A novel cyathane diterpene, cyrneine A, induces neurite outgrowth in a Rac1-dependent mechanism in PC12 cells

We examined the effects of cyrneine A, a novel diterpene isolated from the mushroom Sarcodon cyrneus, on morphology of rat pheochromocytoma cells (PC12). In response to cyrneine A, PC12 cells extended their neurites, an effect partially blocked by the extracellular signal-regulated kinase (ERK) kinase inhibitor PD98059, but not by the protein kinase C inhibitor GF109203X, nor the phosphatidylinositol-3- kinase inhibitor wortmannin. Cyrneine A did not activate ERK at any of the time points tested (5–120 min), indicating that only the basal activity of ERK is required for cyrneine A-induced neurite outgrowth. As transcriptional regulation is required for neurite extension, the activity of three major transcription factors was determined. Cyrneine A enhanced activation of the transcription factors activator protein-1 (AP-1) and nuclear factor-κB, but not CREB, and this was accompanied by enhanced c-fos expression. Moreover, we determined the role of Rac1, a small GTPase protein of the Rho family that regulates actin dynamics, in cyrneine A-induced neurite outgrowth. Treatment with cyrneine A led to actin translocation and subsequently, to accumulation of F-actin at the tip of neurites. Rac1 activity was increased by cyrneine A and expression of a dominant-negative Rac1 mutant significantly inhibited the cyrneine A-induced extension of neurites. These results suggest that cyrneine A induces neurite outgrowth in a Rac1-dependent mechanism.