Sustained GRK2-dependent CREB activation is essential for α-adrenergic receptor-induced PC12 neuronal differentiation.

Many aspects of neuronal development, such as neuronal survival and differentiation, are regulated by the transcription factor cAMP-response element-binding protein (CREB). We have previously reported that α-adrenergic receptors (ARs), members of the G protein-coupled receptor (GPCR) superfamily, induce neuronal differentiation of rat pheochromocytoma (PC)-12 cells in a subtype-specific manner, i.e. α<α<α. Herein, we sought to investigate CREB`s involvement in this αAR-dependent neurogenic process. We used a combination of gene reporter assays and immunoblotting analysis, coupled with co-immunoprecipitation and morphological analysis, in transfected PC12 cell lines. Chronic α- or α-AR activation results in sustained CREB activation, which is both necessary and sufficient for neuronal differentiation of PC12 cells expressing these two αARs. In contrast, chronic α activation only leads to transient CREB activation, insufficient for PC12 neuronal differentiation. However, upon CREB overexpression, α-AR triggers neuronal differentiation similarly to α- or α-ARs. Importantly, NGF (Nerve Growth Factor)`s TrkA receptor transactivation is essential for the sustained activation of CREB by all three α subtypes in PC12 cells, whereas protein kinase A (PKA), the prototypic kinase that phosphorylates CREB, is not. Instead, TrkA-activated GPCR-kinase (GRK)-2 mediates the sustained CREB activation during αAR-induced neuronal differentiation of PC12 cells. In conclusion, catecholaminergic-induced neuronal differentiation of PC12 cells through αARs uses a non-canonical pathway involving TrkA transactivation and subsequent GRK2-dependent, sustained phosphorylation/activation of CREB. These findings provide novel insights into catecholaminergic neurogenesis and suggest that αAR agonists, combined with NGF analogs or GRK2 stimulators, may exert neurogenic/neuroprotective effects.