Abstract Rab3 subfamily small G proteins (Rab3A, Rab3B, Rab3C, and Rab3D) control the regulated exocytosis in neuronal/secretory cells. Rab3B is also detected and upregulated in non-neuronal/non-secretory cells, whereas its function remains elusive. In the present study, we identified growth-arrest-specific gene 8 (Gas8), an evolutionally conserved microtubule-binding protein that is upregulated in growth-arrested NIH 3T3 cells and involved in the dynein motor regulation in flagellar/ciliary axoneme, as a novel Rab3B-binding protein using a yeast two-hybrid system. Rab3B as well as Gas8 was upregulated in growth-arrested NIH 3T3 cells and enriched in testis and lung with well-developed flagella/cilia. Gas8 was specifically interacted with the GTP-bound form of Rab3B and co-localized with Rab3B at the Golgi in NIH 3T3 cells. Furthermore, Rab3B was relocated upon expression of the Rab3B-binding domain of Gas8. These results suggest that Gas8 links Rab3B to microtubules in NIH 3T3 cells.