Binding of a SART3 tumor-rejection antigen to a pre-mRNA splicing factor RNPS1: a possible regulation of splicing by a complex formation

We recently reported the identification of a human SART3 gene that encodes a tumor-rejection antigen recognized by cytotoxic T lymphocytes (CTLs). The squamous-cell carcinoma antigen recognized by T cells-3 (SART3) is an RNA- binding protein expressed in the nucleus of the majority of proliferating c ells, including normal cells and malignant cells, but not in normal tissues except for the testes and fetal liver. To determine its biologic function, we employed a 2-hybrid screening in yeast for proteins interacting with SA RT3, and this method yielded a pre-mRNA splicing factor (RNA-binding protei n prevalent during the S phase or RNA-binding protein with a serine-rich do main [RNPS1]) that activated both constitutive and alternative splicing of pre-mRNA in vitro. Interaction of SART3 with RNPS1 through the physical ass ociation of N-terminal domains of RNPS1 was confirmed by both in vitro pull -down assay and immunoprecipitation assay. Cotransfection of the 2 genes ch anged the distribution pattern of SART3 from diffuse nucleoplasmic spreadin g to nuclear speckled regions in which the RNPS1 was colocalized, suggestin g a complex formation of the 2 proteins. In cooperation with RNPS1, SART3 s timulated the proximal alternative 3' splicing of a calcitonin-dihydrofolat e reductase chimeric minigene pre-mRNA. These results suggest that SART3 is involved in the regulation of mRNA splicing probably via its complex forma tion with RNPS1. (C) 2001 Wiley-Liss, Inc.