Ubiquitination of TrkA by Nedd4-2 regulates receptor lysosomal targeting and mediates receptor signaling

Abstract

The nerve growth factor receptor TrkA (tropomyosin-related kinase receptor) participates in the survival and differentiation of several neuronal populations. The C-terminal tail of TrkA contains a PPXY motif, the binding site of the E3 ubiquitinligase Nedd4-2 (neural precursor cell expressed, developmentally down-regulated 4-2). In order to analyze the role of Nedd4-2 ubiquitination on TrkA function, we generated three TrkA mutants, by introducing point mutations on conserved hydrophobic amino acids – Leu784 and Val790 switched to Ala. TrkA mutants co-localized and co-immunoprecipitated more efficiently with Nedd4-2 and consequently a strong increase in the basal multimonoubiquitination of the mutant receptors was observed. In addition, we found a decrease in TrkA abundance because of the preferential sorting of mutant receptors towards the late endosome/lysosome pathway instead of recycling back to the plasma membrane. Despite the reduction in the amount of membrane receptor caused by the C-terminal changes, TrkA mutants were able to activate signaling cascades and were even more efficient in promoting neurite outgrowth than the wild-type receptor. Our results demonstrate that the C-terminal tail hydrophobicity of TrkA regulates Nedd4-2 binding and activity and therefore controls receptor turnover. In addition, TrkA multimonoubiquitination does not interfere with the activation of signaling cascades, but rather potentiates receptor signaling leading to differentiation.

The work was supported by the Instituto de Salud Carlos III – Ministerio de Sanidad y Consumo (FIS) (PI04/2537 and PS09/ 00140) to ML, the Ministry of Science and Innovation of Spain (SAF2005-02197 and SAF2008-02271) to DS and SAF2007-60287, Programa de Suport a Grups de Recerca from the Government of Catalonia (SGR2005-00628) and Ciberned (CB06/05/1104) to JXC. MVG is supported by ‘Departament d’Universitat, Recerca i Societat de la Informacio’ (DURSI) and the European Social Fund from the Government of Catalonia (DIUE).