Induction of calcium influx through TRPC5 channels by cross-linking of GM1 ganglioside associated with α5β1 integrin initiates neurite outgrowth

Gusheng Wu, Zi Hua Lu, Alexander Obukhov, Martha C. Nowycky, Robert W. Ledeen

Research output: Contribution to journalArticle

79 Citations (Scopus)

Abstract

Previous studies demonstrated that cross-linking of GM1 ganglioside with multivalent ligands, such as B subunit of cholera toxin (CtxB), induced Ca 2+ influx through an unidentified, voltage-independent channel in several cell types. Application of CtxB to undifferentiated NG108-15 cells resulted in outgrowth of axon-like neurites in a Ca2+ influx-dependent manner. In this study, we demonstrate that CtxB-induced Ca 2+ influx is mediated by TRPC5 channels, naturally expressed in these cells and primary neurons. Both Ca2+ influx and neurite induction were blocked by TRPC5 small interfering RNA (siRNA). Pretreatment of NG108-15 cells with neuraminidase increased cell-surface GM1 and greatly enhanced the signal. GM1 was not directly associated with TRPC5 but rather with α5β1 integrin, which opened the channel through a signaling sequence after cross-linking of the GM1/integrin complex. This cascade included autophosphorylation of focal adhesion kinase and subsequent activation of phospholipase Cγ (PLCγ) and phosphoinositide-3 kinase [PI(3)K]. Pharmacological blockers that inhibited tyrosine kinase, PLC, and PI(3)K suppressed both CtxB-induced Ca2+ influx and neurite outgrowth. These were also suppressed by SK&F96365, a nonspecific transient receptor potential channel blocker. Confocal immunocytochemistry revealed that GM1 cross-linking induced colocalization of GM1 with these signaling elements in sprouting regions of plasma membrane. In primary cerebellar granular neurons (CGNs), TRPC5 was detected at 2 d in vitro (2 DIV), a stage corresponding to CtxB-stimulated Ca2+ influx. Neurite outgrowth in CGNs, determined at 3 DIV, was accelerated by CtxB and suppressed by TRPC5 siRNA and the above blockers. The crucial role of GM1 was indicated with CGNs from ganglio-series null mice, in which growth of axons was significantly retarded.

Original languageEnglish
Pages (from-to)7447-7458
Number of pages12
JournalJournal of Neuroscience
Volume27
Issue number28
DOIs
StatePublished - Jul 11 2007

Fingerprint

G(M1) Ganglioside
Integrins
Calcium
Neurons
1-Phosphatidylinositol 4-Kinase
Type C Phospholipases
Neurites
Small Interfering RNA
Transient Receptor Potential Channels
Focal Adhesion Protein-Tyrosine Kinases
Cholera Toxin
Neuraminidase
Protein-Tyrosine Kinases
Axons
Immunohistochemistry
Cell Membrane
Neuronal Outgrowth
Pharmacology
Ligands
Growth

Keywords

  • Calcium signaling
  • Focal adhesion kinase
  • GM1 ganglioside
  • Integrin
  • Neuritogenesis
  • TRPC5 channel

ASJC Scopus subject areas

  • Neuroscience(all)

Cite this

Induction of calcium influx through TRPC5 channels by cross-linking of GM1 ganglioside associated with α5β1 integrin initiates neurite outgrowth. / Wu, Gusheng; Lu, Zi Hua; Obukhov, Alexander; Nowycky, Martha C.; Ledeen, Robert W.

In: Journal of Neuroscience, Vol. 27, No. 28, 11.07.2007, p. 7447-7458.

Research output: Contribution to journalArticle

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AU - Ledeen, Robert W.

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