Biology subjects

Di Stefano, B.

Publications and source records attributed to Di Stefano, B..

4 recordsLinked to original sources

bioRxiv2026

Co-option of ILF2/3 in primates restrains Alu hyper-editing to enable cell fate transitions

Understanding human specific mechanisms of cell fate control is essential for advancing developmental biology and regenerative medicine. Here, we identify the ILF2/3 complex as a critical regulator of primate cell fate transitions. Using genetically and epigenetically engineered gastruloids and adult stem cells, we show that ILF2/3 is required for gastrulation in primates but not mice, and for differentiation of adult progenitor cells. Mechanistically, ILF2/3 directly binds Alu elements in chromatin-associated RNAs and shields them from ADAR1-mediated adenosine-to-inosine (A-to-I) editing. Acute ILF2/3 degradation increases A-to-I editing at Alu elements, but not murine retrotransposons, leading to aberrant splicing and nonsense-mediated decay of transcripts encoding key chromatin regulators in primate cells. This in turn destabilizes the epigenetic landscape and blocks lineage commitment across all three germ layers. Re-expression of correctly spliced chromatin regulators rescues differentiation defects in ILF2/3-deficient cells, functionally linking Alu editing control to chromatin regulation and cell fate. These findings define an evolutionary mechanism that restrains retrotransposon-associated RNA editing to preserve proteome integrity and enable primate-specific developmental programs.

genetics↗

bioRxiv2025

Selective RNA sequestration in biomolecular condensates directs cell fate transitions

Recent studies have emphasized the significance of biomolecular condensates in modulating gene expression through RNA processing and translational control. However, the functional roles of RNA condensates in cell fate specification remains poorly understood. Here, we profiled the coding and non-coding transcriptome within intact biomolecular condensates, specifically P-bodies, in diverse developmental contexts, spanning multiple vertebrate species. Our analyses revealed the conserved, cell type-specific sequestration of untranslated RNAs encoding key cell fate regulators. Notably, P-body contents did not directly reflect active gene expression profiles for a given cell type, but rather were enriched for translationally repressed transcripts characteristic of the preceding developmental stage. Mechanistically, microRNAs (miRNAs) direct the selective sequestration of RNAs into P-bodies in a context-dependent manner, and perturbing AGO2 or alternative polyadenylation profoundly reshapes P-body RNA content. Building on these mechanistic insights, we demonstrate that modulating P-body assembly or miRNA activity dramatically enhances both activation of a totipotency transcriptional program in naive pluripotent stem cells as well as the programming of primed human embryonic cells towards the germ cell lineage. Collectively, our findings establish a direct link between biomolecular condensates and cell fate decisions across vertebrate species and provide a novel framework for harnessing condensate biology to expand clinically relevant cell populations.

cell biology↗

bioRxiv2020

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

In contrast to the extensively studied rewiring of epigenetic and transcriptional programs required for cell reprogramming, the dynamics of post-transcriptional changes and their associated regulatory mechanisms remain poorly understood. Here we have studied the dynamics of alternative splicing (AS) changes occurring during efficient reprogramming of mouse B cells into induced pluripotent stem (iPS) cells. These changes, generally uncoupled from transcriptional regulation, significantly overlapped with splicing programs reported during reprogramming of mouse embryonic fibroblasts (MEFs). Correlation between gene expression of potential regulators and specific clusters of AS changes enabled the identification and subsequent validation of CPSF3 and hnRNP UL1 as facilitators, and TIA1 as repressor of MEFs reprogramming. These RNA-binding proteins control partially overlapping programs of splicing regulation affecting genes involved in developmental and morphogenetic processes. Our results reveal common programs of splicing regulation during reprogramming of different cell types and identify three novel regulators of this process.

molecular biology↗

bioRxiv2018

Single cell expression analysis uncouples transdifferentiation and reprogramming

Many somatic cell types are plastic, having the capacity to convert into other specialized cells (transdifferentiation)(1) or into induced pluripotent stem cells (iPSCs, reprogramming)(2) in response to transcription factor over-expression. To explore what makes a cell plastic and whether these different cell conversion processes are coupled, we exposed bone marrow derived pre-B cells to two different transcription factor overexpression protocols that efficiently convert them either into macrophages or iPSCs and monitored the two processes over time using single cell gene expression analysis. We found that even in these highly efficient cell fate conversion systems, cells differ in both their speed and path of transdifferentiation and reprogramming. This heterogeneity originatesin two starting pre-B cell subpopulations,large pre-BII and the small pre-BII cells they normally differentiate into. The large cells transdifferentiate slowly but exhibit a high efficiency of iPSC reprogramming. In contrast, the small cells transdifferentiate rapidly but are highly resistant to reprogramming. Moreover, the large B cells induce a stronger transient granulocyte/macrophage progenitor (GMP)-like state, while the small B cells undergo a more direct conversion to the macrophage fate. The large cells are cycling and exhibit high Myc activity whereas the small cells are Myc low and mostly quiescent. The observed heterogeneity of the two cell conversion processes can therefore be traced to two closely related cell types in the starting population that exhibit different types of plasticity. These data show that a somatic cells propensity for either transdifferentiation and reprogramming can be uncoupled.\n\nOne sentence summarySingle cell transcriptomics of cell conversions

developmental biology↗