Protein Kinase A Activation Promotes Plasma Membrane Insertion of DCC from an Intracellular Pool: A Novel Mechanism Regulating Commissural Axon Extension

Abstract

Protein kinase A (PKA) exerts a profound influence on axon extension during development and regeneration; however, the molecular mechanisms underlying these effects of PKA are not understood. Here, we show that DCC (deleted in colorectal cancer), a receptor for the axon guidance cue netrin-1, is distributed both at the plasma membrane and in a pre-existing intracellular vesicular pool in embryonic rat spinal commissural neurons. We hypothesized that the intracellular pool of DCC could be mobilized to the plasma membrane and enhance the response to netrin-1. Consistent with this, we show that application of netrin-1 causes a modest increase in cell surface DCC, without increasing the intracellular concentration of cAMP or activating PKA. Intriguingly, activation of PKA enhances the effect of netrin-1 on DCC mobilization and increases axon extension in response to netrin-1. PKA-dependent mobilization of DCC to the plasma membrane is selective, because the distributions of transient axonal glycoprotein-1, neural cell adhesion molecule, and trkB are not altered by PKA in these cells. Inhibiting adenylate cyclase, PKA, or exocytosis blocks DCC translocation on PKA activation. These findings indicate that netrin-1 increases the amount of cell surface DCC, that PKA potentiates the mobilization of DCC to the neuronal plasma membrane from an intracellular vesicular store, and that translocation of DCC to the cell surface increases axon outgrowth in response to netrin-1.

Opposing roles of netrin-1 and the dependence receptor DCC in cancer cell invasion, tumor growth and metastasis

“…Thus, we found that the PLC signaling cascade activated by the physiological agonist adenosine is the critical invasion effector acting through Rho/ROK-independent mechanisms and the Ca 2 þ -dependent myosin light chain kinase that is believed to promote the contractility of the actin cytoskeleton, as shown previously in our laboratory (Nguyen et al, 2005). Both adenosine and the A2B-R are linked to cAMP pathways and to the recruitment of DCC receptors from an intracellular pool to the cell surface, in a netrin-dependent manner (Bouchard et al, 2004). In contrast, we have shown that the proinvasive activity of netrin is determined by PKA, Rho/ROK and PI3K-dependent pathways via a PLC-independent mechanism.…”

Calcium signaling in axon guidance

“…Calcium signaling has an effect on the growth rate of growth cones, first characterized as a narrow extracellular calcium concentration range favorable for growth [43,[60][61][62]. Surprisingly, calcium transients [63,64] and calcium entry through mechanosensitive channels slow axon growth, whereas calcium release from ER stores enhances growth [41]. The relation between calcium transients and cAMP signaling may be different between the central domain of the growth cone and its filopodia [19].…”

14-3-3 Proteins Regulate Protein Kinase A Activity to Modulate Growth Cone Turning Responses

“…PKA can effect the insertion of guidance cue receptors into the plasma membrane (Bouchard et al, 2004). Both NGF and MAG can engage p75 NTR as part of their receptor complex in certain contexts raising the possibility that receptor dynamics may be altered by PKA activation at the site of ligand binding.…”

“…How the activation state of cytoskeletal regulatory proteins is spatially and temporally regulated is an important question which has only been partially elucidated. Local translation, insertion and removal of receptors at the plasma membrane as well as receptor silencing all affect growth cone responses (Stein and TessierLavigne, 2001;Bouchard et al, 2004;Kim et al, 2005;Lin and Holt, 2007). Further, regulating intracellular concentrations of cyclic nucleotides affects whether neuronal processes are attracted or repelled by guidance cues in vitro and in vivo Song et al, 1998;Song and Poo, 1999;Polleux et al, 2000;Shewan et al, 2002;Nishiyama et al, 2003).…”