INHIBITION OF LYSOPHOSPHATIDATE-INDUCED AND THROMBIN-INDUCED NEURITE RETRACTION AND NEURONAL CELL ROUNDING BY ADP-RIBOSYLATION OF THE SMALLGTP-BINDING PROTEIN-RHO

Addition of the bioactive phospholipid lysophosphatidic acid (LPA) or a thrombin receptor-activating peptide (TRP) to serum-starved N1E-115 or NG108-15 neuronal cells causes rapid growth cone collapse, neurite retraction, and transient rounding of the cell body. These shape chang es appear to be driven by receptor-mediated contraction of the cortica l actomyosin system independent of classic second messengers. Treatmen t of the cells with Clostridium botulinum C3 exoenzyme, which ADP-ribo sylates and thereby inactivates the Rho small GTP-binding protein, inh ibits LPA- and TRP-induced force generation and subsequent shape chang es. C3 also inhibits LPA-induced neurite retraction in PC12 cells. Bio chemical analysis reveals that the ADP-ribosylated substrate is RhoA. Prolonged C3 treatment of cells maintained in 10% serum induces the ph enotype of serum-starved cells, with initial cell flattening being fol lowed by neurite outgrowth; such C3-differentiated cells fail to retra ct their neurites in response to agonists. We conclude that RhoA is es sential for receptor-mediated force generation and ensuing neurite ret raction in N1E115 and PC12 cells, and that inactivation of RhoA by ADP -ribosylation abolishes actomyosin contractility and promotes neurite outgrowth.