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The platform was also used to design sequences that directed specificity among closely related reader domains, an application which may have utility in the development of peptidomimetic inhibitors. Furthermore, we used the platform to identify binding determinants of site-specific histone Kme antibodies and surprisingly revealed that only a few amino acids drove epitope recognition. Collectively, these studies introduce and validate a rapid, unbiased, and high-throughput binding assay for Kme readers, and we envision its use as a resource for expanding the study of Kme-driven protein interactions. Copyright © 2020 Kupai, Vaughan, Dickson and Rothbart.The mechanisms leading to NOD-leucine rich repeat and pyrin containing protein 3 (NLRP3) inflammasome activation are still debated. It is well established that oligomerized NLRP3 interacts with apoptosis associated Speck-like protein containing a CARD domain (ASC) which polymerizes into filaments recruiting procaspase-1, leading to its activation. However, pathways triggering NLRP3 activation, such as potassium efflux, ROS production or lysosomal permeabilization, can be required or not, depending on the activators used. Here we proposed to evaluate the importance of Cathepsin B on NLRP3 inflammasome assembly and activation. Using Cathepsin B-/- BMDMs (Bone Marrow-Derived Macrophages), we first show that Cathepsin B is required for caspase-1 activation, IL-1β production and ASC speck formation, upon treatment with different types of NLRP3 activators, i.e., ATP, nigericin or crystals. Moreover, in these conditions, Cathepsin B interacts with NLRP3 at the endoplasmic reticulum (ER) level. To conclude, different NLRP3 activators lead to Cathepsin B interaction with NLRP3 at the ER level and to subsequent caspase-1 activation. Copyright © 2020 Chevriaux, Pilot, Derangère, Simonin, Martine, Chalmin, Ghiringhelli and Rébé.With the ability to switch between proliferative and contractile phenotype, airway smooth muscle (ASM) cells can contribute to the progression of airway diseases such as asthma and chronic obstructive pulmonary disease (COPD), in which airway obstruction is associated with ASM hypertrophy and hypercontractility. A-kinase anchoring proteins (AKAPs) have emerged as important regulatory molecules in various tissues, including ASM cells. AKAPs can anchor the regulatory subunits of protein kinase A (PKA), and guide cellular localization via various targeting domains. Here we investigated whether disruption of the AKAP-PKA interaction, by the cell permeable peptide stearated (st)-Ht31, alters human ASM proliferation and contractility. Treatment of human ASM with st-Ht31 enhanced the expression of protein markers associated with cell proliferation in both cultured cells and intact tissue, although this was not accompanied by an increase in cell viability or cell-cycle progression, suggesting that disruption of AKAP-PKA interaction on its own is not sufficient to drive ASM cell proliferation. Strikingly, st-Ht31 enhanced contractile force generation in human ASM tissue with concomitant upregulation of the contractile protein α-sm-actin. This upregulation of α-sm-actin was independent of mRNA stability, transcription or translation, but was dependent on proteasome function, as the proteasome inhibitor MG-132 prevented the st-Ht31 effect. Collectively, the AKAP-PKA interaction appears to regulate markers of the multi-functional capabilities of ASM, and this alter the physiological function, such as contractility, suggesting potential to contribute to the pathophysiology of airway diseases. Copyright © 2020 Baarsma, Han, Poppinga, Driessen, Elzinga, Halayko, Meurs, Maarsingh and Schmidt.The development of new high-tech systems for screening anticancer drugs is one of the main problems of preclinical screening. Poor correlation between preclinical in vitro and in vivo data with clinical trials remains a major concern. The choice of the correct tumor model at the stage of in vitro testing provides reduction in both financial and time costs during later stages due to the timely screening of ineffective agents. In view of the growing incidence of oncology, increasing the pace of the creation, development and testing of new antitumor agents, the improvement and expansion of new high-tech systems for preclinical in vitro screening is becoming very important. The pharmaceutical industry presently relies on several widely used in vitro models, including two-dimensional models, three-dimensional models, microfluidic systems, Boyden's chamber and models created using 3D bioprinting. This review outlines and describes these tumor models including their use in research, in addition to their characteristics. This review therefore gives an insight into in vitro based testing which is of interest to researchers and clinicians from differing fields including pharmacy, preclinical studies and cell biology. PKM2-IN-1 price Copyright © 2020 Kitaeva, Rutland, Rizvanov and Solovyeva.HIV-1 envelope glycoprotein (Env) remains the most relevant target for the elicitation of functional antibodies to HIV by vaccination. However, soluble Env antigens often do not elicit the desired immune responses. Delivering subunit antigens on particulate nanoparticles is an established approach to improve their immunogenicity. In this study the sequence encoding Zera®, a proline-rich domain derived from the γ-zein storage protein, was fused to either the C- or N-terminus of the superinfecting HIV-1 CAP256 gp140 envelope Zera® generally induces the formation of protein bodies (PBs), which can significantly improve both the immunogenicity and yields of the partner protein. The expression of gp140-Zera® and Zera®-gp140 (N- and C-terminal fusions respectively) in mammalian cells was confirmed by western blot analysis and immunostaining. However, isopycnic ultracentrifugation showed that neither gp140-Zera® nor Zera®-gp140 accumulated in characteristic electron-dense PBs. gp140-Zera® elicited higher binding antibody titers in rabbits to autologous gp140 and V1V2 scaffold than Zera®-gp140. Rabbit anti-gp140-Zera® sera also had significantly higher Tier 1A neutralizing antibody titers than anti-Zera®-gp140 sera. Neither gp140-Zera® nor Zera®-gp140-specific sera neutralized Tier 1B or autologous Tier 2 viruses. These results showed that HIV-1 gp140 tagged with Zera® at either the N- or C-termini elicited high titers of gp140 and V1V2 binding antibodies, and low levels of Tier 1 neutralizing antibodies in rabbits. Copyright © 2020 Ximba, Chapman, Meyers, Margolin, van Diepen, Williamson and Rybicki.