γ-Enolase enhances Trk endosomal trafficking and promotes neurite outgrowth in differentiated SH-SY5Y cells

Background: Neurotrophins can activate multiple signalling pathways in neuronal cells through binding to their cognate receptors, leading to neurotrophic processes such as cell survival and differentiation. gamma-Enolase has been shown to have a neurotrophic activity that depends on its translocation towards the plasma membrane by the scaffold protein gamma 1-syntrophin. The association of gamma-enolase with its membrane receptor or other binding partners at the plasma membrane remains unknown. Methods: In the present study, we used immunoprecipitation and immunofluorescence to show that gamma-enolase associates with the intracellular domain of the tropomyosin receptor kinase (Trk) family of tyrosine kinase receptors at the plasma membrane of differentiated SH-SY5Y cells. Results: In differentiated SH-SY5Y cells with reduced expression of gamma 1-syntrophin, the association of gamma-enolase with the Trk receptor was diminished due to impaired translocation of gamma-enolase towards the plasma membrane or impaired Trk activity. Treatment of differentiated SH-SY5Y cells with a gamma-Eno peptide that mimics gamma-enolase neurotrophic activity promoted Trk receptor internalisation and endosomal trafficking, as defined by reduced levels of Trk in clathrin-coated vesicles and increased levels in late endosomes. In this way, gamma-enolase triggers Rapt activation, which is required for neurotrophic activity of gamma-enolase. Additionally, the inhibition of Trk kinase activity by K252a revealed that increased SH-SY5Y cell survival and neurite outgrowth mediated by the gamma-Eno peptide through activation of signalling cascade depends on Trk kinase activity. Conclusions: These data therefore establish the Trk receptor as a binding partner of gamma-enolase, whereby Trk endosomal trafficking is promoted by gamma-Eno peptide to mediate its neurotrophic signalling.

Automated summary

Neurotrophins like gamma-enolase activate various signaling pathways through specific receptors, promoting neurotrophic processes in neuronal cells. This study found that gamma-enolase associates with the intracellular domain of Trk receptors at the plasma membrane, and its neurotrophic activity is dependent on this association. Additionally, a peptide mimicking gamma-enolase activity promoted Trk receptor internalization and endosomal trafficking, demonstrating the role of Trk kinase activity in mediating neurotrophic signaling.