A FRET-based method for monitoring structural transitions in protein self-organization

Proteins assemble into a variety of dynamic and functional structures. Their structural transitions are often challenging to distinguish inside cells, particularly with a high spatiotemporal resolution. Here, we present a fluorescence resonance energy transfer (FRET)-based method for continuous and high-throughput monitoring of protein self-assemblies to reveal well-resolved transient intermediate states. Intermolecular FRET with both the donor and acceptor proteins at the same target protein provides high sensitivity while retaining the advantage of straightforward ratiometric imaging. We apply this method to monitor self-assembly of three proteins. We show that the mutant Huntingtin exon1 (mHttex1) first forms less-ordered assemblies, which develop into fibril-like aggregates, and demonstrate that the chaperone protein DNAJB6b increases the critical saturation concentration of mHttex1. We also monitor the structural changes in fused in sarcoma (FUS) condensates. This method adds to the toolbox for protein self-assembly structure and kinetics determination, and implementation with native or non-native proteins can inform studies involving protein condensation or aggregation

Combining Individual-Level Discrete Choice Experiment Estimates and Costs to Inform Health Care Management Decisions about Customized Care: The Case of Follow-Up Strategies after Breast Cancer Treatment

AbstractObjectiveCustomized care can be beneficial for patients when preferences for health care programs are heterogeneous. Yet, there is little guidance on how individual-specific preferences and cost data can be combined to inform health care decisions about customized care. Therefore, we propose a discrete choice experiment–based approach that illustrates how to analyze the cost-effectiveness of customized (and noncustomized) care programs to provide information for hospital managers.MethodsWe exploit the fact that choice models make it possible to determine whether preference heterogeneity exists and to obtain individual-specific parameter estimates. We present an approach of how to combine these individual-specific parameter estimates from a random parameter model (mixed logit model) with cost data to analyze the cost-effectiveness of customized care and demonstrate our method in the case of follow-up after breast cancer treatment.ResultsWe found that there is significant preference heterogeneity for all except two attributes of breast cancer treatment follow-up and that the fully customized care program leads to higher utility and lower costs than the current standardized program. Compared with the single alternative program, the fully customized care program has increased benefits and higher costs. Thus, it is necessary for health care decision makers to judge whether the use of resources for customized care is cost-effective.ConclusionsDecision makers should consider using the results obtained from our methodological approach when they consider implementing customized health care programs, because it may help to find ways to save costs and increase patient satisfaction

Erratum

We acknowledge cofunding of the research from the EU ITN project SynCrop (project number 764591), appointment of Luca Mantovanelli

A physicochemical roadmap of yeast replicative aging

Cellular aging is a multifactorial process that is characterized by a decline in homeostatic capacity, best described at the molecular level. Physicochemical properties such as pH and macromolecular crowding, are essential to all molecular processes in cells and require maintenance. Whether a drift in physicochemical properties contributes to the overall decline of homeostasis in aging is not known. Here we show that the cytosol of yeast cells acidifies modestly in early aging and sharply after senescence. Using a macromolecular crowding sensor optimized for long-term FRET measurements, we show the macromolecular crowding changes less in longer-lived cells in contrast to shorter-lived cells. While the average pH and crowding levels change only modestly with aging, we observe drastic changes in organellar volume, leading to crowding on the µm scale, which we term organellar crowding. Our measurements provide an initial framework of physicochemical parameters of replicatively-aged yeast cells

A physicochemical perspective of aging from single-cell analysis of pH, macromolecular and organellar crowding in yeast

Cellular aging is a multifactorial process that is characterized by a decline in homeostatic capacity, best described at the molecular level. Physicochemical properties such as pH and macromolecular crowding are essential to all molecular processes in cells and require maintenance. Whether a drift in physicochemical properties contributes to the overall decline of homeostasis in aging is not known. Here we show that the cytosol of yeast cells acidifies modestly in early aging and sharply after senescence. Using a macromolecular crowding sensor optimized for long-term FRET measurements, we show that crowding is rather stable and that the stability of crowding is a stronger predictor for lifespan than the absolute crowding levels. Additionally, in aged cells we observe drastic changes in organellar volume, leading to crowding on the µm scale, which we term organellar crowding. Our measurements provide an initial framework of physicochemical parameters of replicatively aged yeast cells

A physicochemical perspective of aging from single-cell analysis of ph, macromolecular and organellar crowding in yeast

Cellular aging is a multifactorial process that is characterized by a decline in homeostatic capacity, best described at the molecular level. Physicochemical properties such as pH and macromolecular crowding are essential to all molecular processes in cells and require maintenance. Whether a drift in physicochemical properties contributes to the overall decline of homeostasis in aging is not known. Here we show that the cytosol of yeast cells acidifies modestly in early aging and sharply after senescence. Using a macromolecular crowding sensor optimized for long-term FRET measurements, we show that crowding is rather stable and that the stability of crowding is a stronger predictor for lifespan than the absolute crowding levels. Additionally, in aged cells we observe drastic changes in organellar volume, leading to crowding on the µm scale, which we term organellar crowding. Our measurements provide an initial framework of physicochemical parameters of replicatively aged yeast cells. © 2020, eLife Sciences Publications Ltd. All rights reserved

Evaluating the Age-Based Recommendations for Long-Term Follow-Up in Breast Cancer

Background: After 5 years of annual follow-up following breast cancer, Dutch guidelines are age based: annual follow-up for women &lt;60 years, 60–75 years biennial, and none for &gt;75 years. We determined how the risk of recurrence corresponds to these consensus-based recommendations and to the risk of primary breast cancer in the general screening population. Subjects, Materials, and Methods: Women with early-stage breast cancer in 2003/2005 were selected from the Netherlands Cancer Registry (n = 18,568). Cumulative incidence functions were estimated for follow-up years 5–10 for locoregional recurrences (LRRs) and second primary tumors (SPs). Risks were compared with the screening population without history of breast cancer. Alternative cutoffs for age were determined by log-rank tests. Results: The cumulative risk for LRR/SP was lower in women &lt;60 years (5.9%, 95% confidence interval [CI] 5.3–6.6) who are under annual follow-up than for women 60–75 (6.3%, 95% CI 5.6–7.1) receiving biennial visits. All risks were higher than the 5-year risk of a primary tumor in the screening population (ranging from 1.4% to 1.9%). Age cutoffs &lt;50, 50–69, and &gt; 69 revealed better risk differentiation and would provide more risk-based schedules. Still, other factors, including systemic treatments, had an even greater impact on recurrence risks. Conclusion: The current consensus-based recommendations use suboptimal age cutoffs. The proposed alternative cutoffs will lead to a more balanced risk-based follow-up and thereby more efficient allocation of resources. However, more factors should be taken into account for truly individualizing follow-up based on risk for recurrence. Implications for Practice: The current age-based recommendations for breast cancer follow-up after 5 years are suboptimal and do not reflect the actual risk of recurrent disease. This results in situations in which women with higher risks actually receive less follow-up than those with a lower risk of recurrence. Alternative cutoffs could be a start toward risk-based follow-up and thereby more efficient allocation of resources. However, age, or any single risk factor, is not able to capture the risk differences and therefore is not sufficient for determining follow-up. More risk factors should be taken into account for truly individualizing follow-up based on the risk for recurrence.</p

Power engineering challenges in Zambia

The paper presents a review of current power engineering challenges in Zambia in relevance to the required potential necessary for the country’s modern economic development. An outline of the energy sector is provided. The driving need for enhancing the development of the energy sector is highlighted, accounted by the country’s annual increase of GDP, leading to an increase in electricity demand. Possible mitigations are suggested, with the development of Zambia's rich potential in renewable energy, as well as upgrading the operating power plants and constructing more off-grid and on-grid generating facilities