Ascl1-induced neuronal differentiation of P19 cells requires expression of a specific inhibitor protein of cyclic AMP-dependent protein kinase

作者：Huang Holly S; Turner David L; Thompson Robert C; Uhler Michael D^*

来源：Journal of Neurochemistry, 2012, 120(5): 667-683.

DOI：10.1111/j.1471-4159.2011.07332.x

• cyclic AMP
• differentiation
• shRNA

cAMP-dependent protein kinase (PKA) plays a critical role in nervous system development by modulating sonic hedgehog and bone morphogenetic protein signaling. In the current studies, P19 embryonic carcinoma cells were neuronally differentiated by expression of the proneural basic helix-loop-helix transcription factor Ascl1. After expression of Ascl1, but prior to expression of neuronal markers such as microtubule associated protein 2 and neuronal beta-tubulin, P19 cells demonstrated a large, transient increase in both mRNA and protein for the endogenous protein kinase inhibitor (PKI)beta. PKI beta-targeted shRNA constructs both reduced the levels of PKI beta expression and blocked the neuronal differentiation of P19 cells. This inhibition of differentiation was rescued by transfection of a shRNA-resistant expression vector for the PKI beta protein, and this rescue required the PKA-specific inhibitory sequence of the PKI beta protein. PKI beta played a very specific role in the Ascl1-mediated differentiation process as other PKI isoforms were unable to rescue the deficit conferred by shRNA-mediated knockdown of PKI beta. Our results define a novel requirement for PKI beta and its inhibition of PKA during neuronal differentiation of P19 cells.

• 出版日期2012-3