Activation of RhoA and SAPK/JNK signalling pathways by the RhoA-specific exchange factor mNET1.

We have characterized the DH domain protein mNET1, a Rho-family guanine nucleotide exchange factor (GEF). N-terminal truncation of mNET1 generates an activated transforming form of the protein, mNET1DeltaN, which acts as a GEF for RhoA but not Cdc42 or Rac1. In NIH 3T3 cells, activated mNET1 induces formation of actin stress fibres and potentiates activity of the transcription factor serum response factor. Inhibitor studies show that these processes are dependent on RhoA and independent of Cdc42 or Rac1. In contrast to the GTPase-deficient RhoA.V14 mutant, however, expression of activated mNET1 also activates the SAPK/JNK pathway. This requires mNET1 GEF activity, since it is blocked by point mutations in the mNET1 DH domain and its C-terminal pleckstrin homology (PH) domain, and by the dominant-interfering RhoA mutant RhoA.N19. Although mNET1DeltaN-induced SAPK/JNK activation requires a C3 transferase-sensitive GTPase, it occurs independently of the generation of titratable GTP-bound RhoA. Thus, mNET1 can activate signalling pathways in addition to those directly controlled by activated RhoA.