C53/LZAP (also named as Cdk5rap3) is a putative tumor suppressor that plays important roles in multiple cell signaling pathways, including DNA damage response and NF-kappaB signaling. Yet how its function is regulated remains largely unclear. Here we report the isolation and characterization of two novel C53/LZAP-interacting proteins, RCAD (Regulator of C53/LZAP and DDRGK1) and DDRGK1 (DDRGK domain-containing protein 1). Our co-immunoprecipitation assays confirmed their interactions, while gel filtration assay indicated that C53/LZAP and RCAD may form a large protein complex. Intriguingly, we found that RCAD knockdown led to dramatic reduction of C53/LZAP and DDRGK1 proteins. We also found that C53/LZAP and DDRGK1 became more susceptible to the proteasome-mediated degradation in RCAD knockdown cells, whereas their ubiquitination was significantly attenuated by RCAD overexpression. In addition, we found that RCAD, like C53/LZAP, also plays an important role in regulation of NF-kappaB signaling and cell invasion. Taken together, our findings strongly suggest that RCAD is a novel regulator of C53/LZAP tumor suppressor and NF-kappaB signaling.