The Role Of Scaffold RACK1in The Production Of Proinflammatory Cytokines

RACK1(Receptor for Activated C Kinase1,36kDa) consists of seven highly conserved Trp-Asp40repeats and is therefore a scaffold protein. It is ubiquitiously expressed and has been implicated in diverse cellular activities including embryonic development, cell growth, differentiation, migration, apoptosis, and circadian rhythm clock. Several groups have reported that RACK1regulates the activity of multiple intracellular signaling molecules (such as JNK and GSK3β which might affect the production of proinflammatory cytokines. However, it remains unknown the role of RACK1in proinflammatory cytokine expression.Objective:In this work, we investigated the effects of RACK1knockdown on the production of proinflammatory cytokines and the underlying mechanisms with human monocytes THP1and human primary macrophages as the models.Methods:Cells were transfected with RACK1shRNA by lentivirus-mediated RNA interference, then we detected the expression of proinflammatory cytokines by Elisa and the protein levels by Immunoblot. Other factors were detected by RT-PCR、FACS and so on.Results:We found that LPS-induced IL-6/TNF-a production from THP1human monocytes or human primary macrophages significantly decreased under the condition of RACK1knockdown. RACK1knockdown in THP1cells led to impaired proliferation without inducing terminal differentiation. Further exploration revealed that RACK1not only affected the classic pro-inflammatory JNK and p38MAPK signaling pathways, but also played an important role in the activation of NFκB signaling pathway. RACK1knockdown resulted in reduced phosphorylation of IKK in response to LPS. Then, we found that RACK1affected the protein levels of IKK upstream moleculars IRAK1, TAK1, and TRAF6. RACK1knockdown did not reduce the mRNA levels and the half lives of IRAK1, TAK1, and TRAF6. However, PKCβⅡ specific inhibitor treatment led to reduced IRAK1protein levels.Conclusion:RACK1can lead to the activation of JNK, p38and NF-κB pathway by maintaining the level of IRAK1、TRAF6、TAK1expression, and which finally influences the expression of IL-6/TNF-a. Our results indicated that enhanced PKCβⅡ activity might be involved. There are14pictures,0table and27references in this passage.