bioRxiv : the preprint server for biology最新文献

{"title":"Replication fork remodeling proteins, Smc5/6 and Rtt107, promote palindrome-mediated genome instability.","authors":"Alex B Costa, Anissia Aid Saada, Thy Pham, Jiayi Fan, Rebecca Lever, Xiaolan Zhao, Kirill S Lobachev","doi":"10.64898/2026.04.29.721645","DOIUrl":"https://doi.org/10.64898/2026.04.29.721645","url":null,"abstract":"<p><p>Palindromic sequences are a potent source of genomic instability that can lead to cancer and hereditary diseases in humans. Molecular evidence shows that palindrome instability results from the formation of secondary structures, such as hairpins and cruciforms, which are cleaved by structure-specific nucleases. However, the mechanisms underlying cruciform formation and cleavage at palindromic sequences in eukaryotic cells remain incompletely understood. Here, we describe a pathway for secondary structure formation and chromosomal breakage at palindromes involving DNA helicase Mph1, Rad51 recombinase, Rad54 ATPase DNA strand remodeler, Rtt107 scaffold protein, and the multifunctional Smc5/6 complex. Deletion or mutation of any of these components results in a similar reduction in double-strand breaks at an Alu palindrome and a significant decrease in chromosomal rearrangements. We propose that Mph1, Rad51, and Rad54 work together at stalled replication forks to generate cruciform structures via fork remodeling, while Smc5/6 and Rtt107 mark the cruciforms, indicating an appropriate substrate for nuclease cleavage. As members of this pathway are conserved in humans, the uncovered mechanisms may underlie genomic instability at palindrome sites involved in disease etiology.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13142508/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147848780","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Olfactomedin4 marks luminal progenitor cells that give rise to secretory cell lineage in the mouse cervix.","authors":"ShanmugaPriyaa Madhukaran, Yevgenia Fomina, Govindkumar Balagannavar, Elizabeth Payne, Jason Wilson, Lei Wang, Gary Hon, Maria Florian Rodriguez, Mala Mahendroo","doi":"10.64898/2026.04.28.721179","DOIUrl":"https://doi.org/10.64898/2026.04.28.721179","url":null,"abstract":"<p><p>Development of the female reproductive tract in mice occurs in early postnatal life. The current model identifies Trp63 as the master regulator that initiates differentiation of simple columnar Keratin 8+ epithelium in the cervix and vagina into a stratified squamous epithelium. Thereafter Trp63+ basal progenitors maintain cervicovaginal epithelial cell homeostasis and in the adult serve as the progenitor for hormone-regulated shifts in stratified squamous and secretory luminal cells. This model differs from the human in which two progenitors, one columnar and the other basal gives rise to secretory cells in the endocervix and stratified squamous epithelia in the ectocervix and vagina respectively. In the current study, we identify a population of Krt8+, Tp63- epithelial cells that are retained in the cervicovaginal epithelium during the postnatal developmental period and into adulthood. Single cell datasets from the cervices of adult mice, identify Olfactomedin 4 (Olfm4), as a unique marker of the Krt8+Trp63- population. Adult lineage tracing and reassessment of gene markers during postnatal development support a revised model in which two progenitors are delineated in the mouse cervix and vagina by PND15. Olfactomedin 4+ progenitors give rise to specialized secretory goblet cells, while Trp63+ basal progenitors give rise to stratified squamous luminal cells in the cervix and vagina of nonpregnant and pregnant mice. Consistent with the expansion of goblet cells in pregnancy, the Olfm4+ progenitor is highly proliferative in early pregnancy and progesterone regulates increased goblet cell differentiation. These findings reveal a previously unrecognized species similarity between mice and humans in which goblet cell and squamous keratinized cell subtypes are derived from two progenitor populations respectively.</p><p><strong>Highlights: </strong>Two epithelial progenitors (Trp63 and Olfm4) populations are delineated in the cervix and vagina within the first two weeks of postnatal life.The Trp63+ progenitor gives rise to keratinized epithelial cells, whereas the Olfm4+ progenitor cells give rise to secretory goblet cells.lfm4 is not required for maintenance of the luminal progenitor or differentiation of goblet cells in the cervix and vagina during adulthood and pregnancy.In adults, progesterone promotes differentiation of Olfm4+ progenitors into goblet cells.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13142340/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147848786","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The Nuclear Pore Complex Facilitates Centriole-Nuclear Attachment in Spermatids.","authors":"Danielle B Buglak, Brian J Galletta, Nasser M Rusan","doi":"10.64898/2026.04.28.721503","DOIUrl":"https://doi.org/10.64898/2026.04.28.721503","url":null,"abstract":"<p><p>Proper connection between the sperm head and tail is critical for fertility and is mediated by the head-tail coupling apparatus (HTCA). Recent evidence suggests that the nuclear pore complex (NPC) may be important in male fertility, though a specific role at the HTCA has not been described. To investigate this, we performed a testis-specific RNAi screen targeting nucleoporins of the NPC. We identified Nup133 and Nup107 as regulators of HTCA development. We found that Nup133 and Nup107 were required to form the initial connection between the nucleus and centriole during HTCA establishment. We determined that failure to build the HTCA following Nup133 and Nup107 depletion was due to loss of nuclear envelope dynein/dynactin. Finally, we showed that loss of the NPC cytoplasmic filament component Nup358 results in the most severe centriole detachment phenotype, thus potentially functioning as the dynein anchor. Together, our data indicate that NPCs are critical regulators of early HTCA establishment and are required to recruit dynein to the nuclear envelope to bring the nucleus and centriole together during spermiogenesis.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13142455/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147848792","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Short- and Long-Term Effects of Social Isolation on Adult Murine Bone are Sex-Dependent.","authors":"William Aidan Martel, Stephen Bradley King, Eleanor Buchanan, Brooklynn M Merrill, Julia Patrizia Stohn, Daniel J Brooks, Deborah Barlow, Katherine J Motyl, Rebecca Mountain","doi":"10.64898/2026.04.28.721448","DOIUrl":"https://doi.org/10.64898/2026.04.28.721448","url":null,"abstract":"<p><p>Social isolation is a known modifiable risk factor for many chronic diseases including cardiovascular, metabolic, and brain disorders. Recent research has demonstrated that social isolation is similarly detrimental to skeletal health, but these effects may be sexually dimorphic. In rodents, isolation negatively affects bone in adult male mice, but not in females. However, these sex differences have not been systematically investigated, and it is unknown if they persist with long-term social isolation. The goal of our study was to investigate if isolation-induced bone loss may occur on different timescales between female and male mice, as well as investigate the potential roles of estrogen and testosterone. We examined bone changes in grouped (4 mice/cage) or isolated (1 mouse/cage) female and male 16-week-old C57BL/6J mice after 2, 4, or 8 weeks of treatment. We found that social isolation through single housing significantly reduced bone parameters across treatment lengths in male mice (20% reduction in Tb.BV/TV; 8% reduction in Ct.Th.) but not in females even with prolonged isolation. Isolation also decreased biomechanical properties in the femur of male but not female mice. While the females' overall bone phenotype was unaffected, isolated females did show an increase in bone turnover markers with as little as 2 weeks of isolation. Isolation also altered estrogen-related gene expression in male mice isolated for 4 or 8 weeks. Overall, our results demonstrate that short- and long-term social isolation has sexually dimorphic effects on murine bone. These findings have important clinical implications for individuals at risk for social isolation, as well as for pre-clinical rodent models utilizing single housing.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13142487/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147848832","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A unified photosensitizer platform for <i>in situ</i> DNA-, RNA-, and protein-directed proximity labeling.","authors":"Elijah B Biletch, Conor P Herlihy, Lidan Li, Mary Krebs, Conor J Kelly, Nicolas J Longhi, Olivia Weissenfels, Hanna G Goldberg, Kyle Brandt, Jonathan B Grimm, Luke D Lavis, Edward L Huttlin, Devin K Schweppe, Keriann M Backus, Brian J Beliveau","doi":"10.64898/2026.04.30.721698","DOIUrl":"10.64898/2026.04.30.721698","url":null,"abstract":"<p><p>Cells depend on the spatial organization of proteins, RNA, and DNA into discrete subcellular compartments. Previous methods have largely centered on measuring spatial organization based on only one of these biomolecular classes at a time. Here, we demonstrate that POCA photocatalytic proximity labeling can serve as a unified photosensitizer-based platform for profiling the proximal proteomes of protein, RNA, and DNA targets within a single experimental framework. We show that POCA can harness standard immunofluorescence or <i>in situ</i> hybridization workflows to specifically target organic fluorophore photosensitizers to intracellular targets for proximity labeling in fixed cells. POCA-targeted proximity labeling requires minimal cellular input and does not require genetic engineering. Additionally, POCA photosensitizers are selected to also be fluorescent, enabling direct confirmation of on-target localization by imaging prior to proteomic analysis. To demonstrate broad utility, we apply POCA across multiple molecular targets spanning protein, RNA, and genomic DNA, including components of the nuclear pore complex, nucleolus, nuclear speckles, telomeres, and pericentromeric heterochromatin. By anchoring proximity labeling to both a protein and an RNA within the same nuclear compartment, we resolve shared and distinct proximal proteomes from orthogonal molecular perspectives.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13142541/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147848689","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"DNA-induced conformational changes in SPRTN relieve its auto-inhibitory effect on protease activity.","authors":"Wei Song, Joseph A Newman, Yichen Zhao, Rod Chalk, Christina Redfield, Paul R Elliott, Kristijan Ramadan","doi":"10.64898/2026.04.29.721615","DOIUrl":"https://doi.org/10.64898/2026.04.29.721615","url":null,"abstract":"<p><p>The DNA-dependent metalloprotease SPRTN has emerged as a key enzyme in the proteolysis of DNA-protein crosslinks (DPCs), thereby protecting us against genome instability, accelerated ageing, and cancer. DNA and ubiquitin chains serve as the primary activator and catalyst of SPRTN proteolysis, respectively, but how they promote SPRTN activation and activity remains incompletely understood. To address this question, we developed a highly sensitive multi-turnover fluorescence resonance energy transfer (FRET) assay to monitor SPRTN proteolysis in real time. We found that the auto-cleaved N-terminal SPRTN fragment, comprising the metalloprotease domain (MPD), zinc-binding domain (ZBD), and basic region (BR), is highly stable, enzymatically active, and retains ubiquitin-dependent activation. Interestingly, the MPD alone exhibits basal intrinsic activity that is independent of both DNA activation and ubiquitin avidity effect. We show that ZBD and MPD together exert steric regulation: ZBD maintains MPD in an autoinhibited state, while MPD largely prevents ZBD from binding to DNA. BR, together with DNA, is essential to relieve ZBD-mediated inhibition of MPD. Using a site-trapping approach, we demonstrate that the ZBD-BR- DNA trinity induces an open conformation of the SPRTN N-terminus in cis, thereby releasing autoinhibition. MPD and BR together restrict the DNA-binding stoichiometry of ZBD, enabling SPRTN to function efficiently in proximity to DNA despite its low abundance in vivo. Collectively, our work overturns the long-standing dogma that SPRTN autocleavage inactivates the enzyme and reveals how DNA-induced conformational changes in SPRTN fine-tune its protease activity, providing a prerequisite for subsequent ubiquitin activation and rapid proteolysis of DPCs.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13142381/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147848172","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Microporous Immune-Isolating Capsule with Improved Diffusion for Restored Dynamic Bidirectional Hormone Signaling in a Murine Model of Premature Ovarian Insufficiency.","authors":"Delaney S Sinko, Margaret A Brunette, Despina Ifigenia Pavlidis, Monica A Rionda, Bipasha Ray, Michelle Tong, Suhani Thakur, Brendon Baker, Vasantha Padmanabhan, Ariella Shikanov","doi":"10.64898/2026.04.28.721364","DOIUrl":"https://doi.org/10.64898/2026.04.28.721364","url":null,"abstract":"<p><p>Pediatric cancer survivors treated with gonadotoxic chemotherapy or radiation face lifelong premature ovarian insufficiency (POI), leading to elevated risk of cardiovascular disease, osteoporosis, and metabolic dysfunction. Pharmacological hormone replacement therapy (HRT) cannot replicate the pulsatile, bidirectional signaling of the hypothalamic-pituitary-gonadal (HPG) axis, leaving a critical therapeutic gap. Immune-isolating hydrogel capsules offer a promising strategy for the implantation of donor ovarian tissue without immunosuppression yet they require optimization for human applications. Here, we engineer a microporous immune-isolating capsule by incorporating thermosensitive gelatin microgels as sacrificial porogens. Microfluidic fabrication yielded monodisperse microgels that dissolved at 37°C generating disconnected micropores within a non-degradable poly(ethylene glycol) (PEG) matrix. Critically, the diffusion of FSH-scale analogs (40 kDa) increased by almost two-fold through the microporous capsules relative to nanoporous controls, while antibody-scale molecules (150 kDa) were blocked, demonstrating size-discriminating permeability. In ovariectomized mice implanted with encapsulated ovarian xenografts for 20 weeks, microporous capsules restored dynamic HPG-axis signaling evidenced by elevated levels of estradiol and progesterone, FSH suppression, and fluctuating hormone levels that resembled physiological patterns. Microporosity also improved graft viability, increasing stromal cellularity and reducing follicular apoptosis. These findings support microporous immune-isolating capsules as a platform for physiologically authentic therapy for POI.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13142343/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147848647","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Deep mutational scan of the pore of the cold-sensing TRPM8 channel.","authors":"Averi Pike, Vincent Le, Ashlin Turner, Catalina Galvan, Ashley Vazhavilla, Richard Salinas, Matt Bramble, Dennis Wylie, Jessica Podnar, Ian Hoskins, Can Cenik, Andres Jara-Oseguera","doi":"10.64898/2026.04.28.721489","DOIUrl":"https://doi.org/10.64898/2026.04.28.721489","url":null,"abstract":"<p><p>Members of the Transient Receptor Potential (TRP) family of ion channels have a nearly ubiquitous role in human physiology, tuning cell signaling to remarkably diverse physical and chemical stimuli. Although there is extensive structural data on TRP channels, a systematic and unbiased interrogation of structure-function relations in these proteins is required to fully elucidate their mechanisms of function. By focusing on a critical pore region of the TRPM8 channel, which is the main detector of cold and cooling agents in sensory neurons, we show how deep mutational scanning can be used in combination with the available structural data to understand how TRP channels respond to stimuli. We define a novel mechanism whereby the extracellular pore loop, which has only been resolved in structures representing desensitized states of the channel, plays an essential role in the response of TRPM8 to menthol or cold by coordinating the movement of the S6 helices that line and gate the pore, and the ion-selectivity filter that binds permeant cations. Moreover, our screen reveals sequence determinants along the S6 helices that explain how their architecture sustains gating and, together, provide strong support for a structural mechanism of TRPM8 pore opening in response to menthol and cold.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13142389/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147848648","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Mitigating Family Effects in RNA Secondary-Structure Prediction with Latent-Space Continual Learning.","authors":"Wissal Mokkedem, Giulia Pedrielli, Teresa Wu","doi":"10.64898/2026.04.29.721709","DOIUrl":"https://doi.org/10.64898/2026.04.29.721709","url":null,"abstract":"<p><p>Accurate RNA secondary-structure prediction remains difficult despite decades of thermodynamics-based algorithms and the advent of deep-learning architectures (convolutional networks, Transformers, diffusion models). In fact, the datasets that pair RNA sequences with secondary-structure labels are often low-quality, noisy, and family-imbalanced, which limits out-of-distribution generalization and exacerbates catastrophic forgetting when new data regimes are introduced. We propose a continual-learning approach based on Lifelong Bayesian Optimization (LBO), RNAFoLBO, that treats each class of RNAs obtained from latent-space clustering as a sequential task and jointly orchestrates training and hyperparameter selection of heterogeneous models (UFold, RNA-FM, RNADiffFold), while preserving prior knowledge. Concretely, we apply LBO to 15 clusters obtained by clustering RNAStrAlign in the latent space of RNAGenesis, a model specialized in contextual representation learning and latent-space structuring, achieving a mean <i>F1</i> per cluster of 0.931 (with a range of 0.177). These results surpass the strongest one-shot baseline and mitigate forgetting without full retraining. The gains persist as additional clusters are introduced. Overall, RNAFoLBO delivers higher and more stable performance and practical scalability for integrating new RNA clusters or families, enabling more robust and transferable RNA secondary-structure prediction.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13142540/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147848688","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Pseudouridylation landscape across 42 <i>S. cerevisiae</i> cytosolic tRNA isoacceptors via Nanopore direct RNA sequencing.","authors":"Margaret L Barry, Robin L Abu-Shumays, Lauren E Barnes, Ethan A Shaw, Julia L Reinsch, Abigail L Vaaler, Zachary D Basham, Miten Jain, Kristin S Koutmou, David M Garcia","doi":"10.64898/2026.04.28.721490","DOIUrl":"https://doi.org/10.64898/2026.04.28.721490","url":null,"abstract":"<p><p>Pseudouridine is the most abundant RNA base modification due to its prevalence in tRNA and rRNA, where it serves as a key modulator of structure and function. Yet even in a widely used model organism, the budding yeast <i>Saccharomyces cerevisiae</i> , the positions of all pseudouridines in tRNA have not been completely annotated. Using Nanopore direct RNA sequencing (DRS), an established method for detecting RNA pseudouridylation positions, we sequenced cytosolic tRNA from eight pseudouridine synthase (PUS) knockout <i>S. cerevisiae</i> strains, including deletion strains of Pus1, Pus3, and Pus7. Analysis of these data verified thirty-four existing pseudouridine annotations and predicted eleven previously unannotated pseudouridine sites. Our analysis revealed DRS signal changes at several non-uridine sites with the loss of a PUS, including apparent changes in modification abundances at position 37 upon deletion of Pus3. LC-MS/MS and primer extension assays, however, indicated no change in the abundance of these modifications with the loss of Pus3. Our analysis underscores the need for caution in interpreting DRS-based signal changes, particularly in modification-dense regions. Combining existing modification annotations for the thirty-one isoacceptors in the Modomics database with our dataset that added annotations for the remaining eleven isoacceptors, we created a map of all detected pseudouridines, and the enzymes responsible for their catalysis, across the forty-two <i>S. cerevisiae</i> cytosolic tRNA isoacceptors.</p>","PeriodicalId":519960,"journal":{"name":"bioRxiv : the preprint server for biology","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2026-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13142501/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147848747","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}