Integrated role of U2AF2 and nuclear speckles in the regulation of splicing

• Applicant: Dr. Srimeenakshi Sankaranarayanan, Ph.D.
• Subject Area: General Genetics and Functional Genome Biology ; Bioinformatics and Theoretical Biology
• Term: since 2026
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 584853435

As soon as nascent RNA is transcribed, it undergoes extensive processing to generate functional messenger RNA (mRNA). Central to this maturation is splicing: by removing introns and joining exons, splicing determines mRNA fate and expands transcriptomic and proteomic diversity essential for cellular function. Aberrant splicing is tightly linked to disease and increasingly recognized as a molecular hallmark of cancer. Although splicing has been widely studied, how it is spatially coordinated within the crowded nuclear environment remains less explored. Membraneless organelles like nuclear speckles (NS) are now recognized as dynamic hubs for nuclear RNA processing. Once considered passive storage sites for splicing factors, NS have recently been found to be dynamic compartments that actively influence splicing. For example, proximity to NS correlates with improved splicing efficiency and gene expression. However, a key question remains: why are only some introns spliced in NS, and what determines their selective recruitment? To address this, I focus on U2AF2, a critical factor involved in 3´splice site recognition and early spliceosome assembly. U2AF2 localizes to NS, likely in a splicing-dependent manner. My preliminary data, based on time-course transient transcriptome sequencing (TT-seq) following rapid U2AF2 depletion, show that acute loss of U2AF2 delays splicing of specific introns. Notably, a preliminary analysis of published APEX-seq data revealed that NS-associated introns are preferentially dependent on U2AF2, suggesting that U2AF2 is required for efficient NS-mediated splicing and revealing a previously unrecognized functional link between U2AF2 and NS-mediated splicing. This proposal aims to dissect the role of U2AF2 and NS in nuclear splicing regulation through two objectives: 1. Identifying the role of U2AF2 in regulating splicing dynamics by investigating the influence of U2AF2 on splicing kinetics and classifying introns based on their dependency on U2AF2. 2. Characterizing the integrated roles of U2AF2 and NS and identifying defining features of NS-associated introns, thereby revealing molecular determinants that govern selective recruitment of introns to NS, and the contribution of U2AF2 in NS-mediated splicing. By integrating time-resolved TT-seq with publicly available NS transcriptome datasets such as APEX-seq, ARTR-seq, my work will (i) identify the dynamic role of U2AF2 during early stages of splice site definition, (ii) explain how specific introns are selectively directed to NS, (iii) uncover the spatial role of U2AF2 in NS-mediated splicing. Aligned with the goals of the Walter Benjamin Programme, this project marks a decisive step in my transition from experimental fungal RNA biologist to independent researcher in computational RNA biology, empowering me to investigate key mechanisms of nuclear RNA processing.

• DFG Programme: Position