| Abstract
| - Background. The multidomain protein p62/SQSTM1 modulates important signalling pathways responsible for osteoclastogenesis, cell proliferation and polarity. Furthermore, it also regulates protein degradation by autophagy, the ubiquitin proteasome system or sequestration. Although there is a link between p62 mutations and Paget’s disease of bone, it is unclear how p62 acts as a signaling scaffold between the different pathways and how the single domains coordinate the protein functions and link different processes during chronic inflammation. Methods. Therefore, we generated mice carrying a truncated form of p62, which lacks the binding sites for TRAF6 and aPKC, but still contains a functional ubiquitin binding site (p62Delta69-251). We characterized these mice and crossed them with arthritic hTNFtg animals to study the effects of the p62 signaling domain on the onset, severity and histologicals changes of a TNFalpha dependent arthritis. Isolated cells of these mice were analysed for alterations in their differentiation capacity, ROS production and MAPK signaling. Results. The truncated p62Delta69-251 protein was ubiquitously expressed and was still able to oligomerize and to bind LC3, ubiquitinated proteins, and ERK1 whereas TRAF6 binding was missing. Cells of homozygote p62Delta/69-251mice showed an increased production of ROS and elevated levels of autophagy-related acidic membrane vesicles. Stimulation of p62Delta69-251 cells with the autophagy-inhibitor Bafilomycin and the proteasome-inhibitor MG132 still resulted in an accumulation of p62 and aggregate formation. Furthermore, the lack of the signaling module was sufficient to produce an obese phenotype accompanied by the enlargement of organs, e.g. liver and kidney. PET/CT studies revealed an increased bone metabolism and BMDMs of p62Delta69-251 mice showed a significantly increased osteoclastogenesis in vitro, particularly when stimulated with TNFalpha. WB analysis revealed an increased TNFalpha- induced p38 phosphorylation in cells from p62Delta69-251 compared to wt mice. Crossing of p62Delta69-251mice with hTNFtg animals resulted in a dramatic increase in the severity of joint damage conceivable by an increased number and size of osteoclasts. Interestingly, p62Delta69-251/+hTNFtg mice showed a significant increased weight at week 12 compared to hTNFtg mice. Conclusions. The loss of p62 signaling domains has important consequences both for metabolic activation and for osteoclastogenesis under inflammatory conditions. Disclosure of Interest. None Declared
|
