Anti-PDIA6 monoclonal antibody, clone 4C5

The endoplasmic rcticulum (ER) plays an important role in maintenance of proteostasis through the unfolded protein response (UPR), which is strongly activated in most neurodegencrative disorders. UPR signalling pathways mediated by IRE1 alpha and ATF6 play a crucial role in the maintenance of ER homeostasis through the transactivation of an array of transcription factors. When activated, these transcription factors induce the expression of genes involved in protein folding and degradation with pro-survival effects. However, the specific contribution of these transcription factors to different neurodegenerative diseases remains poorly defined. Here, we characterised 44 target genes strongly influenced by XBP1 and ATF6 and quantified the expression of a subset of genes in the human post-mortem spinal cord from amyotrophic lateral sclerosis (ALS) cases and in the frontal and temporal cortex from frontotemporal lobar degeneration (FTLD) and Alzheimer's disease (AD) cases and controls. We found that IRE1 alpha-XBP1 and ATF6 pathways were strongly activated both in ALS and AD. In ALS, XBP 1 and ATF6 activation was confirmed by a substantial increase in the expression of both known and novel target genes involved particularly in co-chaperone activity and ER-associated degradation (ERAD) such as DNAJB9, SEL1L and OS9. In AD cases, a distinct pattern emerged, where targets involved in protein folding were more prominent, such as CANX, PDIA3 and PDIA6. These results reveal that both overlapping and disease-specific patterns of IRE1 alpha-XBP1 and ATF6 target genes are activated in AD and ALS, which may be relevant to the development of new therapeutic strategies.

Microglial activation in the central nervous system is a key event in the neuroinflammation that accompanies neurodegenerative diseases such as Alzheimers disease (AD). Among cytokines involved in microglial activation, amyloid beta (A beta) peptide is known to be a key molecule in the induction of diverse inflammatory products, which may lead to chronic inflammation in AD. However, proteomic studies of microglia in AD are limited due to lack of proper cell or animal model systems. In this study, we performed a proteomic analysis of A beta-stimulated human microglial cells using SILAC (stable isotope labeling with amino acids in cell culture) combined with LC-MS/MS. Results showed that 60 proteins increased or decreased their abundance by 1.5 fold or greater. Among these, ER-resident proteins such as SERPINH1, PDIA6, PDIA3, and PPIB were revealed to be key molecular biomarkers of human microglial activation by validation of the proteomic results by immunostaining, PCR, ELISA, and Western blot. Taken together, our data suggest that ER proteins play an essential role in human microglial activation by A beta and may be important molecular therapeutic targets for treatment of AD.