Biology subjects

Palancade, B.

Publications and source records attributed to Palancade, B..

6 recordsLinked to original sources

bioRxiv2026

DNA:RNA hybrid mutational landscapes reveal a common route to geneticinstability in evolving genomes

Pervasive DNA:RNA hybrids are recognized as pathological sources of genome instability, yet how their genotoxicity shapes mutational landscapes and genome evolution remains poorly understood. Here, we define the mutational footprint of hybrids by integrating genome-wide mapping, long-term mutation accumulation experiments, and analyses of genetic diversity across thousands of natural yeast genomes. We uncover signatures for embedded ribonucleotides and genic R-loops, together with a composite mutational pattern shared across natural populations and experimental evolution. Using reporters designed to dissect the mechanisms underlying genetic alterations, we find that they arise primarily from targeting by MutL{gamma}-dependent mismatch repair followed by Rad52-mediated recombination. Although hybrids form dynamically across the genome, their genotoxicity is strongly skewed toward repetitive regions susceptible to these error-prone pathways, including microsatellites and retroelements. Our findings reveal a common mechanism of hybrid-associated mutagenesis that can shape genetic variation in evolving genomes.

molecular biology↗

bioRxiv2026

Shining a light on the dark Nt-acetylome by integrating omics data

N-terminal acetylation, catalysed by N-terminal acetyltransferases (Nats), is one of the most prevalent protein modifications and is implicated in human diseases. Yet, despite extensive COFRADIC-based proteomics, only [~]5-10% of the proteome has been interrogated, leaving the majority of the Nt-acetylome unexplored. Here, we combined all major COFRADIC datasets with sel-TRAP, a high-sensitivity, orthogonal approach for profiling co-translational Nat targets via selective ribosome purification. This integrated analysis refined the substrate specificities of NatA, NatB, and NatC/E/F and provided the most comprehensive view to date of the canonical human and yeast Nt-acetylomes. Importantly, we also uncovered hundreds of cryptic Nat substrates arising from alternative translation initiation, with unexpected Nt-proteoforms constituting a previously underappreciated source of Nat targets. Collectively, our results revealed the complex landscape of a "dark" Nt-acetylome, the characterization of which, including its functional roles in regulating protein function and in disease, remains a major challenge for future research.

genomics↗

bioRxiv2024

Different alterations in mRNP composition trigger an increase in harmful R-loops

Understanding how the assembly of nascent mRNA into a ribonucleoprotein (mRNP) influences R-loop homeostasis is crucial for gaining insight into the cellular mechanisms that prevent genome instability. Here, we identify three RNA-binding proteins, Rie1, Rim4 and She2, whose stoichiometry is important to limit R-loop accumulation and, thus, to prevent DNA damage. Interestingly, Rim4 and She2 are overrepresented in mRNPs assembled in the absence of THO. In addition, we found that an excess of Dis3 impairs exosome function, promoting R-loops, particularly from non-coding RNAs, which cause genomic instability. Our results indicate that mRNP assembly is a precise process that, when perturbed by changes in the availability of different RBPs or RNAs, causes R-loop-mediated DNA damage in the cell. These results may help to understand the mechanism that promotes cancer, as several RBPs are overexpressed in different types of tumors. Author summaryR-loops are three-stranded structures formed by a DNA-RNA hybrid and the displaced ssDNA. The accumulation of R-loops damages the genome, highlighting the need for their tight regulation. One of the mechanisms that prevents R-loop formation is the co-transcriptional assembly of the nascent RNA into a messenger ribonucleoprotein (mRNP). Our study reveals that mRNP biogenesis is highly sensitive to perturbations. Not only the absence of mRNP components, but also the overexpression of specific RNA-binding proteins (RBPs) or the persistence of unstable transcripts, significantly stresses mRNP assembly, resulting in R-loop accumulation and consequent DNA damage. R-loops are involved in a number of neurodegenerative diseases and cancer, highlighting the importance of understanding the cellular processes that mitigate their accumulation.

genetics↗

bioRxiv2022

Functional mapping of N-terminal residues in the yeast proteome uncovers novel determinants for mitochondrial protein import.

N-terminal ends of polypeptides are critical for the selective co-translational recruitment of N-terminal modification enzymes. However, it is unknown whether specific N-terminal signatures differentially regulate protein fate according to their cellular functions. In this work, we developed an in-silico approach to detect functional preferences in cellular N-terminomes, and identified in S. cerevisiae more than 200 Gene Ontology terms with specific N-terminal signatures. In particular, we discovered that Mitochondrial Targeting Sequences (MTS) show a strong and specific over-representation at position 2 of hydrophobic residues known to define potential substrates of the N-terminal acetyltransferase NatC. We validated mitochondrial precursors as co-translational targets of NatC by selective purification of translating ribosomes, and found that their N-terminal signature is conserved in Saccharomycotina yeasts. Finally, systematic mutagenesis of the position 2 in a prototypal yeast mitochondrial protein confirmed its critical role in mitochondrial protein import. Our work highlights the hydrophobicity of MTS N-terminal residues and their modification by NatC as critical features for the definition of the mitochondrial proteome, providing a molecular explanation for mitochondrial defects observed in yeast or human NatC-depleted cells. Functional mapping of N-terminal residues thus has the potential to support the discovery of novel mechanisms of protein regulation or targeting.

genomics↗

bioRxiv2021

A nuclear pore sub-complex restricts the propagation of Ty retrotransposons by limiting their transcription

Beyond their canonical function in nucleocytoplasmic exchanges, nuclear pore complexes (NPCs) regulate the expression of protein-coding genes. Here, we have implemented transcriptomic and molecular methods to specifically address the impact of the NPC on retroelements, which are present in multiple copies in genomes. We report a novel function for the Nup84 complex, a core NPC building block, in specifically restricting the transcription of LTR-retrotransposons in yeast. Nup84 complex-dependent repression impacts both Copia and Gypsy Ty LTR-retrotransposons, all over the S. cerevisiae genome. Mechanistically, the Nup84 complex restricts the transcription of Ty1, the most active yeast retrotransposon, through the tethering of the SUMO-deconjugating enzyme Ulp1 to NPCs. Strikingly, the modest accumulation of Ty1 RNAs caused by Nup84 complex loss-of-function is sufficient to trigger an important increase of Ty1 cDNA levels, resulting in massive Ty1 retrotransposition. Altogether, our studies expand our understanding of the complex interactions between retrotransposons and the NPC, and highlight the importance for the cells to keep retrotransposon under tight transcriptional control. AUTHOR SUMMARYRetroelements, which replicate by reverse transcription of their RNA into a cDNA that is integrated into the host genetic material, play an important role in the plasticity of eukaryotic genomes. The study of yeast retrotransposons has led to the identification of host factors that limit retroelement mobility, including components of the nuclear pore complex (NPC), most of them still awaiting mechanistic characterization. Here, we investigated the contribution of the Nup84 complex, a core NPC scaffold, to retrotransposon biology in budding yeast. Our findings uncover that the Nup84 complex restricts the transcription of phylogenetically-distinct Ty retroelements. By focusing on Ty1 retrotransposons, we provide evidence that repression by the Nup84 complex depends on the maintenance at the NPC of the SUMO-protease Ulp1, an essential enzyme of the SUMO pathway with multiple targets in the transcription machinery. We finally show that this transcriptional control is critical for genome dynamics, since a small increase in the accumulation of Ty1 RNAs leads to massive retrotransposition. Our data reveal that although relatively abundant, Ty transcripts are limiting for retrotransposition, underscoring the importance of a tight control of their expression. They also characterize a new non-canonical function of NPCs, confirming their connection with genome expression and stability.

molecular biology↗

bioRxiv2018

The mRNA export adaptor Yra1 contributes to DNA double-strand break repair through its C-box domain

Yra1 is an mRNA export adaptor involved in mRNA biogenesis and export in S. cerevisiae. Yra1 overexpression was recently shown to promote accumulation of DNA:RNA hybrids favoring DNA double strand breaks (DSB), cell senescence and telomere shortening, via an unknown mechanism. Yra1 was also identified at an HO-induced DSB and Yra1 depletion causes defects in DSB repair. Previous work from our laboratory showed that Yra1 ubiquitination by Tom1 is important for mRNA export. Interestingly, we found that Yra1 is also ubiquitinated by the SUMO-targeted ubiquitin ligases Slx5-Slx8 implicated in the interaction of irreparable DSB with nuclear pores. Here we show that Yra1 binds an HO-induced irreparable DSB. Importantly, a Yra1 mutant lacking the evolutionarily conserved C-box is not recruited to an HO-induced irreparable DSB and becomes lethal under DSB induction in a HO-cut reparable system. Together, the data provide evidence that Yra1 plays a crucial role in DSB repair via homologous recombination. Unexpectedly, while the Yra1 C-box is essential, Yra1 sumoylation and/or ubiquitination are dispensable in this process.

molecular biology↗