Suppression of Radixin and Moesin Alters Growth Cone Morphology, Motility, and Process Formation In Primary Cultured Neurons

Abstract

In this study we have examined the cellular functions of ERM proteins in developing neurons. The results obtained indicate that there is a high degree of spatial and temporal correlation between the expression and subcellular localization of radixin and moesin with the morphological development of neuritic growth cones. More importantly, we show that double suppression of radixin and moesin, but not of ezrin–radixin or ezrin–moesin, results in reduction of growth cone size, disappearance of radial striations, retraction of the growth cone lamellipodial veil, and disorganization of actin filaments that invade the central region of growth cones where they colocalize with microtubules. Neuritic tips from radixin–moesin suppressed neurons displayed high filopodial protrusive activity; however, its rate of advance is 8–10 times slower than the one of growth cones from control neurons. Radixin–moesin suppressed neurons have short neurites and failed to develop an axon-like neurite, a phenomenon that appears to be directly linked with the alterations in growth cone structure and motility. Taken collectively, our data suggest that by regulating key aspects of growth cone development and maintenance, radixin and moesin modulate neurite formation and the development of neuronal polarity.

Radixin Knockdown by RNA Interference Suppresses Human Glioblastoma Cell Growth in Vitro and in Vivo

“…An apoptotic property of these cells was observed both in vitro and in tumor tissues. Our results are consistent with previous finding in hippocampal pyramidal cells (Paglini et al, 1998). The observed effects of radixin silencing on the GBM cell transformation indicate a role for radixin in regulating tumor metastasis and progression.…”

Evidence for the Role of MAP1B in Axon Formation

“…Interestingly, immunoelectron microscopy has revealed that microtubules penetrating into the growth cone peripheral domain contain MAP1B and interact with actin filament bundles (Bush et al, 1996;Gordon-Weeks and Fischer, 2000). Therefore, by favoring the existence of dynamic microtubules within growth cones, MAP1B may contribute to the promotion of lamellipodial spreading, an event required for neuronal polarization Dotti, 1997, 1999;Paglini et al, 1998b). In favor of this idea, we observed a significant reduction in the size of the lamellipodial veil and a decrease in the number of actin ribs in axonal growth cones of MAP1B-deficient neurons.…”

Evidence for the Involvement of Tiam1 in Axon Formation

“…C ells were fixed before or after detergent extraction under microtubule-stabilizing conditions and processed for immunofluorescence as described previously (Paglini et al, 1998a). For some experiments a mild extraction protocol that preserves cytoskeletalmembrane interactions was also used (Nakata and Hirokawa, 1987;Brandt et al, 1995;Paglini et al, 1998b).…”

“…In cultured hippocampal pyramidal neurons, axon formation is preceded by the appearance in one of the multiple neurites of a large and highly dynamic growth cone containing a very labile actin network Dotti, 1997, 1999;Paglini et al, 1998a). Thus, the regulation of actin organization and activity within selected growth cones appears to be one of the major factors underlying the establishment of neuronal polarity.…”

“…To test whether the localization of Tiam1 in neurites and growth cones involves an interaction with components of the cytoskeleton, neurons were extracted with Triton X-100 (0.2%) before fixation under microtubule stabilizing conditions (Paglini et al, 1998a). This procedure, which selectively removes cytosolic proteins while preserving microtubules, did not significantly alter the distribution of Tiam1 when compared with that observed in cells fixed before detergent extraction.…”