Proteasome activator PA28c (REGc, Ki antigen) has recently been demonstrated to display anti-apoptotic properties via enhancing Mdm2-p53 interaction, thereby facilitating ubiquitination and down-regulation of the tumor suppressor p53. In this study we demonstrate a correlation between cellular PA28c levels and the sensitivity of cells towards apoptosis in different cellular contexts thereby confirming a role of proteasome activator PA28c as an anti-apoptotic regulator. We investigated the anti-apoptotic role of PA28c upon UV-C stimulation in B8 mouse fibroblasts stably overexpressing the PA28c-encoding PSME3 gene and upon butyrate-induced apoptosis in human HT29 adenocarcinoma cells with silenced PSME3 gene. Interestingly, our results demonstrate that PA28c has a strong influence on different apoptotic hallmarks, especially p53 phosphorylation and caspase activation. In detail, PA28c and effector caspases mutually restrict each other. PA28c is a caspase substrate, if PA28c levels are low. In contrast, PA28c overexpression reduces caspase activities, including the caspase-dependent processing of PA28c. Furthermore, overexpression of PA28c resulted in a nuclear accumulation of transcriptional active p53. In summary, our findings indicate that even in a p53-dominated cellular context, pro-apoptotic signaling might be overcome by PA28c-mediated caspase inhibition.
BackgroundPA28γ (also known as Ki, REG gamma, PMSE3), a member of the ubiquitin-and ATP-independent proteasome activator family 11S, has been proved to show proteasome-dependent and -independent effects on several proteins including tumor suppressor p53, cyclin-dependent kinase inhibitor p21 and steroid receptor co-activator 3 (SCR-3). Interestingly, PA28γ is overexpressed in pathological tissue of various cancers affecting e. g. breast, bowl and thyroids. Furthermore, anti-PA28γ autoantibodies have been linked to several autoimmune disorders. The aim of this study was to develop and evaluate a novel and sensitive PA28γ sandwich ELISA for the quantification of PA28γ serum levels in patients with cancer and autoimmune diseases for diagnostic and prognostic purposes.MethodsPA28γ-specific polyclonal antibodies and recombinant His-tagged PA28γ were purified and used to develop a sandwich ELISA for the detection of circulating PA28γ. With this new assay, PA28γ serum levels of patients with various cancers, rheumatoid arthritis (RA), Sjögren’s syndrome (SS), adult-onset Still’s disease (AOSD) and different connective-tissue diseases (CTD) were compared with healthy control subjects. Anti-PA28γ autoantibodies were additionally confirmed using a newly developed microbead assay.ResultsThe developed PA28γ sandwich ELISA showed a high specificity with a detection limit of 3 ng/ml. A significant up-regulation of circulating PA28γ was detected in the sera of patients with cancer, RA, SS and CTD. A significant correlation was observed dependent on age as well as anti-PA28γ autoantibody levels with circulating PA28γ protein levels. Furthermore, PA28γ serum levels showed a correlation with disease activity in patients with RA under treatment with the T-cell directed biological compound abatacept according to disease activity score 28 (DAS28) and erythrocyte sedimentation rate (ESR).ConclusionThe application of PA28γ as a novel biomarker for diagnostic purposes of a specific disease is limited, since elevated levels were observed in different disorders. However, the correlation with disease activity in patients with RA suggests a prognostic value, which needs to be addressed by further studies. Therefore our results show that PA28γ is a useful marker which should be included in studies related to novel treatments, e.g. abatacept.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2474-15-414) contains supplementary material, which is available to authorized users.
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