RESUMO
The anticancer agent indisulam inhibits cell proliferation by causing degradation of RBM39, an essential mRNA splicing factor. Indisulam promotes an interaction between RBM39 and the DCAF15 E3 ligase substrate receptor, leading to RBM39 ubiquitination and proteasome-mediated degradation. To delineate the precise mechanism by which indisulam mediates the DCAF15-RBM39 interaction, we solved the DCAF15-DDB1-DDA1-indisulam-RBM39(RRM2) complex structure to a resolution of 2.3 Å. DCAF15 has a distinct topology that embraces the RBM39(RRM2) domain largely via non-polar interactions, and indisulam binds between DCAF15 and RBM39(RRM2), coordinating additional interactions between the two proteins. Studies with RBM39 point mutants and indisulam analogs validated the structural model and defined the RBM39 α-helical degron motif. The degron is found only in RBM23 and RBM39, and only these proteins were detectably downregulated in indisulam-treated HCT116 cells. This work further explains how indisulam induces RBM39 degradation and defines the challenge of harnessing DCAF15 to degrade additional targets.
Assuntos
Antineoplásicos/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular/química , Proteínas de Ligação a RNA/química , Sulfonamidas/farmacologia , Motivos de Aminoácidos , Calorimetria , Clonagem Molecular , Fluorometria , Células HCT116 , Células HEK293 , Humanos , Processamento de Imagem Assistida por Computador , Peptídeos e Proteínas de Sinalização Intracelular/genética , Cinética , Proteínas Nucleares/metabolismo , Peptídeos/química , Mutação Puntual , Ligação Proteica , Estrutura Quaternária de Proteína , Estrutura Secundária de Proteína , Proteoma , RNA Interferente Pequeno/metabolismo , Proteínas de Ligação a RNA/genética , Ubiquitina-Proteína Ligases/metabolismoRESUMO
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
RESUMO
The enzyme endo-polygalacturonase A, or PGA, is produced by the fungus, Aspergillus niger, and appears to play a critical role during invasion of plant cell walls. The enzyme has been homologously overexpressed in order to provide sufficient quantities of purified enzyme for structural and biological studies. We have characterized this enzyme in terms of its post-translational modifications (PTMs) and found it to be both N- and O-glycosylated. Additionally, we have characterized the glycosyl moieties using MALDI-TOF and LC-ESI mass spectrometry. The characterization of all PTMs on PGA, along with molecular modeling, allows us to reveal potential roles played by the glycans in modulating the interaction of the enzyme with other macromolecules.
Assuntos
Aspergillus niger/química , Poligalacturonase/química , Poligalacturonase/metabolismo , Polissacarídeos/química , Processamento de Proteína Pós-Traducional , Sequência de Aminoácidos , Glicopeptídeos/química , Glicosilação , Dados de Sequência Molecular , Estrutura Terciária de Proteína , Espectrometria de Massas por Ionização por Electrospray , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
[Correction Notice: An Erratum for this article was reported in Vol 144(4) of Journal of Experimental Psychology: General (see record 2015-33206-002). In the article the labels on the X-axis of Figure 1 "Remove Variance" and "Preserve Variance" should be switched.] Symbolic material objects such as art or certain artifacts (e.g., fine pottery, jewelry) share one common element: The combination of generating an expression, and the materialization of this expression in the object. This explains why people place a much greater value on handmade over machine-made objects, and originals over duplicates. We show that this mechanism occurs when a material object's symbolic property is salient and when the creator (artist or craftsman) is perceived to have agency control over the 1-to-1 materialized expression in the object. Coactivation of these 2 factors causes the object to be perceived as having high value because it is seen as the embodied representation of the creator's unique personal expression. In 6 experiments, subjects rated objects in various object categories, which varied on the type of object property (symbolic, functional, aesthetic), the production procedure (handmade, machine-made, analog, digital) and the origin of the symbolic information (person or software). The studies showed that the proposed mechanism applies to symbolic, but not to functional or aesthetic material objects. Furthermore, they show that this specific form of symbolic object valuation could not be explained by various other related psychological theories (e.g., uniqueness, scarcity, physical touching, creative performance). Our research provides a universal framework that identifies a core mechanism for explaining judgments of value for one of our most uniquely human symbolic object categories.