Cellular functions of cyclophilin A in actin stability and cyclosporine A induced-gingival overgrowth

Cyclophilin A (CypA) is a peptidyl-prolyl cis-trans isomerase that is involved in multiple signaling events in eukaryotic cells either by acting as a catalyst for prolyl bond isomerization or by forming stoichiometric complexes with target proteins. CypA was originally identified as a cellular binding protein for the immunosuppressant Cyclosporine A (CsA) which has gingival overgrowth as one of its potential side effects. Despite the fact that CypA is a ubiquitous cellular protein, our understanding of its cellular functions is limited. Therefore, we examined the role of CypA in the regulation of actin polymerization, and since CypA was originally identified as the major target for CsA, we also examined CsA's role in gingival overgrowth.;In our experiments, we demonstrate that knockdownCypA expression using RNAi in U2OS cells resulted in disruption of the F-actin structure, as well as decreased anchorage-independent growth, proliferation, and migration. Wild-type U2OS cells treated with cyclosporine A (CsA), a peptidyl-prolyl isomerase inhibitor, displayed the same phenotype as knockdown CypA cells, suggesting that the isomerase activity of CypA is required to maintain a normal phenotype. In vitro and in vivo binding assays revealed that CypA binds to N-WASP, which functions in the nucleation of actin via the Arp2/3 complex. Pulse-chase labeling study indicated an enhanced degradation of N-WASP in cell lacking CypA, suggesting that CypA is required for stabilizing N-WASP to form a N-WASP/Arp2/3 complex for the nucleation/initiation of F-actin polymerization.;In our examination of the molecular mechanisms of CsA-induced gingival overgrowth, we show that MMP's-1, 2 and 9 expressions are inhibited by Cyclosporine (CsA) treatment. The decrease in MMP-1 expression by CsA appears to be due to the destabilization of MMP-1 mRNA while the decrease in MMP-2 was found to be limited to the secreted protein expression suggesting that CsA's affects are limited to the secretion of MMP-2. CsA also appeared to inhibit NFkappaB, which is known to transcriptionally activate MMP-9 expression. These results suggest that MMP-1, -2, and -9 all may contribute to the pathogenesis of CsA induced gingival overgrowth by different mechanisms.