Deciphering the role of the putative RNA-Binding Protein TFIP11 in regulating tissue-specific alternative splicing

RNA-binding proteins (RBPs) comprise a large class of proteins that regulate the metabolism of RNA transcripts throughout their life cycle. Many regulatory RBPs function in a cell-, tissue-, or condition-specific manner and are capable of regulating variety of molecular processes such as alternative splicing of messenger RNAs that is critical for appropriate protein expression in the corresponding tissues. The proper activity of RBPs is essential for human physiology, and their dysregulation has been implicated in diseases including neurodegeneration, autoimmune disorders, and cancer. RNA-binding domains (RBDs) in RBPs are the functional units responsible for RNA binding. Multiple RBDs are often present in a single RBP and these modular domains can coordinate and enhance binding to RNA in a sequence and/or structure-specific manner. Intrinsically disordered regions (IDRs) often behave as linkers connecting RBDs within a single RBP but these regions also have the ability to mediate interactions with RNA directly. Tuftelin-Interacting Protein 11 (TFIP11) is a spliceosomal protein that harbors two putative RBDs: an N-terminal G-patch domain and a C-terminal double-stranded RNA-binding domain (dsRBD), both flanked by IDRs. While we have previously shown that TFIP11 plays critical roles in spliceosome assembly and activation, the RNA-binding functionality of its individual domains and their roles in splicing regulation remained uncharacterized. In this study, we employed a multidisciplinary strategy combining in cellulo, in vitro, in silico, and in vivo approaches to investigate the RNA-binding properties of TFIP11’s RBDs and IDRs, and to define their functional relevance in alternative splicing regulation. Our findings provide novel mechanistic insights into TFIP11’s role as a splicing regulator and support its classification as a previously unrecognized RNA-binding protein with broad implications for RNA biology and disease.