Abstract
IL-33 is a nuclear cytokine from the IL-1 family that plays important roles in health and disease. Extracellular IL-33 activates a growing number of target cells, including group 2 innate lymphoid cells, mast cells and regulatory T cells, but it remains unclear whether intracellular nuclear IL-33 has additional functions in the nucleus. Here, we used a global proteomic approach based on high-resolution mass spectrometry to compare the extracellular and intracellular roles of IL-33 in primary human endothelial cells, a major source of IL-33 protein in human tissues. We found that exogenous extracellular IL-33 cytokine induced expression of a distinct set of proteins associated with inflammatory responses in endothelial cells. In contrast, knockdown of endogenous nuclear IL-33 expression using two independent RNA silencing strategies had no reproducible effect on the endothelial cell proteome. These results suggest that IL-33 acts as a cytokine but not as a nuclear factor regulating gene expression in endothelial cells.
Highlights
Nuclear localization of IL-33 is a fundamental property of the protein that has been observed in all producing cells, both in human and mouse tissues[21,22,23]
For defining expression changes in pairwise comparisons, a Student t-test was calculated in the Perseus software and a permutation-based approach was used to account for multiple testing and adjust the final false discovery rate at 5%. Based on this stringent selection procedure, 56 proteins were reproducibly found to exhibit a significant variation in endothelial cells stimulated with IL-3395–270 during 24 h (Fig. 2a,b and Supplementary Table 1). 17 proteins were already modulated after 6 h of stimulation with IL-3395–270 (Supplementary Fig. 2)
We addressed an important question in IL-33 biology, the relative roles of extracellular IL-33 cytokine and endogenous intracellular nuclear IL-33 in the regulation of protein expression
Summary
Nuclear localization of IL-33 is a fundamental property of the protein that has been observed in all producing cells, both in human and mouse tissues[21,22,23]. To date, no large scale study has been performed to demonstrate a global role of endogenous nuclear IL-33 in the regulation of gene or protein expression. Knockdown of endogenous nuclear IL-33 using two independent RNA silencing strategies did not induce reproducible changes in the endothelial cell proteome. Together, these proteome-wide analyses do not support the previous proposal that IL-33 is a dual function protein. These proteome-wide analyses do not support the previous proposal that IL-33 is a dual function protein They suggest that the main purpose of IL-33 nuclear localization is the regulation of its extracellular cytokine activity through nuclear sequestration, rather than the regulation of gene or protein expression
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