Induction of filopodia-like protrusions by transmembrane agrin: Role of agrin glycosaminoglycan chains and Rho-family GTPases

Abstract

Filopodia sense the extracellular environment and direct movement in many cell types, including neurons. Recent reports suggest that the transmembrane form of the widely- expressed proteoglycan agrin (TM-agrin) regulates formation and stability of neuronal filopodia. In order to elucidate the mechanism by which TM-agrin regulates filopodia, we investigated the role of agrin’s glycosaminoglycan (GAG) chains in the induction of filopodia formation by TM-agrin over-expression in hippocampal neurons, and in the induction of filopodia-like processes in COS7 cells. Deletion of the GAG chains of TM-agrin sharply reduced formation of filopodia-like branched retraction fibers (BRFs) in COS7 cells, with deletion of the heparan sulfate GAG chains being most effective, and eliminated filopodia induction in hippocampal neurons. GAG chain deletion also reduced the activation of Cdc42 and Rac1 resulting from TM-agrin over-expression. Moreover, dominant negative Cdc42 and Rac1 inhibited BRF formation. Lastly, over-expression of TM-agrin increased the adhesiveness of COS7 cells and this increase was reduced by deletion of the GAG chains. Our results suggest that TM-agrin regulates actin-based protrusions in large part through interaction of its GAG chains with extracellular or transmembrane proteins, leading to the activation of Cdc42 and Rac1.

CHCHD2 Is Coamplified with EGFR in NSCLC and Regulates Mitochondrial Function and Cell Migration

“…For example, Agrin protein expression increased 2-fold upon CHCHD2 knockdown and this effect was reversed by CHCHD2 rescue. Overexpression of Agrin increased substrate adhesion in COS7 cells (47). We therefore speculate that elevated Agrin may have stimulated cell adhesion, and consequently hindered cell migration, in response to CHCHD2 knockdown.…”

mDia1 and WAVE2 Proteins Interact Directly with IRSp53 in Filopodia and Are Involved in Filopodium Formation

“…This set of criteria was used to distinguish them from neurites, which typically span the length of at least one cell body in N1E115 cells (greater than 15 m in length), as well as retraction fibers, which we have observed by live imaging to often be tapered (wider at the base than at the distal end closer to the tip) and branched, and sometimes appear in clusters or with several of them emerging from the same angle with respect to the cell perimeter (aligned parallel to one another). Fascin and myosin X were not used as markers for identifying filopodia as they have been shown to be present in retraction fibers (34). Only peripheral filopodia were scored, as the dorsal protrusions observed in both fixed and live N1E115 cells overexpressing activated Rif (9) were not seen in the same type of cells overexpressing IRSp53.…”

Emerging roles for MAP kinases in agrin signaling

“…Antibody-induced clustering or overexpression of transmembrane agrin in neuronal and non-neuronal cells induces filopodia. [40][41][42][43] This activity depends on domains in the N-terminal third of the agrin protein, 43,44 not overlapping with the C-terminal domains responsible for AChR clustering. ERK1/2 is likely to mediate filopodia induction by transmembrane agrin as PD98059, a MEK1/2 inhibitor, 45 blocked this activity.…”