Catalytic Subunits Of Immunoproteasome Research Articles

Upregulation of immunoproteasome PSMB8 is associated with Parkinson’s disease

BackgroundImmunoproteasome, a part of ubiquitin–proteasome system, is involved in immune response as well as protein degradation. However, the relationship between immunoproteasome and Parkinson's disease (PD) was not evaluated clearly. We hypothesized that the shift of immunoproteasome attributes to PD pathogenesis due to its role in inflammation and protein homeostasis. ObjectiveTo determine whether immunoproteasome in peripheral blood mononuclear cells (PBMC) and brain is expressed differently between patients with PD and healthy controls (HC). MethodsBlood samples were collected from 19 HC to 40 patients with PD of comparable ages. Peripheral blood mononuclear cells were isolated and followed by RT-qPCR to measure the mRNA levels of three catalytic subunits of immunoproteasome, namely, PSMB8, PSMB9, and PSMB10. Then, the protein levels of each subunit were measured by western blot. Finally, we confirmed the altered immunoproteasome subunit in the post-mortem human brain of PD. ResultsIn PBMCs, PSMB8 mRNA expression of PD group significantly increased compared to HC (p = 0.004), whereas PSMB9 and PSMB10 mRNA were not different between the PD and HC. The ratio of PSMB10 and PSMB8 mRNA (PSMB10/8 ratio) also reflected the significant difference between the PD and HC (p = 0.002). The PSMB10/8 ratio was well correlated with the UPDRS total and Part III score in the early stage of PD (Hoehn and Yahr ≤2.5) or drug-naïve PD subgroups. In terms of the protein level of immunoproteasome subunits in PBMCs, the increase of PSMB8 protein was observed in PD compared to HC (p = 0.0009), while PSMB9 and PSMB10 were not different between groups. Finally, we confirmed that immunoproteasome PSMB8 was expressed abundantly in the postmortem PD brain compared with normal control. ConclusionOur novel findings implicate that immunoproteasome PSMB8 is engaged in PD pathomechanism.

Evolution of dimorphisms of the proteasome subunit beta type 8 gene (PSMB8) in basal ray-finned fish

The proteasome subunit beta type 8 (PSMB8) gene encodes a catalytic subunit of immunoproteasome that plays a central role in the processing of antigenic peptides presented by major histocompatibility complex class I molecules. The A- and F-type alleles defined by the 31st amino acid residue determining cleaving specificity have been identified from ray-finned fish, amphibia, and reptiles. These two types show extremely long-term trans-species polymorphism in Polypteriformes, Cypriniformes, and Salmoniformes, suggesting the presence of very ancient lineages termed A and F. To elucidate the evolution of the PSMB8 dimorphism in basal ray-finned fish, we analyzed Pantodon buchholzi (Osteoglossiformes), seven species of Anguilliformes, and Hypomesus nipponensis (Osmeriformes). Both A and F lineage sequences were identified from P. buchholzi and H. nipponensis, confirming that these two lineages have been conserved by basal ray-finned fish. However, both the A- and F-type alleles found in Anguilliformes species belonged to the F lineage irrespective of their types. This apparently suggests that the A lineage was lost in the common ancestor of Anguilliformes, and recovery of the A type within the F lineage occurred in Anguilliformes. The apparent loss of the F lineage and recovery of the F type within the A lineage have already been reported from tetrapods and higher teleosts. However, this is the first report on the reverse situation and reveals the dynamic evolution of the PSMB8 dimorphism.

CDNA microarray analysis to compare HCV subgenomic replicon cells with their cured cells

The hepatitis C virus (HCV) replicon system carrying autonomously replicating HCV subgenomic RNA in human hepatocyte cells is a potent tool for basic studies of HCV, such as viral replication and drug development. Recently, we developed two HCV subgenomic replicons (50-1 and 1B-2R1) derived from two HCV strains, 1B-1 and 1B-2, respectively. Since the expression of HCV proteins is thought to affect the host cells’ gene expression profiles, we attempted to identify target genes of HCV proteins using microarray analysis (9970 genes) by comparing 50-1 and 1B-2R1 replicon cells with their “cured cells”, from which the replicons had been eliminated by prolonged treatment with interferon-α. The results showed that HCV replicons could have a variety of expression profiles in human hepatocytes. The results also showed that 2 and 6 genes were commonly up-regulated (more than 2.0-fold) and down-regulated (less than 0.50-fold), respectively, in both 50-1 and 1B-2R1 replicon cells compared with their cured cells. The differential expression profiles of genes selected by the microarray analysis were confirmed with standard RT-PCR and real-time LightCycler PCR. It was noteworthy that the commonly down-regulated genes contained large multifunctional proteases 2 and 7, which are known as catalytic subunits of immunoproteasome, and serine proteinase inhibitor clade C. Our microarray analysis demonstrated that HCV subgenomic replicons can change the gene expression profiles of host cells, and it allowed us to compile the first list of genes that the replicons transcriptionally regulate.