Protein methylation is one of the most important protein post-translational modifications. Protein arginine methyltransferase 5 (PRMT5) catalyzes the mono- and symmetric dimethylation of arginine residues of a wide range of proteins that play critical roles in various cellular processes. It is demonstrated that PRMT5 directly interacts with commonly misregulated or mutated proteins in cancer and PRMT5 plays an important role in cancer as a potential oncogene. RioK1 is required for cytoplasmic maturation of the small (40S) subunit of ribosome. Increased ribosome biogenesis is a critical requirement for all cancers and RIO kinases play an important role in efficient ribosome biogenesis. It is also reported that RioK1 plays distinct roles in cancer-supportive processes such as proliferation and migration. The PRMT5-MEP50 core complex binds to pICln or RioK1 in a mutually exclusive fashion to alter the specificity of substrate recognition and subsequent recruitment. RioK1 serves as adaptor protein of PRMT5 to recruit Nucleolin for methylation. Despite the fact that structural information for each component of the PRMT5-RioK1-Nucleolin complex is available, little is known about the molecular details of the interaction between the components. In this project, we report our attempt at the structural characterization of the PRMT5-RioK1-Nucleolin complex. Firstly, we investigated the interaction of PRMT5 and RioK1 homolog in Chaetomium thermophilum and solved the structure of ctPRTM5. CtPRMT5 adopts a tetrameric overall structure and an elongated α helix in the N-terminal region compared to human PRMT5. Secondly, we investigated the effect of RioK1 knockdown on ovarian cancer cells. Our results show that RioK1 function is positively related to the migration rate of ovarian cancer cells. Finally, we pulled out endogenous PRMT5 complexes from HEK293F cells. We report here that RNA integrity is required for the interaction between RioK1 and Nucleolin. We find evidence that indicates a direct interaction between human PRMT5 and the 80S ribosome particle. We also identified a novel assembly between human PRMT5 and the nucleosome.