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DJ Biros, AW Taylor; Altered Protein Ubiquitination In Regulatory T Cells Induced By The Ocular Immunosuppressive Cytokine -Melanocyte Stimulating Hormone . Invest. Ophthalmol. Vis. Sci. 2002;43(13):1536.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Recently we have reported that the immunosuppressive neuropeptide α-melanocyte stimulating hormone (α-MSH) induces the activation of regulatory T cells (Tr cells). This induction requires the T cells to be primed and restimulated through their T cell receptor in the presence of α-MSH. The lymphokine profile of the α-MSH-induced Tr cells showed suppressed IFN-γ and enhanced TGF-ß1 production; however, the levels of mRNA expression for both of these lymphokines did not change. Therefore, to evaluate the mechanisms of α-MSH induction of Tr cells, we examined total RNA through microarray gene expression comparing α-MSH-induced Tr cells with untreated T cells. Methods: In the presence of α-MSH primed T cells were activated in vitro by antigen presenting cells (APC) pulsed with antigen. Total RNA was collected 24 hours post activation and was examined through Affymetrix microarray gene analysis. For comparison, mRNA was also collected from untreated primed T cells activated by APC and antigen. Based on the microarray results, immunoblotting for ubiquitinated proteins was done on cell lysates of 48-hour cultures of α-MSH treated and untreated primed T cells activated by APC and antigen. Results: We found significant changes in mRNA of 371 genes with 9 genes significantly increased in the α-MSH treated T cells. Among the genes with an increased message was f-box only protein 15 (3 fold increase). F-box proteins enable Skp1/Cdc53/F-box (SCF) complexes to ubiquitinate specific proteins for degradation. Immunoblots of ubiquitinated proteins from 48-hour-old cultures of α-MSH induced Tr cells revealed a 2-fold increase in 42-210 kDa ubiquitinated proteins and a 2.5-fold decrease in free ubiquitin (8.7 kDa). Conclusion: The results suggest the observed α-MSH suppression of inflammation by effector T cells is through altered protein ubiquitination. This implies that a significant effect of the ocular microenvironment on T cell functionality is a potential post-translational modification of inflammatory factors preventing their secretion.
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