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PLoS One. 2014 Apr 21;9(4):e95863. doi: 10.1371/journal.pone.0095863. eCollection 2014.

LAMTOR2-mediated modulation of NGF/MAPK activation kinetics during differentiation of PC12 cells.

Author information

1
Division of Neurobiochemistry, CCB-Biocenter, Medical University of Innsbruck, Innsbruck, Austria.
2
Division of Neurobiochemistry, CCB-Biocenter, Medical University of Innsbruck, Innsbruck, Austria; Division of Cell Genetics, Department for Pharmacology and Genetics, Medical University of Innsbruck, Innsbruck, Austria.
3
Division of Cell Genetics, Department for Pharmacology and Genetics, Medical University of Innsbruck, Innsbruck, Austria.
4
Division of Cell Biology, CCB-Biocenter, Medical University of Innsbruck, Innsbruck, Austria.

Abstract

LAMTOR2 (p14), a part of the larger LAMTOR/Ragulator complex, plays a crucial role in EGF-dependent activation of p42/44 mitogen-activated protein kinases (MAPK, ERK1/2). In this study, we investigated the role of LAMTOR2 in nerve growth factor (NGF)-mediated neuronal differentiation. Stimulation of PC12 (rat adrenal pheochromocytoma) cells with NGF is known to activate the MAPK. Pharmacological inhibition of MEK1 as well as siRNA-mediated knockdown of both p42 and p44 MAPK resulted in inhibition of neurite outgrowth. Contrary to expectations, siRNA-mediated knockdown of LAMTOR2 effectively augmented neurite formation and neurite length of PC12 cells. Ectopic expression of a siRNA-resistant LAMTOR2 ortholog reversed this phenotype back to wildtype levels, ruling out nonspecific off-target effects of this LAMTOR2 siRNA approach. Mechanistically, LAMTOR2 siRNA treatment significantly enhanced NGF-dependent MAPK activity, and this effect again was reversed upon expression of the siRNA-resistant LAMTOR2 ortholog. Studies of intracellular trafficking of the NGF receptor TrkA revealed a rapid colocalization with early endosomes, which was modulated by LAMTOR2 siRNA. Inhibition of LAMTOR2 and concomitant destabilization of the remaining members of the LAMTOR complex apparently leads to a faster release of the TrkA/MAPK signaling module and nuclear increase of activated MAPK. These results suggest a modulatory role of the MEK1 adapter protein LAMTOR2 in NGF-mediated MAPK activation required for induction of neurite outgrowth in PC12 cells.

PMID:
24752675
PMCID:
PMC3994133
DOI:
10.1371/journal.pone.0095863
[Indexed for MEDLINE]
Free PMC Article

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