Control of Growth Cone Motility and Morphology by LIM Kinase and Slingshot via Phosphorylation and Dephosphorylation of Cofilin

Abstract

Growth cone motility and morphology are based on actin-filament dynamics. Cofilin plays an essential role for the rapid turnover of actin filaments by severing and depolymerizing them. The activity of cofilin is repressed by phosphorylation at Ser3 by LIM kinase (LIMK, in which LIM is an acronym of the three gene products Lin-11, Isl-1, and Mec-3) and is reactivated by dephosphorylation by phosphatases, termed Slingshot (SSH). We investigated the roles of cofilin, LIMK, and SSH in the growth cone motility and morphology and neurite extension by expressing fluorescence protein-labeled cofilin, LIMK1, SSH1, or their mutants in chick dorsal root ganglion (DRG) neurons and then monitoring live images of growth cones by time-lapse video fluorescence microscopy. The expression of LIMK1 remarkably repressed growth cone motility and neurite extension, whereas the expression of SSH1 or a nonphosphorylatable S3A mutant of cofilin enhanced these events. The fan-like shape of growth cones was disorganized by the expression of any of these proteins. The repressive effects on growth cone behavior by LIMK1 expression were significantly rescued by the coexpression of S3A-cofilin or SSH1. These findings suggest that LIMK1 and SSH1 play critical roles in controlling growth cone motility and morphology and neurite extension by regulating the activity of cofilin and may be involved in signaling pathways that regulate stimulus-induced growth cone guidance. Using various mutants of cofilin, we also obtained evidence that the actin-filament-severing activity of cofilin is critical for growth cone motility and neurite extension.

Phosphorylation of Actin-Depolymerizing Factor/Cofilin by LIM-Kinase Mediates Amyloid -Induced Degeneration: A Potential Mechanism of Neuronal Dystrophy in Alzheimer's Disease

“…Because ADF/cofilin is the only known substrate for LIMK1 in nerve cells (Bamburg and Wiggan, 2002;Meyer and Feldman, 2002), its inactivation by LIMK1, might result in actin filament accumulation. This interpretation is consistent with previously reported effects of LIMK1 (Endo et al, 2003;Rosso et al, 2004). Importantly, the nontoxic soluble-monomeric form of A␤1-40 did not induce dystrophy (Busciglio et al, 1992;Heredia et al, 2004), nor did it promote actin filament accumulation or increase the P-cofilin level, indicating that the conversion of A␤ to the fibrillar form is required to activate LIMK1 and to promote actin remodeling and dystrophy.…”

“…LIM kinase 1 (LIMK1) phosphorylates and thereby inhibits ADF/cofilin (Arber et al, 1998). LIMK1 is widely expressed in tissues, including the nervous system (Proschel et al, 1995;Foletta et al, 2004), and its activity modulates neuritogenesis and synaptic plasticity (Meng et al, 2002;Endo et al, 2003;Rosso et al, 2004). LIMK1 is a key downstream effector of the Rho family of small GTPases and is activated by phosphorylation on Thr508 by Rho kinase ROCK, or by Rac-Cdc42 p21-activated kinase (PAK) (Edwards et al, 1999).…”

The Interaction of mPar3 with the Ubiquitin Ligase Smurf2 Is Required for the Establishment of Neuronal Polarity

“…mPar3 controls tight junction assembly by regulating cofilin through the inhibition of LIM kinase 2 (26). The regulation of cofilin may have a similar function in neurons to control actin dynamics and neurite extension (31). A misregulation of LIM kinase 2 and cofilin could also contribute to the reduction of neurite extension after suppression of mPar3.…”

A proposed mechanism for cell polarization with no external cues

“…The inactivating phosphorylation is reversed by broadly acting phosphatases (PP1 and PP2B) [Meberg et al, 1998;Ambach et al, 2000] or is reversed presumably more specifically by slingshot. Slingshot, with at least 6 mammalian isoforms [Ohta et al, 2003], is a phosphatase first discovered in Drosophila [Niwa et al, 2002;Endo et al, 2003]. The localization of this "phosphocycling" during migration and possible higher order regulation by the protein 14-3-3 [Gohla and Bokoch 2002;Birkenfeld et al, 2002] are under investigation.…”

“…Such alkalization would be expected through ADF/cofilin proteins to create new fast-growing filament ends and initially a larger monomer actin pool for directed lamellipodial expansion by enhanced addition at the fast-growing ends (barbed ends). Barbed-end creation through cofilin severing is essential for initiation of epithelial cell migration with guidance cues [Chan et al, 2000;Zebda et al, 2000], and regulation of cofilin is essential for motility of neuronal growth cones [Endo et al, 2003].…”