Proteins of the SURF6 family are evolutionary conservative house-keeping nucleolar proteins, but the functional role of human SURF6 and its protein partners are still unknown. In this study, we use the affinity adsorption (GST pull-down) assay in order to answer these questions. The recombinant human SURF6 and the recombinant conservative C-terminal domain of the mouse Surf6 (Surf6-dom), which has 85% homology with the human SURF6 domain, were fused with glutathione-S-transferase (GST) and used as acceptors. We demonstrated that GST-SURF6 interacts with several RNA processing factors (B23/nucleophosmin, nucleolin, and EBP2), and also with the cofactor of RNA-polymerase I, the UBF protein. The same protein partners were bound to GST-Surf6-dom. These data are the first experimental evidences in favor of the participation of human SURF6 in ribosomal biogenesis, including transcription of rDNA and processing of rRNA. In addition, the set of the protein partners of GST-Surf6-dom in the HeLa cells pointed to a possible interaction of human SURF6 with the nucleolar and nuclear proteins of other functional groups, i.e.; to its multifunctionality. © 2014 Pleiades Publishing, Ltd.