The RNA-binding protein TIA-1 is a novel mammalian splicing regulator acting through intron sequences adjacent to a 5 ' splice site

Splicing of the K-SAM alternative exon of the fibroblast growth factor rece ptor 2 gene is heavily dependent on the U-rich sequence IAS1 lying immediat ely downstream from its 5' splice site. We show that IAS1 can activate the use of several heterologous 5' splice sites in vitro. Addition of the RNA-b inding protein TIA-1 to splicing extracts preferentially enhances the use o f 5' splice sites linked to IAS1. TIA-1 can provoke a switch to use of such sites on pre-mRNAs with competing 5' splice sites, only one of which is ad jacent to IAS1. Using a combination of UV cross-linking and specific immuno precipitation steps, we show that TIA-1 binds to IAS1 in cell extracts. Thi s binding is stronger if IAS1 is adjacent to a 5' splice site and is U1 snR NP dependent. Overexpression of TIA-I in cultured cells activates K-SAM exo n splicing in an IAS1-dependent manner. If IAS1 is replaced with a bacterio phage MS2 operator, splicing of the K-SAM exon can no longer be activated b y TIA-1. Splicing can, however, be activated by a TIA-1-MS2 coat protein fu sion, provided that the operator is close to the 5' splice site. Our result s identify TIA-I as a novel splicing regulator, which acts by binding to in tron sequences immediately downstream from a 5' splice site in a U1 snRNP-d ependent fashion. TIA-1 is distantly related to the yeast U1 snRNP protein Nam8p, and the functional similarities between the two proteins are discuss ed.