Exchange Protein Directly Activated by cAMP (EPAC) Regulates Neuronal Polarization through Rap1B.
Acquisition of neuronal polarity is a complex process involving cellular and molecular events. The second messenger cAMP is involved in axonal specification through activation of protein kinase A. However, an alternative cAMP-dependent mechanism involves the exchange protein directly activated by cAMP (EPAC), which also responds to physiological changes in cAMP concentration, promoting activation of the small Rap GTPases. Here, we present evidence that EPAC signaling contributes to axon specification and elongation. In primary rat hippocampal neurons, EPAC isoforms were expressed differentially during axon specification. Furthermore, 8-pCPT, an EPAC pharmacological activator, and genetic manipulations of EPAC in neurons induced supernumerary axons indicative of Rap1b activation. Moreover, 8-pCPT-treated neurons expressed ankyrin G and other markers of mature axons such as synaptophysin and axonal accumulation of vGLUT1. In contrast, pharmacological inhibition of EPAC delayed neuronal polarity. Genetic manipulations to inactivate EPAC1 using either shRNA or neurons derived from EPAC1 knock-out (KO) mice led to axon elongation and polarization defects. Interestingly, multiaxonic neurons generated by 8-pCPT treatments in wild-type neurons were not found in EPAC1 KO mice neurons. Altogether, these results propose that EPAC signaling is an alternative and complementary mechanism for cAMP-dependent axon determination.


CONCLUSIONS
This study identifies the guanine exchange factor responsible for Rap1b activation during neuronal polarization and provides an alternate explanation for cAMP-dependent acquisition of neuronal polarity.
Laboratory of Cell and Neuronal Dynamics, Department of Biology, Faculty of Sciences, Universidad de Chile, 7800024 Santiago, Chile, Department of Molecular Pharmacology, University of Groningen, The Netherlands.
Pablo
http://orcid.org/0000-0001-9280-1590
P
Muñoz-Llancao
Laboratory of Cell and Neuronal Dynamics, Department of Biology, Faculty of Sciences, Universidad de Chile, 7800024 Santiago, Chile.
Daniel R
http://orcid.org/0000-0002-1761-1217
DR
Henríquez
Laboratory of Cell and Neuronal Dynamics, Department of Biology, Faculty of Sciences, Universidad de Chile, 7800024 Santiago, Chile.
Carlos
C
Wilson
Laboratory of Cell and Neuronal Dynamics, Department of Biology, Faculty of Sciences, Universidad de Chile, 7800024 Santiago, Chile.
Felipe
F
Bodaleo
Department of Medical Physiology, University Medical Center Groningen, University of Groningen, The Netherlands.
Erik W
http://orcid.org/0000-0002-5058-9648
EW
Boddeke
Inserm UMR-1048, Institut des Maladies Métaboliques et Cardiovasculaires, 31432 Toulouse, France, and Université de Toulouse III, Paul Sabatier, 31062 Toulouse, France.
Frank
F
Lezoualc'h
Department of Molecular Pharmacology, University of Groningen, The Netherlands.
Martina
M
Schmidt
Laboratory of Cell and Neuronal Dynamics, Department of Biology, Faculty of Sciences, Universidad de Chile, 7800024 Santiago, Chile, chrgonza@uchile.cl.
Christian
C
González-Billault
D000818:Animals; D001369:Axons / Q000378:metabolism; D016764:Cell Polarity / Q000502:physiology*; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D006624:Hippocampus / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D020033:Protein Isoforms / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000502:physiology; D020668:rap GTP-Binding Proteins / Q000378:metabolism*
D020662:Guanine Nucleotide Exchange Factors; D020033:Protein Isoforms; C526973:Rapgef3 protein, rat; C492135:Rap1b protein, rat; D020668:rap GTP-Binding Proteins
EPAC signaling; Rap1b signaling; axon; axon initial segment; cytoskeleton; neuronal polarity