Structure-Activity Relationship of Acetylenes from Galls of Hedera rhombea as Plant Growth Inhibitors

The structure-activity relationship of 12 isolated acetylenes from galls of Hedera rhombea (Araliaceae) induced by Asphondylia sp. (Cecidomyiidae) and their derivatives has been studied for the inhibition of the shoot and root growth of rice, perennial ryegrass, cockscomb, lettuce, and cress. Almost all acetylenes generally showed growth inhibitory activity. The diacetylenes exhibited higher activity than the monoacetylenes, suggesting that a conjugated diyne segment is essential for the activity. On the other hand, the acetylenes with a nonoxidated methylene group at C-8 showed stronger activity comparing with those possessing hydroxy and acetoxy groups at C-8. Furthermore, it has been demonstrated that the acetylenes bearing a terminal olefinic group at C-16,C-17 enhanced the activity. It is thus clarified that important sites for the activity of the acetylenes from galls of H. rhombea are a conjugated diyne and a terminal olefinic group connecting to the aliphatic chain and that less oxidated compounds show more activity

MCC950/CRID3 potently targets the NACHT domain of wild-type NLRP3 but not disease-associated mutants for inflammasome inhibition

The nucleotide-binding-domain (NBD)-and leucine-rich repeat (LRR)-containing (NLR) family, pyrin-domain-containing 3 (NLRP3) inflammasome drives pathological inflammation in a suite of autoimmune, metabolic, malignant, and neurodegenerative diseases. Additionally, NLRP3 gain-of-function point mutations cause systemic periodic fever syndromes that are collectively known as cryopyrin-associated periodic syndrome (CAPS). There is significant interest in the discovery and development of diarylsulfonylurea Cytokine Release Inhibitory Drugs (CRIDs) such as MCC950/CRID3, a potent and selective inhibitor of the NLRP3 inflammasome pathway, for the treatment of CAPS and other diseases. However, drug discovery efforts have been constrained by the lack of insight into the molecular target and mechanism by which these CRIDs inhibit the NLRP3 inflammasome pathway. Here, we show that the NAIP, CIITA, HET-E, and TP1 (NACHT) domain of NLRP3 is the molecular target of diarylsulfonylurea inhibitors. Interestingly, we find photoaffinity labeling (PAL) of the NACHT domain requires an intact (d)ATP-binding pocket and is substantially reduced for most CAPS-associated NLRP3 mutants. In concordance with this finding, MCC950/CRID3 failed to inhibit NLRP3-driven inflammatory pathology in two mouse models of CAPS. Moreover, it abolished circulating levels of interleukin (IL)-1 beta and IL-18 in lipopolysaccharide (LPS)-challenged wild-type mice but not in Nlrp3(L351P) knock-in mice and ex vivo-stimulated mutant macrophages. These results identify wild-type NLRP3 as the molecular target of MCC950/CRID3 and show that CAPS-related NLRP3 mutants escape efficient MCC950/CRID3 inhibition. Collectively, this work suggests that MCC950/CRID3-based therapies may effectively treat inflammation driven by wild-type NLRP3 but not CAPS-associated mutants

細胞の接着と増殖を促進するダンベル型小分子化合物

京都大学0048新制・課程博士博士(医科学)甲第15273号医科博第14号新制||医科||2(附属図書館)27751京都大学大学院医学研究科医科学専攻(主査)教授 成宮 周, 教授 鈴木 茂彦, 教授 瀬原 淳子学位規則第4条第1項該当Doctor of Medical ScienceKyoto UniversityDA

Heterobifunctional Molecules Induce Dephosphorylation of Kinases - a Proof of Concept Study

Previous work has shown that heterodimeric molecules, in some instances referred to as PROteolysis Targeting Chimeras, promote proximity of an E3-ligase to a target of interest to induce target ubiquitination. We hypothesize that many other PTMs, beyond ubiquitination, can be catalyzed in a similar matter. This work utilizes recruitment of PP1 (protein phosphatase 1) to induce de phosphorylation of target proteins (AKT and EGFR). To our knowledge, this work represents the first examples of small molecules recruiting non-native partners to induce removal of a PTM. <br /

Nitroreductase-Activatable Morpholino Oligonucleotides for <i>in Vivo</i> Gene Silencing

Phosphorodiamidate morpholino oligonucleotides are widely used to interrogate gene function in whole organisms, and light-activatable derivatives can reveal spatial and temporal differences in gene activity. We describe here a new class of caged morpholino oligonucleotides that can be activated by the bacterial nitroreductase NfsB. We characterize the activation kinetics of these reagents <i>in vitro</i> and demonstrate their efficacy in zebrafish embryos that express NfsB either ubiquitously or in defined cell populations. In combination with transgenic organisms, such enzyme-actuated antisense tools will enable gene silencing in specific cell types, including tissues that are not amenable to optical targeting

Basal constriction during midbrain–hindbrain boundary morphogenesis is mediated by Wnt5b and focal adhesion kinase

Basal constriction occurs at the zebrafish midbrain–hindbrain boundary constriction (MHBC) and is likely a widespread morphogenetic mechanism. 3D reconstruction demonstrates that MHBC cells are wedge-shaped, and initially constrict basally, with subsequent apical expansion. wnt5b is expressed in the MHB and is required for basal constriction. Consistent with a requirement for this pathway, expression of dominant negative Gsk3β overcomes wnt5b knockdown. Immunostaining identifies focal adhesion kinase (Fak) as active in the MHB region, and knockdown demonstrates Fak is a regulator of basal constriction. Tissue specific knockdown further indicates that Fak functions cell autonomously within the MHBC. Fak acts downstream of wnt5b, suggesting that Wnt5b signals locally as an early step in basal constriction and acts together with more widespread Fak activation. This study delineates signaling pathways that regulate basal constriction during brain morphogenesis

Post-transcriptional mechanisms contribute to Etv2 repression during vascular development

Abstractetv2 is an endothelial-specific ETS transcription factor that is essential for vascular differentiation and morphogenesis in vertebrates. While recent data suggest that Etv2 is dynamically regulated during vascular development, little is known about the mechanisms involved in this process. Here, we find that etv2 transcript and protein expression are highly dynamic during zebrafish vascular development, with both apparent during early somitogenesis and subsequently down-regulated as development proceeds. Inducible knockdown of Etv2 in zebrafish embryos prior to mid-somitogenesis stages, but not later, caused severe vascular defects, suggesting a specific role in early commitment of lateral mesoderm to the endothelial linage. Accordingly, Etv2-overexpressing cells showed an enhanced ability to commit to endothelial lineages in mosaic embryos. We further find that the etv2 3' untranslated region (UTR) is capable of repressing an endothelial autonomous transgene and contains binding sites for members of the let-7 family of microRNAs. Ectopic expression of let-7a could repress the etv2 3'UTR in sensor assays and was also able to block endogenous Etv2 protein expression, leading to concomitant reduction of endothelial genes. Finally, we observed that Etv2 protein levels persisted in maternal-zygotic dicer1 mutant embryos, suggesting that microRNAs contribute to its repression during vascular development. Taken together, our results suggest that etv2 acts during early development to specify endothelial lineages and is then down-regulated, in part through post-transcriptional repression by microRNAs, to allow normal vascular development