Chemical Biology of Dynein

Cytoplasmic dynein is a AAA (ATPase Associated with various Activities) motor protein that transports cellular cargoes towards the microtubule minus-end. Despite its essential role in intracellular transport, dynein remains the least understood cytoskeletal motor. With speeds \u3e25 μm/min in cells, dynein’s cellular functions are challenging to study using genetic approaches, such as CRISPR and RNA interference, as the perturbation timescales far exceed those of dynein action. Fast-acting small molecule probes can be powerful tools to study dynein’s many cellular mechanisms, but the design of potent and selective inhibitors of dynein remains challenging. Inhibitors of dynein’s motor domain have been reported, such as ciliobrevins, dynapyrazoles and dynarrestin, but the inhibitor-binding site(s) have yet to be elucidated. Though inhibitor-bound dynein structures have been long-sought after, efforts have been curtailed by difficulties in obtaining high-resolution dynein structures and the lack of an inhibitor with the requisite chemical properties, such as compound solubility and stability, for structural studies. The work presented here describes the design and characterization of a class of dynein probes, one of which we use to obtain an inhibitor-bound structure of dynein. The first chapter “Chemical inhibitors of AAA proteins” provides a brief overview of AAA proteins in general and the inhibitors that have been designed. The chapter emphasizes on the design and characterization of ciliobrevins, dynapyrazoles and dynarrestin, three classes of dynein inhibitors with distinct mechanisms of action. Their limitations as cellular probes are discussed and motivates the need for compounds with improved potency and selectivity. In the second chapter, “Dynapyrazoles acutely inhibit intraflagellar transport”, I characterize dynapyrazole-A in a cell-based assay that can measure the effect of the compound on intraflagellar transport (IFT). Dynapyrazole-A is an acute reversible inhibitor of dynein 2-dependent retrograde transport and can be a useful probe to study IFT function. However, as dynapyrazole-A inhibits both isoforms of dynein, its use as a chemical probe is limited due to its cytotoxic effects at treatment times \u3e1 hour. This chapter motivates the need for dynein-2 specific inhibitors. Finally, I present a third chapter, “Structural insights into the chemical inhibition of dynein”, that discusses the design of a dynapyrazole derivative, compound 20, that inhibits the basal ATPase activity of human and S. cerevisiae dynein. I used cryo-EM to obtain a structure of S. cerevisae dynein’s motor domain in the presence of the dynapyrazole derivative and find that the compound binds to the regulatory ATPase sites in the AAA3 and AAA4 domains, rather than the main catalytic site in the AAA1 domain. This finding addresses a major gap in our knowledge, as inhibitors of dynein’s ATPase activity have been assumed to target the AAA1 domain. Inhibitor design efforts can now be focused on the regulatory ATPase sites to obtain potent and selective small molecule probes of dynein

Line-start permanent-magnet motor single-phase steady-state performance analysis

This paper describes an efficient calculating procedure for the steady-state operation of a single-phase line-start capacitor-run permanent-magnet motor. This class of motor is beginning to be applied in hermetic refrigerator compressors as a high-efficiency alternative to either a plain induction motor or a full inverter-fed drive. The calculation relies on a combination of reference-frame transformations including symmetrical components to cope with imbalance, and dq axes to cope with saliency. Computed results are compared with test data. The agreement is generally good, especially in describing the general properties of the motor. However, it is shown that certain important effects are beyond the limit of simple circuit analysis and require a more complex numerical analysis method

Effects of the COVID-19 Pandemic on Physical Activity in Children: A Systematic Rapid Review

Background: Prolonged lockdowns, put in place to mitigate the spread of COVID-19, may have altered physical activity (PA). The goal of this systematic rapid review was to synthesize the global impact of the COVID-19 pandemic on PA of children. Methods: Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines were used to conduct a database search on 03/29/2021. Included studies were peer- reviewed, in English language, and encompassed both a measure of PA during the COVID-19 pandemic and PA data for children aged 18 and younger. The database search yielded 677 unique citations and, ultimately, 69 articles were included in the review. Results: During the COVID-19 pandemic, frequently reported PA included walking, unstructured play, and virtual PA through online platforms. Of the articles that reported changes in PA of children during the COVID-19 pandemic, 89% reported PA (e.g., frequency, intensity, duration, or the percentage of children who met PA recommendations) of children decreased. Boys, younger children, children who lived with other children, and children with more outdoor play space had higher levels of PA. Conclusions: The COVID-19 pandemic resulted in decreased PA among children around the world. Further work is needed to develop policies to support global PA increases

Five key attributes can increase marine protected areas performance for small-scale fisheries management

Marine protected areas (MPAs) have largely proven to be effective tools for conserving marine ecosystem, while socio-economic benefits generated by MPAs to fisheries are still under debate. Many MPAs embed a no-take zone, aiming to preserve natural populations and ecosystems, within a buffer zone where potentially sustainable activities are allowed. Small-scale fisheries (SSF) within buffer zones can be highly beneficial by promoting local socio-economies. However, guidelines to successfully manage SSFs within MPAs, ensuring both conservation and fisheries goals, and reaching a win-win scenario, are largely unavailable. From the peer-reviewed literature, grey-literature and interviews, we assembled a unique database of ecological, social and economic attributes of SSF in 25 Mediterranean MPAs. Using random forest with Boruta algorithm we identified a set of attributes determining successful SSFs management within MPAs. We show that fish stocks are healthier, fishermen incomes are higher and the social acceptance of management practices is fostered if five attributes are present (i.e. high MPA enforcement, presence of a management plan, fishermen engagement in MPA management, fishermen representative in the MPA board, and promotion of sustainable fishing). These findings are pivotal to Mediterranean coastal communities so they can achieve conservation goals while allowing for profitable exploitation of fisheries resources

Chemical structure-guided design of dynapyrazoles, potent cell-permeable dynein inhibitors with a unique mode of action

Cytoplasmic dyneins are motor proteins in the AAA+ superfamily that transport cellular cargos toward microtubule minus-ends. Recently, ciliobrevins were reported as selective cell-permeable inhibitors of cytoplasmic dyneins. As is often true for first-in-class inhibitors, the use of ciliobrevins has in part been limited by low potency. Moreover, suboptimal chemical properties, such as the potential to isomerize, have hindered efforts to improve ciliobrevins. Here, we characterized the structure of ciliobrevins and designed conformationally constrained isosteres. These studies identified dynapyrazoles, inhibitors more potent than ciliobrevins. At single-digit micromolar concentrations dynapyrazoles block intraflagellar transport in the cilium and lysosome motility in the cytoplasm, processes that depend on cytoplasmic dyneins. Further, we find that while ciliobrevins inhibit both dynein's microtubule-stimulated and basal ATPase activity, dynapyrazoles strongly block only microtubule-stimulated activity. Together, our studies suggest that chemical-structure-based analyses can lead to inhibitors with improved properties and distinct modes of inhibition

Cerebral vein thrombosis in patients with Philadelphia-negative myeloproliferative neoplasms : an European Leukemia Net study

To investigate the characteristics and clinical course of cerebral vein thrombosis (CVT) in patients with myeloproliferative neoplasms (MPN) we compared 48 patients with MPN and CVT (group MPN-CVT) to 87 with MPN and other venous thrombosis (group MPN-VT) and 178 with MPN and no thrombosis (group MPN-NoT) matched by sex, age at diagnosis of MPN (\ub15 years) and type of MPN. The study population was identified among 5,500 patients with MPN, from January 1982 to June 2013. Thrombophilia abnormalities were significantly more prevalent in the MPN-CVT and MPN-VT than in MPN-NoT group (P = 0.015), as well as the JAK2 V617F mutation in patients with essential thrombocythemia (P = 0.059). Compared to MPN-VT, MPN-CVT patients had a higher rate of recurrent thrombosis (42% vs. 25%, P = 0.049) despite a shorter median follow-up period (6.1 vs. 10.3 years, P = 0.019), a higher long-term antithrombotic (94% vs. 84%, P = 0.099) and a similar cytoreductive treatment (79% vs. 70%, P = 0.311). The incidence of recurrent thrombosis was double in MPN-CVT than in MPN-VT group (8.8% and 4.2% patient-years, P = 0.022), and CVT and unprovoked event were the only predictive variables in a multivariate model including also sex, blood count, thrombophilia, cytoreductive, and antithrombotic treatment (HR 1.97, 95%CI 1.05-3.72 and 2.09, 1.09-4.00, respectively). \ua9 2014 Wiley Periodicals, Inc

Alternative circuits on agro-food markets

The increasing development of alternative forms in production, marketing and consumption in the agrifood sector make us to reflect on how these different modalities can be explained with a model of production and consumption in which the neoclassical theory of supply and demand is still central in explaining individual and collective behavior. The analysis starts by developing a taxonomy of "alternative" market systems to identify specific characteristics of producers and consumers. The taxonomy of "alternative" systems market concerns the identification and description of "alternative" markets and the definition of features that may characterize the markets in order to put them in a taxonomic classification. Among the common features of the markets we find the type of product, usually food, whose quality characteristics are defined by the producer\u2010consumer interaction. The alternative marketing channels are usually locally based. The experiences analyzed are developed according to the logic of communicative rationality which is explain by the economic theory of the agency. In some cases the communicative rationality is more evident, in other less, but in all cases is based on direct contact between producers and consumers. In this analysis, the need to make a critical reading of the market economic theory, in the light of the diversity that exists in human behavior, has been central, trying to build an interpretation model of the consumption and production systems. Starting from the analysis of the mechanisms leading to the choice of alternative agri\u2010food markets, it has been possible to develop an explanation for the differences in the behavior of producers and consumers based on the theories of the Gaussian distribution of populations. This idea was verified by direct analysis, through questionnaires given to the users of different way of commerce like buying groups, adoption of product, Farmer's Market, Earth Markets. For the formulation of the questionnaires were considered different socio\u2010economic variables useful to create the bases to understand social behavior of both producers and consumers. This paper highlights heterogeneity in the behavior of actors in the economic system showing experiences where the communication and definition of quality criteria, shared by producer and consumer, allows the survival of small local farms

Common Visions Between Farmers and Consumers as a Tool to Characterize \u201cNew\u201d Agro-Food Markets - European Society for Ecological Economics conference ESEE Instanbul 2011

Several alternative forms of trade in the agro-food market are spreading in Italy at local and global levels, namely: Farmers\u2019 Market; Solidarity Purchasing Groups, Organized Demand and Supply Groups; Product Adoptions; Solidarity Trade Districts; Farm Shops. These have been introduced as a reaction to the global standardization and homologation of agricultural products leading to biodiversity loss. This research aims at examining more in detail how social and economic relations are connected in the new food market channels. The main result focuses on the central role of relational communication between the subjects involved in the process. Communication means the sharing of information about products, producers and farming systems. In the different forms of alternative circuits in the agro-food system, the attention is mainly focused on a particular type of product trade, the Solidarity Purchasing Groups. Solidarity Purchasing Groups consist of people meeting to make collective purchases of different food and non-food products. People who are part of the buying groups are linked by shared consumption choices. The producers and products to be bought are chosen carefully according to farming methods, normally organic, and producer characteristics favouring small farmers. The case study analysis was developed from a direct survey. The questionnaires were sent by mail to the representatives of Solidarity Purchasing Groups of Northeastern Italy (Veneto, Friuli Venezia Giulia and Trentino). 202 questionnaires were collected from 24 Solidarity Purchasing Groups registered on the official SPG web site and 5 from unregistered SPG sites. The analysis of the answers given by the consumers shows the role of relational communication which is the basis of the social network built up through consumers and producers on the local scale. The results of this research were interpreted by using economic theories based on communication and relationship such as the Stewardship Theory (Davis et al., 1997) Relational communication is the taxonomical key which makes the Solidarity Purchasing Groups different from the other alternative circuits in agro-food market