GSK3β phosphorylation modulates CLASP–microtubule association and lamella microtubule attachment

Abstract

Polarity of the microtubule (MT) cytoskeleton is essential for many cell functions. Cytoplasmic linker–associated proteins (CLASPs) are MT-associated proteins thought to organize intracellular MTs and display a unique spatiotemporal regulation. In migrating epithelial cells, CLASPs track MT plus ends in the cell body but bind along MTs in the lamella. In this study, we demonstrate that glycogen synthase kinase 3β (GSK3β) directly phosphorylates CLASPs at multiple sites in the domain required for MT plus end tracking. Although complete phosphorylation disrupts both plus end tracking and association along lamella MTs, we show that partial phosphorylation of the identified GSK3β motifs determines whether CLASPs track plus ends or associate along MTs. In addition, we find that expression of constitutively active GSK3β destabilizes lamella MTs by disrupting lateral MT interactions with the cell cortex. GSK3β-induced lamella MT destabilization was partially rescued by expression of CLASP2 with mutated phosphorylation sites. This indicates that CLASP-mediated stabilization of peripheral MTs, which likely occurs in the vicinity of focal adhesions, may be regulated by local GSK3β inactivation.

Plus-end tracking proteins, CLASPs, and a viral Akt mimic regulate herpesvirus-induced stable microtubule formation and virus spread

“…Depletion of CRMP2 had no effect on MT stabilization or virus spread, demonstrating specificity in the GSK3β-regulated +TIPs that contribute to the biology of infection. CLASPs mediate MT stabilization in response to GSK3β inactivation and MT orientation toward the leading edge of motile cells and neuronal growth cones (29,(41)(42)(43). Similar to HSV-1-infected cells, depletion of CLASPs results in MT looping (43) and reduced levels of Ac-MTs (29,41,43,44).…”

Collapsin Response Mediator Proteins (CRMPs) Are a New Class of Microtubule-associated Protein (MAP) That Selectively Interacts with Assembled Microtubules via a Taxol-sensitive Binding Interaction

“…To date, these include MAP1b, Tau, ACF7, and CLASP2 (49,50,52,53). It has been suggested that the polarized migration of cells in general requires local GSK3␤ inhibition at the leading edge (52,54,55).…”

Identification of a Role for CLASP2 in Insulin Action

“…This finding does not imply that all phosphorylation of CLASP2 is controlled by the MAPK pathway, raising the possibility that CLASP2 may undergo PI3K-controlled phosphorylation in response to insulin as well. In support of this hypothesis, it has been shown that GSK3␤, a kinase known to be deactivated by insulin-stimulated Akt (25,26), phosphorylates CLASP2, leading to disruption of the association of CLASP2 with MTs (13). In this case, insulin would suppress GSK3␤ phosphorylation sites within CLASP2.…”

“…Current indirect evidence supports a role for CLASP2 in insulin action. Glycogen synthase kinase 3␤ (GSK3␤), a kinase known to be deactivated by insulin-stimulated Akt, phosphorylates CLASP2, leading to disruption of the association of CLASP2 with MTs (13,14). CLASP2 associates with LL5␤, a pleckstrin homology (PH) domain-containing protein that interacts with PI3K-generated PIP 3 (9,15).…”