Microtubule and Cell Contact Dependency of ER-bound PTP1B Localization in Growth Cones

Abstract

PTP1B is an ER-bound protein tyrosine phosphatase implied in the regulation of cell adhesion. Here we investigated mechanisms involved in the positioning and dynamics of PTP1B in axonal growth cones and evaluated the role of this enzyme in axons. In growth cones, PTP1B consistently localizes in the central domain, and occasionally at the peripheral region and filopodia. Live imaging of GFP-PTP1B reveals dynamic excursions of fingerlike processes within the peripheral region and filopodia. PTP1B and GFP-PTP1B colocalize with ER markers and coalign with microtubules at the peripheral region and redistribute to the base of the growth cone after treatment with nocodazole, a condition that is reversible. Growth cone contact with cellular targets is accompanied by invasion of PTP1B and stable microtubules in the peripheral region aligned with the contact axis. Functional impairment of PTP1B causes retardation of axon elongation, as well as reduction of growth cone filopodia lifetime and Src activity. Our results highlight the role of microtubules and cell contacts in the positioning of ER-bound PTP1B to the peripheral region of growth cones, which may be required for the positive role of PTP1B in axon elongation, filopodia stabilization, and Src activity. INTRODUCTIONProper discrimination and response to extracellular signals by axonal growth cones are crucial for the wiring of the nervous system. The sensing activity is performed by receptors in the plasma membrane of growth cones, from which cell adhesion receptors represent a major category. In part, information relied by these receptors regulates the behavior of the cytoskeleton and other macromolecular complexes (Suter and Forscher, 2000;Huber et al., 2003). Growth cone responses vary, but depend on the dynamic properties of microfilaments and microtubules. Microtubules explore dynamically the periphery of growth cones and occasionally penetrate into filopodia (Gordon-Weeks, 1991;Tanaka and Kirschner, 1991;Schaefer et al., 2002;Brown and Bridgman, 2003;Schober et al., 2007). It was early proposed that a subset of exploratory microtubules become selectively stabilized in response to extracellular cues (Kirschner and Mitchison, 1986;Mitchison and Kirschner, 1988). Indeed, imaging of Ti1 pioneer neurons in live grasshopper embryonic limb buds shows the selective invasion of microtubules toward growth cone branches that contact guidepost cells (Sabry et al., 1991). Furthermore, studies in bag cell neurons of Aplysia show that microtubules quickly invade regions of contact between growth cone filopodia and neighboring cells or beads coated with ApCAM (Lin and Forscher, 1993;Suter et al., 2004). This process would facilitate the translocation of vesicular and other membranous compartments to contact sites (Waterman-Storer et al., 2000;Ligon and Holzbaur, 2007; Shaw et al., 2007;Spiliotis et al., 2008). Some degree of microtubule dynamics, however, seems to be required for events induced by target interactions of growth cones, including morphological changes, the assem...

Protein tyrosine phosphatase PTP1B in cell adhesion and migration

“…Consistently, axons also showed reduced net elongation rate and increased pause times compared with axons of control neurons. 3 Collectively, these results strongly suggest that PTP1B promotes cell migration by mechanisms that stabilize lamellar protrusions.…”

“…35,36 In neurons, microtubule-dependent localization of ER-bound PTP1B at the peripheral region of axonal growth cones is critical for Src activation and filopodia stability. 3 In migrating fibroblasts, the ER network extends toward the leading edge, positioning GFP-PTP1B over new adhesions assembled at the advancing lamella ( Fig. 1A and B).…”

“…PTP1B was shown to promote cell adhesion and motility in fibroblasts, 17,18 neurons, 3,19 platelets, 4 and several tumor cell lines. 12,[20][21][22] An inhibitory role has been described in fibroblasts, 23,24 primary aortic smooth muscle cells, 25 ovarian cancer cells, 26 and in glioblastoma multiforme tumor cell invasion in mice.…”

“…27 Remarkably, the positive role of PTP1B in cell motility was frequently associated with the stimulation of integrin-dependent signaling. 3,4,12,17,18,22 In contrast, the negative effect of PTP1B on cell motility was related to antagonizing signaling from growth factor receptor tyrosine kinases. [24][25][26][27] In a recent quantitative study, we analyzed the intrinsic motility and migration parameters of PTP1B-null cells (KO cells) in an isotropic 2-D fibronectin substratum.…”

“…Extension of the ER toward the cell periphery allows PTP1B to dephosphorylate substrates located at the plasma membrane, for example Src, EphA3, and proteins associated to cadherin and integrin complexes. [2][3][4][5][6][7][8] Alternatively, PTP1B can dephosphorylate the cytosolic domains of endocytosed receptors, which are brought into contact with the ER, as, for example, activated platelet-derived growth factor and epidermal growth factor receptors. 9 Through the proline-rich motif, PTP1B has also been shown to interact directly with components of focal adhesions, such as the SH3 domain of adaptor protein p130Cas.…”