Biology subjects

Alepuz, P.

Publications and source records attributed to Alepuz, P..

7 recordsLinked to original sources

bioRxiv2026

Efficient translation of chaperones by eIF5A safeguards proteostasis under heat stress

The increase in global temperature poses a major threat to cells and organisms because it disrupts cellular proteostasis. We discovered that the evolutionarily conserved and essential translation elongation factor eIF5A is necessary for a rapid response to heat-stress in yeast cells. Lack of eIF5A results in ribosome stalling on heat-stress-induced mRNAs, reducing the synthesis of chaperones, primarily impacting those that undergo robust synthesis via high codon optimality, such as Hsp70 Ssa1 and Hsp12 chaperones. This new eIF5A role in translation appears to be direct, as eIF5A binds preferentially to induced mRNAs with high codon optimality and its polysome association increases under heat-shock. Low levels of Hsp70 in eIF5A mutants delay Hsf1 feedback inhibition during stress adaptation and cause the accumulation of protein aggregates, resulting in low cellular viability to heat-stress. Overall, our results show that eIF5A is crucial for heat-stress resistance by facilitating rapid and efficient translation of high codon-optimal chaperones. Therefore, eIF5A emerges as a key regulator of cellular proteostasis, providing a novel perspective on its significant involvement in neurodegenerative diseases and aging.

molecular biology↗

bioRxiv2026

eIF5A is an indispensable protein for eukaryotic cells

eIF5A is an evolutionarily conserved protein found in all eukaryotes, and in Archaea and bacteria. It functions as a translation factor promoting ribosomal elongation, and it can also bind to genes in the nucleus and to mRNA. eIF5A is encoded by two paralogous genes in most eukaryotes, with one copy (TIF51A in yeast and EIF5A1 in humans) highly expressed and essential, and the other (TIF51B in yeast and EIF5A2 in humans) repressed in most cells and conditions. eIF5A is linked to viral infection, aging, diabetes and neurodevelopmental disorders. Whilst derepression of the duplicated silent gene EIF5A2 is associated with several cancers, promoting metastasis. To expand our knowledge of eIF5As function, we searched for suppressors of yeast temperature-sensitive mutants of TIF51A, which cannot grow at restrictive temperature. All suppressors contained mutations in the transcriptional repressors Rox1 and Mot3, resulting in the upregulation of the paralogous gene TIF51B. Next, we searched for suppressors of TIF51A temperature-sensitive mutants in yeast lacking the TIF51B gene. The frequency of suppression was 20-times lower and all suppressors were revertants or contained intragenic mutations in the Tif51A protein that conferred stability. Our results suggest that the duplicated eIF5A gene serves as a non-conventional backup system that rescues mutations in the first copy, but acting at the population level. Furthermore, our results expand our understanding of the repression mechanisms that keep the second eIF5A gene silent. Lastly, our results demonstrate that eIF5A is an indispensable protein in eukaryotic cells, whose function cannot be substituted by mutations in other proteins or pathways. Article summaryeIF5A is an evolutionary conserved protein encoded by two paralogous genes in eukaryotes: one highly expressed and essential, and the other silent. eIF5A promotes translation elongation, and its deficiency is linked to diabetes, aging and neurodevelopmental disorders; while derepression of the silent gene promotes metastatic cancers. We isolated suppressors of conditional mutants of the first eIF5A gene in yeast cells. Suppressors either up-regulated the second eIF5A gene or reverted first copy mutations, restoring eIF5A protein levels and survival. Our work deepens our understanding of the eIF5A paralogue gene repression, and demonstrates that eIF5A is an indispensable protein for eukaryotic cells.

genetics↗

bioRxiv2025

eIF5A coordinates the transcription and translation of its target genes

Maintaining balanced cellular protein levels requires precise control of gene expression and effective coordination between the various stages of the process, from transcription to translation. In recent years, several components of the translation apparatus have been found in the nuclei of various eukaryotes, where they regulate transcription, mRNA processing or export, thereby integrating different stages of gene expression. eIF5A is an essential and evolutionarily conserved translation elongation factor that is involved in viral infection and in the development of diseases such as cancer and neurodevelopmental disorders. eIF5A promotes translation elongation by binding to ribosomes that stall at codons encoding problematic amino acids for peptide bond formation, such as consecutive prolines, also known as polyproline motifs. Although eIF5A shuttles between the nucleus and cytoplasm, its specific nuclear roles remain poorly defined. Here, we demonstrate that nuclear yeast eIF5A binds to chromatin and represses gene transcription by preventing the binding of RNA polymerase II. Importantly, chromatin binding and transcriptional repression by eIF5A have a higher impact on genes encoding its own translational targets. The presence of polyproline motifs in genes imposes both translation and transcriptional control by eIF5A. Furthermore, eIF5As active engagement in cytoplasmic translation is necessary for its role in repressing transcription. Our results suggest that eIF5A coordinates gene expression by promoting the cytoplasmic translation of specific genes while repressing their transcription in the nucleus, thus ensuring efficient final protein synthesis. Significance StatementOur study provides genome-wide and gene-specific evidence supporting the role of the translation elongation factor eIF5A in transcription. eIF5A is essential in eukaryotes, facilitating the translation of mRNAs encoding stretches of problematic amino acids, such as consecutive prolines. Through its role in the synthesis of specific proteins, eIF5A has been linked to development and different diseases, including cancer and diabetes. We have now discovered that eIF5A also controls the transcription of its translation target genes and this effect is driven by the presence of eIF5A-dependent motifs at their sequences. In the nucleus, eIF5A binds to specific genes and attenuates the binding of RNA polymerase II. By negatively regulating transcription and positively regulating translation, eIF5A coordinates gene expression, fine-tuning protein levels.

molecular biology↗

bioRxiv2023

eIF5A controls mitoprotein import by relieving ribosome stalling at the TIM50 translocase mRNA

The efficient import of nuclear-encoded proteins into mitochondria is crucial for proper mitochondrial function. The conserved translation factor eIF5A is primarily known as an elongation factor which binds ribosomes to alleviate ribosome stalling at sequences encoding polyprolines or combinations of proline with glycine and charged amino acids. eIF5A is known to impact the mitochondrial function across a variety of species although the precise molecular mechanism underlying this impact remains unclear. We found that depletion of eIF5A in yeast drives reduced translation and levels of TCA cycle and oxidative phosphorylation proteins. We further found that loss of eIF5A leads to the accumulation of mitoprotein precursors in the cytosol as well as to the induction of a mitochondrial import stress response. Here we identify an essential polyproline-containing protein as a direct eIF5A target for translation: the mitochondrial inner membrane protein Tim50, which is the receptor sub-unit of the TIM23 translocase complex. We show how eIF5A directly controls mitochondrial protein import through the alleviation of ribosome stalling along TIM50 mRNA at the mitochondrial surface. Removal of the polyprolines from Tim50 rescues the mitochondrial import stress response, as well as the translation of oxidative phosphorylation reporter genes in an eIF5A loss of function. Overall, our findings elucidate how eIF5A impacts the mitochondrial function by reducing ribosome stalling and facilitating protein translation, thereby positively impacting the mitochondrial import process.

cell biology↗

bioRxiv2021

Hypusinated eIF5A is required for the translation of collagen

The evolutionary conserved elongation factor eIF5A is required for the translation of mRNAs that encode protein sequences with consecutive prolines or combined with glycine and charged amino acids. Mammalian collagens are enriched in putative eIF5A-dependent Pro-Gly-containing tripeptides. Here, we show that eIF5A is needed for heterologous expression of collagen in yeast, and using a dual luciferase reporter system we confirmed that eIF5A depletion interrupts translation at Pro-Gly-collagenic motifs. Using mouse fibroblasts, we showed that depletion of active eIF5A reduced collagen 1 (Col1a1) content, which became concentrated around the nuclei, in contrast to a stronger and all over the cell collagen signal in untreated cells. Active eIF5A-depleted mouse fibroblast showed upregulation of endoplasmic reticulum (ER) stress markers, suggesting retention of partially synthesized Col1a1 in the ER. A dramatically lower level of Col11 protein was also observed in functional eIF5A-depleted human hepatic stellate cells treated with the profibrotic cytokine TGF-{beta}1. Our results show that collagen expression requires eIF5A and imply its potential as a target for regulating collagen production in fibrotic diseases.

molecular biology↗

bioRxiv2020

Xrn1 influences RNA pol II-dependent transcription elongation rates across the yeast genome and this control is particularly relevant for late elongation of regulatory genes

mRNA homeostasis is favored by crosstalk between transcription and degradation machineries. Both the Ccr4-Not and the Xrn1-decaysome complexes have been described to influence transcription. While Ccr4-Not has been shown to directly stimulate transcription elongation, the information available on how Xrn1 influences transcription is scarce and contradictory. In this study we have addressed this issue by mapping RNA polymerase II (RNA pol II) at high resolution, using CRAC and BioGRO-seq techniques in Saccharomyces cerevisiae. We found significant effects of Xrn1 perturbation on RNA pol II profiles across the genome. RNA pol II profiles at 5 exhibited significant alterations that were compatible with decreased elongation rates in the absence of Xrn1. Nucleosome mapping detected altered chromatin configuration in the gene bodies. We also detected accumulation of RNA pol II shortly upstream of polyadenylation sites by CRAC, although not by BioGRO-seq, suggesting higher frequency of backtracking before pre-mRNA cleavage. This phenomenon was particularly linked to genes with poorly positioned nucleosomes at this position. Accumulation of RNA pol II at 3 was also detected in other mRNA decay mutants. According to these and other pieces of evidence, Xrn1 seems to influence transcription elongation at least in two ways: by directly favoring elongation rates and by a more general mechanism that connects mRNA decay to late elongation.

genomics↗

bioRxiv2018

The 5’ untranslated region of the EFG1 transcript promotes its translation to regulate hyphal morphogenesis in Candida albicans

Extensive 5 untranslated regions (UTR) are a hallmark of transcripts determining hyphal morphogenesis in Candida albicans. The major transcripts of the EFG1 gene, which are responsible for cellular morphogenesis and metabolism, contain a 5 UTR of up to 1170 nt. Deletion analyses of the 5 UTR revealed a 218 nt sequence that is required for production of the Efg1 protein and its functions in filamentation, without lowering the level and integrity of the EFG1 transcript. Polysomal analyses revealed that the 218 nt 5 UTR sequence is required for efficient translation of the Efg1 protein. Replacement of the EFG1 ORF by the heterologous reporter gene CaCBGluc confirmed the positive regulatory importance of the identified 5 UTR sequence. In contrast to other reported transcripts containing extensive 5 UTR sequences, these results indicate the positive translational function of the 5 UTR sequence in the EFG1 transcript, which is observed in context of the native EFG1 promoter. The results suggest that the 5 UTR recruits regulatory factors, possibly during emergence of the native transcript, which aid in translation of the EFG1 transcript.\n\nIMPORTANCEMany of the virulence traits that make Candida albicans an important human fungal pathogen are regulated on a transcriptional level. Here we report an important regulatory contribution of translation, which is exerted by the extensive 5 untranslated regulatory sequence (5 UTR) of the transcript for the protein Efg1, which determines growth, metabolism and filamentation in the fungus. Presence of the 5 UTR is required for efficient translation of Efg1, to promote filamentation. Because transcripts for many relevant regulators contain extensive 5 UTR sequences, it appears that virulence of C. albicans depends on the combination of transcriptional and translation regulatory mechanisms.

microbiology↗