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Measuring Composition of CD95 Death-Inducing Signaling Complex and Processing of Procaspase-8 in this Complex. | LitMetric

AI Article Synopsis

  • - Extrinsic apoptosis is triggered by death receptors like CD95 and TRAIL receptors, which, when activated by their ligands, form a signaling complex called the death-inducing signaling complex (DISC).
  • - The DISC includes key proteins such as FADD and procaspases-8/-10, and is crucial for activating procaspase-8 through a series of steps involving dimerization within the complex.
  • - A new experimental workflow using immunoprecipitation and western blot techniques is outlined in the text, enabling researchers to study the formation of the DISC and the activation of procaspase-8 in detail.

Article Abstract

Extrinsic apoptosis is mediated by the activation of death receptors (DRs) such as CD95/Fas/APO-1 or tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-receptor 1/receptor 2 (TRAIL-R1/R2). Stimulation of these receptors with their cognate ligands leads to the assembly of the death-inducing signaling complex (DISC). DISC comprises DR, the adaptor protein Fas-associated protein with death domain (FADD), procaspases-8/-10, and cellular FADD-like interleukin (IL)-1β-converting enzyme-inhibitory proteins (c-FLIPs). The DISC serves as a platform for procaspase-8 processing and activation. The latter occurs via its dimerization/oligomerization in the death effector domain (DED) filaments assembled at the DISC. Activation of procaspase-8 is followed by its processing, which occurs in several steps. In this work, an established experimental workflow is described that allows the measurement of DISC formation and the processing of procaspase-8 in this complex. The workflow is based on immunoprecipitation techniques supported by western blot analysis. This workflow allows careful monitoring of different steps of procaspase-8 recruitment to the DISC and its processing and is highly relevant for investigating molecular mechanisms of extrinsic apoptosis.

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Source
http://dx.doi.org/10.3791/62842DOI Listing

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