Scaffolding proteins play a critical role in conferring specificity and fidelity to signaling pathways. The JNK-interacting leucine zipper protein (JLP) has been identified as a scaffolding protein involved in linking components of the JNK signaling module. Galpha(12) and Galpha(13), the alpha-subunits of heterotrimeric G proteins G12 and G13, respectively, stimulate the JNK module in diverse cell types. Here, we report that Galpha(13) physically interacts with JLP, and this interaction enhances Galpha(13)-mediated JNK activation. We also demonstrate endogenous interaction between JLP and Galpha(13) in MCF-7 cells. JLP interaction is specific to the G12 family of alpha-subunits via its C-terminal domain (termed GID-JLP), spanning amino acids 1165-1307, and this interaction is more pronounced with the mutationally or functionally activated form of Galpha(13) compared to that of wild-type Galpha(13). The presence of a ternary complex consisting of Galpha(13), JLP, and JNK suggests a role for JLP in tethering Galpha(13) to the signaling components involved in JNK activation. Coexpression of GID-JLP disrupts ternary complex formation in addition to attenuating Galpha(13)-stimulated JNK activity. These findings identify JLP as a novel scaffolding protein in the Galpha(13)-mediated JNK signaling pathway.