Targeted proteolysis of endogenous proteins is desirable as a research toolkit and in therapeutics. CRISPR/Cas9-mediated gene knockouts are irreversible and often not feasible for many genes. Similarly, RNA interference approaches necessitate prolonged treatments, can lead to incomplete knockdowns and are often associated with off-target effects. Targeted proteolysis can overcome these limitations. In this report, we describe an affinity-directed protein missile (AdPROM) system that harbours the von Hippel–Lindau (VHL) protein, the substrate receptor of the Cullin2 (CUL2) E3 ligase complex, tethered to polypeptide binders that selectively bind and recruit endogenous target proteins to the CUL2-E3 ligase complex for ubiquitination and proteasomal degradation. By using synthetic monobodies that selectively bind the protein tyrosine phosphatase SHP2 and a camelid-derived VHH nanobody that selectively binds the human ASC protein, we demonstrate highly efficient AdPROM-mediated degradation of endogenous SHP2 and ASC in human cell lines. We show that AdPROM-mediated loss of SHP2 in cells impacts SHP2 biology. This study demonstrates for the first time that small polypeptide binders that selectively recognize endogenous target proteins can be exploited for AdPROM-mediated destruction of the target proteins.
The BMP and Wnt signalling pathways determine axis specification during embryonic development. Our previous work has shown that PAWS1 (also known as FAM83G) interacts with SMAD1 and modulates BMP signalling. Here, surprisingly, we show that overexpression of PAWS1 in Xenopus embryos activates Wnt signalling and causes complete axis duplication. Consistent with these observations in Xenopus, Wnt signalling is diminished in U2OS osteosarcoma cells lacking PAWS1, while BMP signalling is unaffected. We show that PAWS1 interacts and co‐localises with the α isoform of casein kinase 1 (CK1), and that PAWS1 mutations incapable of binding CK1 fail both to activate Wnt signalling and to elicit axis duplication in Xenopus embryos.
Wireless Power Transfer (WPT) offers a viable means of charging Electric Vehicles (EV)'s whilst in a dynamic state (DWPT), mitigating issues concerning vehicle range, the size of on-board energy storage and the network distribution of static based charging systems. Such Charge While Driving (CWD) technology has the capability to accelerate EV market penetration through increasing user convenience, reducing EV costs and increasing driving range indefinitely, dependent upon sufficient charging infrastructure. This paper reviews current traction battery technologies, conductive and inductive charging processes, influential parameters specific to the dynamic charging state as well as highlighting notable work undertaken within the field of WPT charging systems. DWPT system requirements, specific to the driver, vehicle and infrastructure interaction environment are summarised and international standards highlighted in order to acknowledge the work that must be done within this area. It is important to recognise that the gap is not currently technological; instead, it is an implementation issue. Without the necessary standardisation, system architectures cannot be developed and implemented without fear of interoperability issues between countries or indeed systems. For successful deployment, the technologies impact should be maximised with the minimum quantity of infrastructure and technology use, deployment scenarios and locations are discussed that have the potential to bring this to fruition. IntroductionThe electrification of road transport provides a viable means of reducing fossil fuel consumption and environmental pollution, hence the recent advancements in Electric Vehicle (EV) design and performance [1]. However, the high costs and poor specific energy densities of batteries compared to fossil fuels results in a less than ideal scenario [2]. Due to their relatively shorter range, EV's require more frequent charging (than refuelling of Internal Combustion Engine (ICE) vehicles) to maintain a desirable range and with long charge times (compared with conventional refuelling times) or potential battery degradation that occurs during rapid charging; battery charging technology has restricted EV development. With no significant advancements in battery technology that would bring EV range in line with comparable ICE vehicles forecasted within the foreseeable future [3] this has resulted in substantial research into alternative charging methods. Whilst conventional plug in charging is the most common form, there are still conductive energy losses within the system resulting in an overall efficiency of around 86% [4] and potentially lower for rapid chargers. In addition, the high power transfer, human handling and the ability for the user to forget to plug in/out result in a pragmatic scenario.Wireless Power Transfer (WPT) technology is capable of mitigating the issues of plug in charging; the EV is parked over a coil that inductively transfers electrical energy to a receiver coil positioned on the vehicle. ...
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