally, we tried to identify predictors of influenza-associated mortality.In this prospective observational multi-center-study we included all influenza-positive patients ≥ 18 years who were hospitalized and treated on flu-isolation-wards in three hospitals in Vienna during the 2018/19 influenza season. Diagnoses were made via Cobas490 Patients (48.8% female) tested positive for influenza A. Female patients were older (median age 76 years vs. 70 years, p 0.001). Male patients had a higher rate of chronic liver disease in history (8.8% vs. 2.9%, p = 0.006), myositis (11.7% vs. 3.1%, p 0.001) and ICU admissions (9.6% vs. 4.6%, p = 0.03). The in-hospital mortality rate was 4.3% and increased to 9.5% during the 90-day follow-up period. Female patients 75 years had a significantly higher in-hospital mortality rate than ≤ 75-year-old females (9.2% vs. 1.7%, p = 0.019). This effect was not observed in male patients (5.4% vs. 1.9%, p = ns). Age 75 years (OR 5.49, 95% CI 1.10-27.43), acute heart failure (OR 3.56, 95% CI 1.03-12.05) and ICU admission (OR 6.1, 95% CI 0.98-37.91) were predictors for in-hospital mortality for female patients, while any malignancy (OR 9.4, 95% CI 1.90-46.54) and ICU admission (OR 7.05, 95% CI 1.44-34.55) were predictors in male patients.Gender is associated with differences in clinical presentation and complications of influenza A virus infection. Women with acute heart failure or aged 75 years have an increased risk of influenza associated in-hospital mortality, while ICU admission and any malignancy are predictors for male patients. Mortality rates in patients 75 years are 5-10 times higher compared to their non-hospitalized influenza-negative Austrian counterparts.

Bacterial cells have characteristic spatial and temporal scales. For instance, Escherichia coli, the typical rod-shaped bacteria, always maintains a relatively constant cell width and cell division time. However, whether the external physical perturbation of cell width has an impact on cell division time remains largely unexplored. In this work, we developed two microchannel chips, namely straight channels and ‘necked’ channels, to precisely regulate the width of E. coli cells and to investigate the correlation between cell width and division time of the cells. Our results show that, in the straight channels, the wide cells divide much slower than narrow cells. In the ‘necked’ channels, the cell division is remarkably promoted compared to that in straight channels with the same width. Besides, fluorescence time-lapse microscopy imaging of FtsZ dynamics shows that the cell pre-constriction time is more sensitive to cell width perturbation than cell constriction time. Finally, we revealed a significant anticorrelation between the death rate and the division rate of cell populations with different widths. Our work provides new insights into the correlation between the geometrical property and division time of E. coli cells and sheds new light on the future study of spatial–temporal correlation in cell physiology.

As the second year of the COVID-19 pandemic begins, it remains clear that a massive increase in the ability to test for SARS-CoV-2 infections in a myriad of settings is critical to controlling the pandemic and to preparing for future outbreaks. The current gold standard for molecular diagnostics is the polymerase chain reaction (PCR), but the extraordinary and unmet demand for testing in a variety of environments means that both complementary and supplementary testing solutions are still needed. This review highlights the role that loop-mediated isothermal amplification (LAMP) has had in filling this global testing need, providing a faster and easier means of testing, and what it can do for future applications, pathogens, and the preparation for future outbreaks. This review describes the current state of the art for research of LAMP-based SARS-CoV-2 testing, as well as its implications for other pathogens and testing. The authors represent the global LAMP (gLAMP) Consortium, an international research collective, which has regularly met to share their experiences on LAMP deployment and best practices; sections are devoted to all aspects of LAMP testing, including preanalytic sample processing, target amplification, and amplicon detection, then the hardware and software required for deployment are discussed, and finally, a summary of the current regulatory landscape is provided. Included as well are a series of first-person accounts of LAMP method development and deployment. The final discussion section provides the reader with a distillation of the most validated testing methods and their paths to implementation. This review also aims to provide practical information and insight for a range of audiences: for a research audience, to help accelerate research through sharing of best practices; for an implementation audience, to help get testing up and running quickly; and for a public health, clinical, and policy audience, to help convey the breadth of the effect that LAMP methods have to offer.

Evolution is often an obstacle to the engineering of stable biological systems due to the selection of mutations inactivating costly gene circuits. Gene overlaps induce important constraints on sequences and their evolution. We show that these constraints can be harnessed to increase the stability of costly genes by purging loss-of-function mutations. We combine computational and synthetic biology approaches to rationally design an overlapping reading frame expressing an essential gene within an existing gene to protect. Our algorithm succeeded in creating overlapping reading frames in 80% of E. coli genes. Experimentally, scoring mutations in both genes of such overlapping construct, we found that a significant fraction of mutations impacting the gene to protect have a deleterious effect on the essential gene. Such an overlap thus protects a costly gene from removal by natural selection by associating the benefit of this removal with a larger or even lethal cost. In our synthetic constructs, the overlap converts many of the possible mutants into evolutionary dead-ends, reducing the evolutionary potential of the system and thus increasing its stability over time., Author summary Genomes are translated by triplets of nucleotides on two different strands, allowing for six different reading frames. This permits the existence of gene overlaps, often observed in microbial genomes, where two different proteins are encoded on the same piece of DNA, but in different reading frames. Gene overlaps are classically considered an obstacle for both evolution and genetic engineering, as mutations in overlapping regions likely have pleitrotropic effects on several genes. In 2013, we identified specific evolutionary scenarios where the decrease in evolutionary potential caused by gene overlaps could instead be advantageous and selected for. In this work, we demonstrate the use of gene overlaps in another context where reducing evolutionary potential can be useful: preventing evolution from inactivating synthetic circuits. We show that gene overlaps can be engineered to increase the evolutionary stability of genes that are costly to their hosts, by entangling these costly genes with essential genes.

In 1850, an Imperial firmän was circulated by the Ottoman authorities regarding the Protestant religion. It stated ' . [l]et, then, a respectable and trustworthy person, acceptable to and chosen by themselves, from among their own number, be appointed, with the title of "Agent of the Protestants," who shall be attached to the department of the Minister of Police,' and thus serve as the official representative for the Protestant community to the Porte. This order was pronounced during the Tan;imät, an important period of reform when the Ottoman goverment sought to reconfigure its relationship with its diverse subjects (Hanssen, Philipp and Weber 2002:6- 21; Hanssen 2002:49-74; Makdisi 2008:184-186). This specific decree was a response to a series of events, whereby different groups of individuals, around the Empire, appropriated Euro-American theology and church structuring to their own social and religious practices. Through this and subsequent pronouncements, the Ottoman government recognized and incorporated the different pockets of Protestantism that emerged throughout the Empire and integrated this new religious identity into official structures.

International audience; The purpose of biomedicine is to serve society, yet its hierarchical and closed structure excludes many citizens from the process of innovation. We propose a collection of reforms to better integrate citizens within the research community, reimagining biomedicine as more participatory, inclusive, and responsive to societal needs.

Whole-cell screening for Mycobacterium tuberculosis (Mtb) inhibitors is complicated by the pathogen’s slow growth and biocontainment requirements. Here we present a synthetic biology framework for assaying Mtb drug targets in engineered E. coli. We construct Target Essential Surrogate E. coli (TESEC) in which an essential metabolic enzyme is deleted and replaced with an Mtb-derived functional analog, linking bacterial growth to the activity of the target enzyme. High throughput screening of a TESEC model for Mtb alanine racemase (ALR) revealed benazepril as a targeted inhibitor. In vitro biochemical assays indicated a noncompetitive mechanism unlike that of clinical ALR inhibitors. This is the first report of an antimicrobial activity in an approved Angiotensin Converting Enzyme (ACE) inhibitor and may explain clinical data associating use of ACE inhibitors with reduced Mtb infection risk. We establish the scalability of TESEC for drug discovery by characterizing TESEC strains for four additional targets.SIGNIFICANCE STATEMENTThe challenge of discovering new antibiotics is both scientific and economic. No simple test can determine if a given molecule will be safe and effective in real human patients. Many drug candidates must therefore be advanced for each new antibiotic that reaches the market - a risky and expensive process.In this work we use synthetic biology to engineer the common laboratory model bacterium E. coli as a tool for early stage antibiotic discovery. As a proof of concept we expressed a known tuberculosis drug target and found a novel inhibitor: benazepril.Many other drug targets could be screened similarly using the system that we describe. Because E. coli can be grown safely and cheaply, this approach may help to reduce costs and make drug discovery more accessible.

The coronavirus disease 2019 (COVID-19) pandemic has highlighted bottlenecks in large-scale, frequent testing of populations for infections. Polymerase chain reaction (PCR)-based diagnostic tests are expensive, reliant on centralized labs, can take days to deliver results, and are prone to backlogs and supply shortages. Antigen tests that bind and detect the surface proteins of a virus are rapid and scalable but suffer from high false negative rates. To address this problem, an inexpensive, simple, and robust 60-minute do-it-yourself (DIY) workflow to detect viral RNA from nasal swabs or saliva with high sensitivity (0.1 to 2 viral particles/μL) and specificity (&gt, 97% true negative rate) utilizing reverse transcription loop-mediated isothermal amplification (RT-LAMP) was developed. ALERT (Accessible LAMP-Enabled Rapid Test) incorporates the following features: (1) increased shelf-life and ambient temperature storage, compared to liquid reaction mixes, by using wax layers to isolate enzymes from other reagents, (2) improved specificity compared to other LAMP end-point reporting methods, by using sequence-specific QUASR (quenching of unincorporated amplification signal reporters), (3) increased sensitivity, compared to methods without purification through use of a magnetic wand to enable pipette-free concentration of sample RNA and cell debris removal, (4) quality control with a nasopharyngeal-specific mRNA target, and (5) co-detection of other respiratory viruses, such as influenza B, by multiplexing QUASR-modified RT-LAMP primer sets. The flexible nature of the ALERT workflow allows easy, at-home and point-of-care testing for individuals and higher-throughput processing for labs and hospitals. With minimal effort, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific primer sets can be swapped out for other targets to repurpose ALERT to detect other viruses, microorganisms, or nucleic acid-based markers.

Most mental health questionnaires only probe behavior and emotion over one domain (e.g., frequency) by using an adverbial phrase such as 'How often...?'. This single-domain approach to behavior sampling may limit the scope of psychiatric evidence that these instruments can capture. We investigated whether adding domains such as intensity, duration, ability and context to mental health survey instruments garner responses that better characterize or measure relevant symptoms or behaviors, testing its association with relevant outcomes. We tested whether our multidomain instrument could be associated with hospitalization, self-harm, education, and disability. 1,504 participants took an augmented version of the SWAN questionnaire (‘SWAN+’), where 4 domains were added to the ability domain already present (frequency, duration, intensity, and context). First, we used structural equation models and the tri-factor modelling to analyze if the latent construct of domains was independently associated with the outcomes. Our results suggest that adding domains such as frequency and context could help characterize disability beyond the domain originally probed by the SWAN questionnaire. Different domains had different predictive effects across outcomes measured in the study. We also complemented results from the tri-factor modelling with a more classical approach, by performing regressions on the summed scores on each domain and the entire scale, adjusted by age and gender. Second, analyses were performed to identify the best predictor domain within each question to predict disability. Significant results were found for the ability and context domains on practically all questions (p

Citizen science has expanded rapidly over the past decades. Yet, defining citizen science and its boundaries remained a challenge, and this is reflected in the literature—for example in the proliferation of typologies and definitions. There is a need for identifying areas of agreement and disagreement within the citizen science practitioners community on what should be considered as citizen science activity. This paper describes the development and results of a survey that examined this issue, through the use of vignettes—short case descriptions that describe an activity, while asking the respondents to rate the activity on a scale from ‘not citizen science’ (0%) to ‘citizen science’ (100%). The survey included 50 vignettes, of which five were developed as clear cases of not-citizen science activities, five as widely accepted citizen science activities and the others addressing 10 factors and 61 sub-factors that can lead to controversy about an activity. The survey has attracted 333 respondents, who provided over 5100 ratings. The analysis demonstrates the plurality of understanding of what citizen science is and calls for an open understanding of what activities are included in the field.

Background: SARS-CoV-2 belongs to a subgroup of coronaviruses rampant in bats for centuries. It has caused the COVID-19 pandemic. Most patients recover, but a minority of severe cases experience acute respiratory distress or an inflammatory storm devastating many organs that can lead to patient death. The spread of SARS-CoV-2 has been facilitated by the increasing intensity of air travel, urban congestion and human contact during the last half century. Until therapies and vaccines are available, tests for virus and exposure, confinement measures and physical distancing have helped curb the pandemic. Vision: The COVID-19 pandemic calls for safeguards and remediation measures through a systemic response. A myriad of self-organizing initiatives by scientists and citizens is developing an advanced collective intelligence response to the coronavirus crisis. Their integration forms Olympiads of Solidarity and Health. Their ability to optimize our response to COVID-19 could serve as a model to trigger a global metamorphosis of our societies with far-reaching consequences for attacking fundamental challenges facing humanity in the 21st century. Mission: For COVID-19 and these other challenges, there is no alternative but action. Meeting in Paris in 2003, we set out to "rethink research to understand life and improve health." We have now formed an international coalition of academia and industry ecosystems taking a systems medicine approach to understanding COVID-19 by thoroughly characterizing viruses, patients and populations during the pandemic, using openly shared tools. All results will be publicly available with no initial claims for intellectual property rights. This World Alliance for Health and Wellbeing will catalyze the creation of medical and health products such as diagnostic tests, drugs and vaccines that become common goods accessible to all, while seeking further alliances with civil society to bridge with socio-ecological and technological approaches that characterise urban systems, for a collective response to future health emergencies. Funding: This work was supported during its preparation phases by the State Key Laboratory of Medical Genomics, Overseas Expertise Introduction Project for Discipline Innovation (111 Project, B17029) to SJC, LH, CA; European Union grants CASyM (FP7-n°305033) to CA, RB, JR, DR, DS, JT, OW; eTRIKS (IMI-1-n°115446) to CA, RB, YG; PREPARE (FP7n°602525) to CA, PO, SV; and initial funds in support of the deep phenotyping clinical study from philanthropic donors, Gilead, Merck and Novartis to JG, JH, LH; the Ricerca Corrente Program by the Italian Ministry of Health to AC, CPr, GS, AU, VV; the National Megaprojects of China for Infectious Diseases (2017ZX10103009-001, 2018ZX10305409-001-005), Double Firstclass projects from the Ministry of Education to the National Research Center for Translational Medicine Shanghai to SJC; Shanghai Municipal Science and Technology Major Project (2017SHZDZX01) to LJ. Conflict of Interest: Rudi Balling reports being a founder/shareholder of ITTM S.A. and Megeno S.A., outside the submitted work, he is co-inventor and holder of two issued patents; James Heath reports personal fees and other from PACT Pharma, outside the submitted work; Hiroaki Kitano reports grants from Minister of Health, Welfare, and Labor of Japanese Government, grants from Okinawa Prefecture Government, outside the submitted work; Doron Lancet reports personal fees from Weizmann Institute of Science and LifeMap Science Inc., outside the submitted work; Catherine Larue reports personal fees from Genfit, outside the submitted work; Peter Openshaw reports personal fees for consultancy work with Janssen, JJ Susanna Palkonen reports grants from Aimmune, Astra Zeneca, Boehringer Ingelheim, Chiesi, DBV Technologies, GSK, Leo Pharma, Novartis, Pfizer, Regeneron Pharmaceuticals, Sanofi Genzyme, outside the submitted work, is employed by the EFA who receives unrestricted grants from corporate entities as above, represents EFA in the following advisory groups of these entities: Astra Zeneca COPD Advisory Committee, GSK Health Advisory Board and Novartis European Patient Advisory Group, receives no personal gain, honoraria, when applicable expenses are payed to her organization, is in a voluntary function the Chair of the Patient Access Partnership PACT who receives unrestricted grants from EFPIA, MedTech Europe, EUCOPE and Medicines for Europe for core operations, and project grants from individual companies, outside the submitted work; Christophe Pison reports personal fees and non-financial support from AstraZeneca, Boehringer Ingelheim, GSK, Novartis, outside the submitted work; Mihael Polymeropulos reports other from Vanda Pharmaceuticals, outside the submitted work; Ulrike Protzer reports grants from VIR Biotechnology, grants and non-financial support from Roche, grants from Alios-JJ Peter Sterk reports other from Breathomix BV, outside the submitted work; Giulio Superti-Furga reports being a shareholder of Proxygen GmbH, Haplogen GmbH and Allcyte GmbH, outside the submitted work; Vincenzo Valentini reports personal fees from Merck Serono, Betaglue, outside the submitted work; Sylvie van der Werf reports other from Sanofi Pasteur, outside the submitted work; she is the holder of four issued patents. The other authors have declared no conflict of interest.

BackgroundUniversal access to assessment and treatment of mental health and learning disorders remains a significant and unmet need. There is a vast number of people without access to care because of economic, geographic, and cultural barriers as well as limited availability of clinical experts who could help advance our understanding of mental health.ObjectiveTo create an open, configurable software platform to build clinical measures, mobile assessments, tasks, and interventions without programming expertise. Specifically, our primary requirements include: an administrator interface for creating and scheduling recurring and customized questionnaires where end users receive and respond to scheduled notifications via an iOS or Android app on a mobile device. Such a platform would help relieve overwhelmed health systems, and empower remote and disadvantaged subgroups in need of accurate and effective information, assessment, and care. This platform has potential to advance scientific research by supporting the collection of data with instruments tailored to specific scientific questions from large, distributed, and diverse populations.MethodsWe conducted a search for tools that satisfy the above requirements. We designed and developed a new software platform called “MindLogger” that exceeds the above requirements. To demonstrate the tool’s configurability, we built multiple “applets” (collections of activities) within the MindLogger mobile application and deployed several, including a comprehensive set of assessments underway in a large-scale, longitudinal, mental health study.ResultsOf the hundreds of products we researched, we found 10 that met our primary requirements above with 4 that support end-to-end encryption, 2 that enable restricted access to individual users’ data, 1 that provides open source software, and none that satisfy all three. We compared features related to information presentation and data capture capabilities, privacy and security, and access to the product, code, and data. We successfully built MindLogger mobile and web applications, as well as web browser-based tools for building and editing new applets and for administering them to end users. MindLogger has end-to-end encryption, enables restricted access, is open source, and supports a variety of data collection features. One applet is currently collecting data from children and adolescents in our mental health study, and other applets are in different stages of testing and deployment for use in clinical and research settings.ConclusionsWe have demonstrated the flexibility and applicability of the MindLogger platform through its deployment in a large-scale, longitudinal, mobile mental health study, and by building a variety of other mental health-related applets. With this release, we encourage a broad range of users to apply the MindLogger platform to create and test applets to advance health care and scientific research. We hope that increasing availability of applets designed to assess and administer interventions will facilitate access to health care in the general population.

BACKGROUND Universal access to assessment and treatment of mental health and learning disorders remains a significant and unmet need. There are many people without access to care because of economic, geographic, and cultural barriers, as well as the limited availability of clinical experts who could help advance our understanding and treatment of mental health. OBJECTIVE This study aims to create an open, configurable software platform to build clinical measures, mobile assessments, tasks, and interventions without programming expertise. Specifically, our primary requirements include an administrator interface for creating and scheduling recurring and customized questionnaires where end users receive and respond to scheduled notifications via an iOS or Android app on a mobile device. Such a platform would help relieve overwhelmed health systems and empower remote and disadvantaged subgroups in need of accurate and effective information, assessment, and care. This platform has the potential to advance scientific research by supporting the collection of data with instruments tailored to specific scientific questions from large, distributed, and diverse populations. METHODS We searched for products that satisfy these requirements. We designed and developed a new software platform called MindLogger, which exceeds the requirements. To demonstrate the platform’s configurability, we built multiple applets (collections of activities) within the MindLogger mobile app and deployed several of them, including a comprehensive set of assessments underway in a large-scale, longitudinal mental health study. RESULTS Of the hundreds of products we researched, we found 10 that met our primary requirements with 4 that support end-to-end encryption, 2 that enable restricted access to individual users’ data, 1 that provides open-source software, and none that satisfy all three. We compared features related to information presentation and data capture capabilities; privacy and security; and access to the product, code, and data. We successfully built MindLogger mobile and web applications, as well as web browser–based tools for building and editing new applets and for administering them to end users. MindLogger has end-to-end encryption, enables restricted access, is open source, and supports a variety of data collection features. One applet is currently collecting data from children and adolescents in our mental health study, and other applets are in different stages of testing and deployment for use in clinical and research settings. CONCLUSIONS We demonstrated the flexibility and applicability of the MindLogger platform through its deployment in a large-scale, longitudinal, mobile mental health study and by building a variety of other mental health–related applets. With this release, we encourage a broad range of users to apply the MindLogger platform to create and test applets to advance health care and scientific research. We hope that increasing the availability of applets designed to assess and administer interventions will facilitate access to health care in the general population.

RNA-based regulation offers a promising alternative of protein-based transcriptional networks. However, designing synthetic riboregulators with desirable functionalities using arbitrary sequences remains challenging, due in part to insufficient exploration of RNA sequence-to-function landscapes. Here we report that CRISPR-Csy4 mediates a nearly all-or-none processing of precursor CRISPR RNAs (pre-crRNAs), by profiling Csy4 binding sites flanked by > 1 million random sequences. This represents an ideal sequence-to-function space for universal riboregulator designs. Lacking discernible sequence-structural commonality among processable pre-crRNAs, we trained a neural network for accurate classification (f1-score ≈ 0.93). Inspired by exhaustive probing of palindromic flanking sequences, we designed anti-CRISPR RNAs (acrRNAs) that suppress processing of pre-crRNAs via stem stacking. We validated machine-learning-guided designs with >30 functional pairs of acrRNAs and pre-crRNAs to achieve switch-like properties. This opens a wide range of plug-and-play applications tailored through pre-crRNA designs, and represents a programmable alternative to protein-based anti-CRISPRs.

International audience

Biochemical processes often require spatial regulation and specific microenvironments. The general lack of organelles in bacteria limits the potential of bioengineering complex intracellular reactions. Here, we demonstrate synthetic membraneless organelles in Escherichia coli termed transcriptionally engineered addressable RNA solvent droplets (TEARS). TEARS are assembled from RNA-binding protein recruiting domains fused to poly-CAG repeats that spontaneously drive liquid-liquid phase separation from the bulk cytoplasm. Targeting TEARS with fluorescent proteins revealed multilayered structures with composition and reaction robustness governed by non-equilibrium dynamics. We show that TEARS provide organelle-like bioprocess isolation for sequestering biochemical pathways, controlling metabolic branch points, buffering mRNA translation rates, and scaffolding protein-protein interactions. We anticipate TEARS to be a simple and versatile tool for spatially controlling E. coli biochemistry. Particularly, the modular design of TEARS enables applications without expression fine-tuning, simplifying the design-build-test cycle of bioengineering.

Contributing to the global turn of WWI studies, this chapter explores a unique relief effort that entwined Middle Eastern migrants in the United States and American missionaries in the Ottoman Empire. Cut off from established banking channels to send money to their families, migrants from Ottoman Syria living in the Mahjar employed the Presbyterian Church’s American Mission Press in Beirut to facilitate “remittance relief” efforts during and after the Great War. Contextualized within the entangled history of migration between the Middle East and the Americas, this chapter employs records held at the Near East School of Theology in Beirut, Lebanon and the Presbyterian Historical Society in Philadelphia, PA to illuminate the process through which the remittances were organized and transferred from New York to Beirut, and point further connected. The chapter includes a preliminary study of the remittances themselves, which reveals important trends in Syrian migration to the United States leading up to and during the Great War, and how these trends impacted the transnational relief whereby Middle Easterners abroad saved Middle Easterners back home.

Biochemical processes often require spatial regulation and specific microenvironments. The general lack of organelles in bacteria limits the potential of bioengineering complex intracellular reactions. Here we demonstrate Transcriptionally Engineered Addressable RNA Solvent droplets (TEARS) as synthetic microdomains within theEscherichia coli.TEARS are assembled from RNA-binding protein recruitment domains fused to poly-CAG repeats that spontaneously drive liquid-liquid phase separation from the bulk cytoplasm. Targeting TEARS with fluorescent proteins revealed multilayered structures and a non-equilibrium mechanism controlling their composition and reaction robustness. We show that TEARS provide organelle-like bioprocess isolation for sequestering biochemical pathways, controlling metabolic branch points, buffering mRNA translation rates and scaffolding protein-protein interactions. TEARS are a simple and versatile tool for spatially controllingE. colibiochemistry.