The deubiquitinase Usp27x as a novel regulator of cFLIPL protein expression and sensitizer to death-receptor-induced apoptosis

Abstract

Death receptors are transmembrane proteins that can induce the activation of caspase-8 upon ligand binding, initiating apoptosis. Recent work has highlighted the great molecular complexity of death receptor signalling, in particular through ubiquitination/deubiquitination. We have earlier defined the deubiquitinase Ubiquitin-Specific Protease 27x (Usp27x) as an enzyme capable of stabilizing the pro-apoptotic Bcl-2 family member Bim. Here, we report that enhanced expression of Usp27x in human melanoma cells leads to the loss of cellular FLICE-like inhibitory protein (cFLIP) and sensitizes to Tumor necrosis factor receptor 1 (TNF-R1) or Toll-like receptor 3 (TLR3)-induced extrinsic apoptosis through enabling enhanced processing of caspase-8. The loss of cFLIPL upon overexpression of Usp27x was not due to reduced transcription, could be partially counteracted by blocking the ubiquitin proteasome system and was independent of the known cFLIPL destabilizing ubiquitin E3-ligases Itch and DTX1. Instead, Usp27x interacted with the E3-ligase TRIM28 and reduced ubiquitination of TRIM28. Reduction of cFLIPL protein levels by Usp27x-induction depended on TRIM28, which was also required for polyI:C-induced cell death. This work defines Usp27x as a novel regulator of cFLIPL protein expression and a deubiquitinase in fine tuning death receptor signalling pathways to execute apoptosis.

USP27Xvariants underlying X-linked intellectual disability disrupt protein function via distinct mechanisms

“…To investigate how USP27X variants affect USP27X protein localization and stability, we introduced human USP27X into Usp27x −/y mouse embryonic stem cells (ESCs) (Atanassov et al, 2016). WT USP27X is localized mainly to the nucleus with some cytoplasmic expression (Fig 2A ) as previously described (Atanassov et al, 2016;Dold et al, 2022). Similar to the WT, XLID105 mutants largely localize to the nucleus (Fig 2A).…”

“…Inserts in the USP domain can also contribute to specificity and regulation of USP27X catalytic activity (Tencer et al, 2016). Particularly for USP27X, an amino terminal extension resulting from a noncanonical translation start that occurs in some cell types (Atanassov et al, 2016;Dold et al, 2022) may play a role in regulation of catalytic activity. Furthermore, we performed in vitro assays using recombinant proteins to assess the impact of variants on USP27X catalysis.…”

“…Ubiquitin-specific protease 27 X-linked (USP27X) is a DUB that has been associated to important cellular functions such as cell proliferation and immune response. (Atanassov et al, 2016;Weber et al, 2016;Dong et al, 2018;Guo et al, 2019;Lambies et al, 2019;Seo et al, 2020;Alam et al, 2022;Dold et al, 2022). Furthermore, USP27X targets the developmental regulator HES1 to regulate neuronal differentiation (Kobayashi et al, 2015).…”

The Emerging Role of Deubiquitinases in Cell Death

“…The overexpression of DUB ubiquitin-specific protease 27 X-Linked (USP27X) leads to the loss of the CFLAR L protein and sensitizes extrinsic apoptosis in melanoma cells. USP27X interacts with the E3-ligase tripartite motif containing 28 (TRIM28) and reduces the ubiquitination of E3-ligases TRIM28, but not ITCH and DTX1, which leads to decreased CFLAR protein [ 17 ]. The DUB ubiquitin-specific protease 2 (USP2) also promotes MAPK8-mediated and TNF-induced activation of ITCH and subsequent CFLAR L/S degradation.…”

Death receptor 3 is involved in preeclampsia through regulating placental trophoblast cell physiology by inactivating the PI3K/AKT pathway

“… 8 , 9 Under pathological conditions, once this balance is broken, it will eventually lead to apoptosis. 10 There are three pathways of apoptosis, including death receptor induced apoptosis, 11 mitochondrial permeability induced apoptosis, 12 and endoplasmic reticulum pathway. 13 DR‐3, a member of TNFRSF, contains a death domain with proapoptotic effects and is able to activate caspase 8 and NF‐κB signals apoptosis by signaling cell survival.…”