The potassium channel Kv1.3 plays roles in immunity, neuronal development and sensory discrimination. Regulation of Kv1.3 by kinase signaling has been studied. In this context, EGF binds to specific receptors (EGFR) and triggers tyrosine kinase-dependent signaling, which down-regulates Kv1.3 currents. We show that Kv1.3 undergoes EGF-dependent endocytosis. This EGF-mediated mechanism is relevant because is involved in adult neural stem cell fate determination. We demonstrated that changes in Kv1.3 subcellular distribution upon EGFR activation were due to Kv1.3 clathrin-dependent endocytosis, which targets the Kv1.3 channels to the lysosomal degradative pathway. Interestingly, our results further revealed that relevant tyrosines and other interacting motifs, such as PDZ and SH3 domains, were not involved in the EGF-dependent Kv1.3 internalization. However, a new, and yet undescribed mechanism, of ERK1/2-mediated threonine phosphorylation is crucial for the EGF-mediated Kv1.3 endocytosis. Our results demonstrate that EGF triggers the down-regulation of Kv1.3 activity and its expression at the cell surface, which is important for the development and migration of adult neural progenitors.

The voltage-dependent potassium channel Kv1.3 plays essential physiological functions in the immune system. Kv1.3, regulating the membrane potential, facilitates downstream Ca2+ -dependent pathways and becomes concentrated in specific membrane microdomains that serve as signaling platforms. Increased and/or delocalized expression of the channel is observed at the onset of several autoimmune diseases. In this work, we show that adenosine (ADO), which is a potent endogenous modulator, stimulates PKC, thereby causing immunosuppression. PKC activation triggers down-regulation of Kv1.3 by inducing a clathrin-mediated endocytic event that targets the channel to lysosomal-degradative compartments. Therefore, the abundance of Kv1.3 at the cell surface decreases, which is clearly compatible with an effective anti-inflammatory response. This mechanism requires ubiquitination of Kv1.3, catalyzed by the E3 ubiquitin-ligase Nedd4-2. Postsynaptic density protein 95 (PSD-95), a member of the MAGUK family, recruits Kv1.3 into lipid-raft microdomains and protects the channel against ubiquitination and endocytosis. Therefore, the Kv1.3/PSD-95 association fine-tunes the anti-inflammatory response in leukocytes. Because Kv1.3 is a promising multi-therapeutic target against human pathologies, our results have physiological relevance. In addition, this work elucidates the ADO-dependent PKC-mediated molecular mechanism that triggers immunomodulation by targeting Kv1.3 in leukocytes.

In fluorescence spectroscopy and imaging experiments with the fluorophores laurdan and 1,6-diphenyl-1,3,5-hexatriene (DPH) we found that phenol derivatives modulate membrane properties. We propose that activation of TRPA1 by non-electrophilic compounds may arise from the induction of changes in membrane properties and suggest that chemosensation may result from primary mechanosensory mechanisms.

The voltage-dependent potassium channel Kv1.3 participates in the immune system physiology. Kv1.3 regulates the membrane potential triggering downstream Ca 2þ -dependent pathways. This channel concentrates in specific membrane microdomains serving as signaling platforms. Altered expression of Kv1.3 is observed at the onset of several autoimmune diseases. We show that adenosine (ADO), acting as a potent endogenous modulator, stimulates PKC, thereby causing immunosuppression. PKC activation down-regulates Kv1.3 triggering a clathrin-mediated internalization that targets the channel to lysosomes. Thus, the amount of Kv1.3 at the plasma membrane decreases, which is clearly compatible with an effective anti-inflammatory response. This mechanism involves ubiquitination of Kv1.3, catalyzed by the E3 ubiquitin-ligase Nedd4-2. Postsynaptic density protein 95 (PSD-95), member of the MAGUK family, situates Kv1.3 into lipid-raft microdomains impairing the ubiquitination and endocytosis of the channel. Therefore, the association of PSD-95 with Kv1.3 would modulate the anti-inflammatory response in leukocytes. This work elucidates the PKC-dependent molecular mechanisms that target Kv1.3 during immunomodulation in leukocytes.