Overexpression of BETA2/NeuroD induces neurite outgrowth in F11 neuroblastoma cells.

BETA2/NeuroD, a basic helix-loop-helix (bHLH) transcription factor, has been shown to play important roles in the development of the nervous system and the maintenance and formation of pancreatic and enteroendocrine cells. The gain of function of BETA2/NeuroD in neurogenesis has been shown in Xenopus embryos. In this study, we investigated the neurogenic potential of BETA2/NeuroD using neuroblastoma cell line, F11, which could be induced to differentiate into neurons in the presence of cAMP. To induce or block the expression of BETA2/NeuroD, expression vectors for the full-length and a C-terminal deletion mutant of BETA2 were constructed and their transactivation potential was verified using reporter genes containing the insulin promoter sequences. Overexpression of BETA2 with full-length construct induced neurite outgrowth in F11 cells in the absence of cAMP. In contrast, the C-terminal deletion mutant, BETA2(1--233), which has dominant negative activity, inhibited neurite outgrowth induced by cAMP in F11 cells. These results indicate that BETA2/NeuroD plays an important role in terminal differentiation of neuroblastoma cells. They also imply that BETA2/NeuroD or related bHLH factors plays an essential role for differentiation of F11 neuroblastoma cells.

Neurite outgrowth induced by cyclic AMP can be modulated by the alpha subunit of Go.

Although abundant Go has been found in nervous tissues and it has been implicated in neuronal differentiation, the mechanism of how Go modulates neuronal differentiation has not been defined. Here, we report that the alpha subunit of Go (alphao) modulates neurite outgrowth by interfering with the signaling pathway initiated by cyclic AMP (cAMP). In F11 cells, cAMP induced neurite outgrowth and activated cAMP-responsive element binding protein (CREB). Specific inhibition of cAMP-dependent protein kinase reduced both CREB activity and neurite outgrowth (NOG). Interestingly, cAMP reduced phosphorylation of extracellular signal-regulated kinase (Erk). Neither a dominant negative form nor an active form of Ras altered neurite outgrowth. Expression of alphao (alphao(wt)) decreased the average length of neurites but increased the number of neurites per cell. An active mutant, alphaoQ205L, which lost GTPase activity and thus could not bind to Gbetagamma, gave similar results, suggesting that the effect of alphao is not mediated through Gbetagamma. Expression of ao(wt) or alphaoQ205L also prohibited CREB activation. Thus, activation of Erk may not be essential for neuronal differentiation in F11 cells and alphao may cause changes in NOG by inhibiting CREB activation.