Immunoproteasome Deficiency Modifies the Alternative Pathway of NFκB Signaling

Abstract

Immunoproteasome is a protease abundant in immune cells and also present, albeit at lower concentrations, in cells outside the immune system. Recent evidence supports a novel role for the immunoproteasome in the cellular stress response potentially through regulation of NFκB signaling, which is the primary response to multiple stressors. The current study tests whether the Classical or Alternative Pathways are regulated by immunoproteasome following chronic TNFα exposure in cultured retinal pigment epithelial cells isolated from wild-type mice and mice deficient in one (LMP2, L2) or two (LMP7 and MECL-1, L7M1) immunoproteasome subunits. Assays were performed to assess the expression of NFκB responsive genes, the content and activity of NFκB transcription factors (p65, p50, p52, cRel, RelB), and expression and content of regulatory proteins (IκBα, A20, RPS3). Major findings include distinct differences in expression of NFκB responsive genes in both KO cells. The mechanism responsible for the altered gene expression could not be established for L7M1 since no major differences in NFκB transcription factor content or activation were observed. However, L2 cells exhibited substantially higher content and diminished activation of NFκB transcription factors associated with the Alternative Pathway and delayed termination of the Classical Pathway. These results provide strong experimental evidence supporting a role for immunoproteasome in modulating NFκB signaling.

Immunoproteasome subunit deficiency has no influence on the canonical pathway of NF-κB activation

“…While free IB␣ can indeed be degraded in a ubiquitin-independent fashion (Mathes et al, 2008) this degradation pathway might not be representative for the canonical ubiquitin-dependent degradation of NF-B-bound IB␣ via the 26S proteasome. In contrast, a recent report by Maldonado and colleagues found that activation of the canonical pathway was not affected in LMP2 −/− and L7M −/− cells while the alternative pathway seemed to be "aberrant" in LMP2 −/− cells (Maldonado et al, 2013). Furthermore, it has been proposed that the immunoproteasome has a higher capacity to clear ubiquitylated proteins accumulating after stimulation with IFN-␥ (Seifert et al, 2010).…”

“…Ultimately depending on the proteasome for activation, NF-B is an important transcription factor in inflammatory conditions. Still, previous studies concerned with the effect of immunoproteasome subunits on NF-B activation yielded contradictory results Faustman, 2000, 1999;Hensley et al, 2010;Kessler et al, 2000;Maldonado et al, 2013;Visekruna et al, 2006).…”

Therapeutic Potential of Immunoproteasome Inhibition in Duchenne Muscular Dystrophy

“…[6][7][8] Interestingly, local intramuscular inhibition of proteasome in murine and canine animal models of DMD leads to dystrophin rescue and DGC proteins expression. 7,[9][10][11] The immune cytokine interferon gamma (IFN-γ) can induce the expression of three other catalytically active β-subunits of the 20S proteasome -β1i (PSMB9), β2i (PSMB10), and β5i (PSMB8) to form the induced-proteasome, also referred as immunoproteasome (i-proteasome). 6 i-proteasome is involved in the generation of peptides presented by the MHC-I to the antigen receptors of cytotoxic T cells.…”

Denervation-Induced Activation of the Standard Proteasome and Immunoproteasome

“…Under basal conditions, the immunoproteasome exists at low concentrations, roughly contributing 5% to the total proteasome population in skeletal muscle [15]. The immunoproteasome is well-known for its role in immune function, specifically generation of antigenic peptides [16] as part of immune surveillance [17–19]. Interestingly, under catabolic conditions such as aging [20,21], muscular dystrophy [22], and denervation [11], the immunoproteasome increases, which suggests a link between atrophy and the immunoproteasome may exist in skeletal muscle.…”