Dynamics of alternative splicing during somatic cell reprogramming reveals functions for RNA-binding proteins CPSF3, hnRNP UL1, and TIA1

Abstract

Background Somatic cell reprogramming is the process that allows differentiated cells to revert to a pluripotent state. In contrast to the extensively studied rewiring of epigenetic and transcriptional programs required for reprogramming, the dynamics of post-transcriptional changes and their associated regulatory mechanisms remain poorly understood. Here we study the dynamics of alternative splicing changes occurring during efficient reprogramming of mouse B cells into induced pluripotent stem (iPS) cells and compare them to those occurring during reprogramming of mouse embryonic fibroblasts. Results We observe a significant overlap between alternative splicing changes detected in the two reprogramming systems, which are generally uncoupled from changes in transcriptional levels. Correlation between gene expression of potential regulators and specific clusters of alternative splicing changes enables the identification and subsequent validation of CPSF3 and hnRNP UL1 as facilitators, and TIA1 as repressor of mouse embryonic fibroblasts reprogramming. We further find that these RNA-binding proteins control partially overlapping programs of splicing regulation, involving genes relevant for developmental and morphogenetic processes. Conclusions Our results reveal common programs of splicing regulation during reprogramming of different cell types and identify three novel regulators of this process and their targets.

Deficiency of T-Cell Intracellular Antigen 1 in Murine Embryonic Fibroblasts Is Associated with Changes in Mitochondrial Morphology and Respiration

“…TIA1 regulates the decay and translation of mRNAs encoding a diverse class of proteins, including inducible proinflammatory cytokines, constitutive survival factors, and angiogenic growth-associated proteins [ 12 , 13 , 14 , 15 , 16 , 17 ]. There are no RNA binding maps of TIA1 in MEFs, but previous findings and the present study (by using large-scale binding and functional maps of human RBPs in human cells) suggest that TIA1 could regulate mRNA stability and RNA decay through varying regulatory mechanisms that likely involve cell-type-specific co-factors [ 18 , 19 , 20 , 21 , 22 ]. Thus, TIA1 could have a functionally versatile role, acting as a dual agent pro-growth or pro-senescence factor during embryonic development, cellular homeostasis and stress, or tumorigenesis in a cellular context-dependent manner [ 2 , 24 , 25 , 26 , 27 , 28 ].…”

“…Changes in the expression and/or subcellular localization of TIA1 have been associated with important pathophysiological consequences in human biology and disease, including embryogenesis [ 14 , 21 , 22 ], inflammation [ 14 , 23 , 24 ], tumorigenesis [ 25 , 26 , 27 ], neuronal homeostasis [ 28 , 29 ], tauopathies [ 30 ], myopathies [ 31 , 32 , 33 , 34 , 35 ], cell stress [ 10 , 11 , 12 ] and viral infections [ 2 , 36 ]. The participation of TIA1 in these complex programs points to its direct involvement in the regulation of myriad cellular pathways among other apoptosis [ 1 , 25 ], autophagy/mitophagy [ 21 , 26 , 37 ], immune system [ 13 , 24 , 38 ], membrane dynamics [ 39 ], axonal regeneration [ 40 ], activity and localization of cellular translational machinery [ 17 , 41 ], cell cycle [ 2 , 19 , 24 , 29 ], proteostasis [ 42 , 43 ], dynamics of stress granules during environmental challenges (oxidative, heat, osmotic, etc.)…”

The Multifunctional Faces of T-Cell Intracellular Antigen 1 in Health and Disease

“…One of the nuclear functions of TIA1 is to regulate the alternative splicing (and also constitutive splicing) of some pre-mRNAs to favor the inclusion [19,[33][34][35] or selective exclusion [36] of exons (Figures 5 and 6). For splicing to occur, the spliceosome must first be assembled, which begins with the recognition of the 5 intronic splicing site (5 ss) by the nuclear ribonucleoprotein U1 (snRNP U1).…”

“…It is in this context that TIA1 exerts its function. To promote exon inclusion, TIA1 binds to uridine-rich RNA sequences located near both constitutive and alternative 5 ss with weakly conserved consensus sequences for U1 snRNA hybridization and U1 snRNP anchoring and subsequently binds to the U1-C protein of the U1 snRNP [13,19,[33][34][35][36][37][38][39]. In this process, RRM2 binds to the uridine-rich regions near the 5 ss of the intron, favored by RRM3, and the Q/N-rich domain binds to the N-terminal region of the U1-C subunit of the U1 snRNP by an RRM1-favored process [38,39].…”

“…In this process, RRM2 binds to the uridine-rich regions near the 5 ss of the intron, favored by RRM3, and the Q/N-rich domain binds to the N-terminal region of the U1-C subunit of the U1 snRNP by an RRM1-favored process [38,39]. Its role as a splicing repressor is linked to competition for uridine-rich sequences within the polypyrimidine-rich regions located at positions near the 3 ss of introns to interfere with auxiliary-factor (perhaps U2AF65) binding by preventing U2 snRNP recruitment [35,36].…”