Cortactin-binding protein 2 (CTTNBP2) interacts with cortactin to regulate cortactin mobility and control dendritic spine formation. CTTNBP2 has also been associated with autistic spectrum disorder. The regulation of dendritic spinogenesis could explain the association of CTTNBP2 with autism. Sequence comparison has indicated that CTTNBP2 N-terminal like protein (CTTNBP2NL) is a CTTNBP2 homolog. To confirm the specific effect of CTTNBP2 on dendritic spinogenesis, here we investigated whether CTTNBP2NL has a similar function to CTTNBP2. Although both CTTNBP2 and CTTNBP2NL interacted with cortactin, CTTNBP2NL was associated with stress fibers, whereas CTTNBP2 was distributed to the cortex and intracellular puncta. We also provide evidence that CTTNBP2, but not CTTNBP2NL, is predominantly expressed in the brain. CTTNBP2NL did not show any activity in the regulation of dendritic spinogenesis. In addition to spine morphology, CTTNBP2 was also found to regulate the synaptic distribution of striatin and zinedin (the regulatory B subunits of protein phosphatase 2A (PP2A)), which interact with CTTNBP2NL in HEK293 cells. The association between CTTNBP2 and striatin/zinedin suggests that CTTNBP2 targets the PP2A complex to dendritic spines. Thus, we propose that the interactions of CTTNBP2 and cortactin and the PP2A complex regulate spine morphologenesis and synaptic signaling.