Interleukin-32 alpha modulates promyelocytic leukemia zinc finger gene activity by inhibiting protein kinase C epsilon-dependent sumoylation

Abstract

Interleukin-32 (IL-32) is a proinflammatory cytokine. However, there is growing evidence that IL-32 also plays a mediatory role intracellularly. In this study, we present evidence that IL-32 alpha modifies and inhibits promyelocytic leukemia zinc finger (PLZF), a sequence-specific transcriptional regulator that regulates the expression of a subset of interferon (IFN)-stimulated genes (ISGs). We screened IL-32 alpha-interacting proteins in a human spleen cDNA library using the yeast two-hybrid assay, and investigated the functional relevance of the interaction between IL-32 alpha and PLZF. We demonstrated that IL-32 alpha interacts with protein kinase C (PKC)delta and PKC epsilon in a phorbol 12-myristate 13-acetate (PMA) dependent way, and that PKC epsilon regulates the interaction of IL-32 alpha with PLZF. We verified the involvement of PKC epsilon in the interaction between these proteins by using various PKC inhibitors. PLZF is known to be modified by small ubiquitin-like modifier (SUMO)-1, but it is unclear whether SUMO-2 conjugation of PLZF occurs. We showed that IL-32 alpha inhibited SUMO-2-conjugation of PLZF. Further, we demonstrated that sumoylated PLZF decreased when IL-32 alpha was co-expressed. PKC epsilon affected the sumoylation of PLZF only in the presence of IL-32 alpha because PKC inhibitor treatment did not reduce PLZF sumoylation in the absence of IL-32 alpha. We finally investigated whether IL-32 alpha-mediated inhibition of PLZF sumoylation affected the transcriptional activity of PLZF, and demonstrated that the inhibition of sumoylation of PLZF by IL-32 alpha down-regulated ISGs induced by PLZF. Together, our data suggest that IL-32 alpha associates with PLZF and PKC epsilon, and then inhibits PLZF sumoylation, resulting in suppression of the transcriptional activity of PLZF. (C) 2014 Elsevier Ltd. All rights reserved.