Molecular components involved in MAPK activation mediated by CRH in the central nervous system.

SILBERSTEIN, S.; BONFIGLIO, J. J.; GIACOMINI, D.; REFOJO, D.; HOLSBOER, F.; ARZT, E.

Buenos Aires - Argentina

Congreso; III Iberoamerican Congress on Neuroimmunomodulation.; 2009

Corticotropin-releasing hormone (CRH) plays a key role in stress. Disregulation of CRH receptor 1 (CRHR1) in limbic structures is associated to anxiety/depression. We previously demonstrated that icv CRH administration in mice leads, through CRHR1, to ERK1/2 activation in specific limbic areas. We decided to characterize the CRH signaling pathways in hippocampal cells, and to develop studies aimed to identify physiological targets of the CRH action. We generated HT22 cell lines stably overexpressing CRHR1, in which CRH stimulates CRE-LUC and increases pERK1/2 levels in a dose- and time-dependent manner. We characterized the B-Raf interactome, the MAPKKK of the B-Raf-MEK1/2-ERK1/2 cascade in neurons. B-Raf co-immunoprecipitates were resolved by SDS/PAGE and protein bands were identified by mass spec. We identified vimentin associated to B-Raf. Since B-Raf and 14-3-3 proteins are abundant in brain, we investigated its interaction. Our results show that 14-3-3, vimentin and B-Raf form a complex in CRH-stimulated cells. By immunofluorescence, we started the temporo-spatial analysis of CRH activated ERK and its subcellular localization in relation to CRHR1 internalization. CRH-activated pERK1/2 was mainly located in the cytosol. Our results contribute to the understanding of the physiological relevance of the identified complex in CRH signaling in hippocampal cells