Abstract
The ubiquitin-proteasome system (UPS) plays a crucial part in normal cell function by mediating intracellular protein clearance. We have previously shown that UPS-mediated protein degradation varies in a cell type-specific manner in C. elegans. Here, we use formalin-fixed, paraffin-embedded C. elegans sections to enable studies on endogenous proteasome tissue expression. We show that the proteasome immunoreactivity pattern differs between cell types and within subcellular compartments in adult wild-type (N2) C. elegans. Interestingly, widespread knockdown of proteasome subunits by RNAi results in tissue-specific changes in proteasome expression instead of a uniform response. In addition, long-lived daf-2(e1370) mutants with impaired insulin/IGF-1 signaling (IIS) display similar proteasome tissue expression as aged-matched wild-type animals. Our study emphasizes the importance of alternate approaches to the commonly used whole animal lysate-based methods to detect changes in proteasome expression occurring at the sub-cellular, cell or tissue resolution level in a multicellular organism.
Highlights
Correct maintenance of the lifecycle of proteins, i.e., from their synthesis to their degradation, is critical for all organisms
We have demonstrated that long-lived daf-2 (e1370) mutants with reduced insulin/IGF-1 signaling (IIS) display increased in vivo ubiquitinproteasome system (UPS) activity in body-wall muscle cells and intestinal cells, without any detectable difference in proteasome amount in whole animal lysates [7]
We demonstrate that impaired insulin/IGF-1 signaling of daf-2 (e1370) mutants does not affect tissue expression of the proteasome, when compared to wildtype animals, supporting a role for tissue-specific post-translational regulation of UPS activity
Summary
Correct maintenance of the lifecycle of proteins, i.e., from their synthesis to their degradation, is critical for all organisms. Intracellular protein turnover is mediated by two evolutionarily conserved systems: the autophagy-lysosome pathway and the ubiquitinproteasome system (UPS). In the UPS, polyubiquitin-marked proteins are routed to a large protease complex, the 26S proteasome, for their proteolytic degradation [1]. The 26S proteasome is a multi-subunit protease complex consisting of a proteolytic core particle known as the 20S proteasome capped by one or two 19S regulatory particles [2]. The 20S can associate with the 11S complexes or PA200/Blm (proteasome activator 200 kDa/bleomycin resistance protein-10) [3]. The proteolytic activity of the proteasome is conveyed by the β-rings [4]. Proteasome genes are highly conserved and the Caenorhabditis elegans (C. elegans) orthologues of human 20S proteasome subunits are referred to as proteasome alpha subunits 1 to 7 (pas-1-7) and proteasome beta subunits 1 to 7 (pbs-1-7) [5]
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