Biology subjects

Cakir, U.

Publications and source records attributed to Cakir, U..

9 recordsLinked to original sources

bioRxiv2026

A single-nucleus regulatory atlas links transposable elements to adult hippocampal neurogenesis

Adult hippocampal neurogenesis can be enhanced or even triggered by extrinsic stimuli, but the lineage-specific regulatory changes that accompany a stimulated neurogenic response remain undefined. We used recombinant human erythropoietin (rhEPO), a defined influencing factor that enriches newly formed pyramidal neurons by ~2 fold in this paradigm, to profile ~35,000 nuclei by single-nucleus ATAC sequencing (snATAC-seq) from rhEPO- and placebo-treated adult mouse hippocampi and to integrate them with matched single-nucleus RNA sequencing (snRNA-seq) of ~120,000 nuclei. Thirty-five accessibility-defined clusters resolved 11 major lineages. Of 39,404 differentially accessible regions (DARs), more than half arose in pyramidal neurons, whereas dentate gyrus granule neurons and interneurons showed predominantly reduced accessibility; within the pyramidal lineage, remodelling was predominant in newly-formed rather than mature neurons. About 20% of hippocampal candidate cis-regulatory elements (cCREs) overlapped transposable elements (TEs) at baseline, rising to about 50% among regions that gained accessibility upon stimulation, and multiple families were significantly bound above the expected threshold by neurogenic transcription factors, whereby the many neighbouring genes have established roles for neuronal differentiation and synapse formation. These data define TE-derived sequences as candidate components of stimulus-responsive neurogenic regulatory networks and provide a cell-type-specific chromatin resource for the adult hippocampus at single-nucleus resolution.

neuroscience↗

bioRxiv2026

HIF1A recruits primate-specific endogenous retroviruses into the human hypoxic and immune responses

Oxygen availability varies profoundly across the human body and changes further during inflammation, infection, tissue injury and disease. Immune cells must therefore continuously adapt their transcriptional and metabolic state based on the oxygen availability to them. Hypoxia-inducible factor 1 (HIF1A) is central to this adaptation and a marker of the cellular response to low oxygen, yet its genomic targets have been assembled from a non-repetitive fraction of the genome, leaving nearly half of the human genome largely unexplored. Here we define the gene and transposable-element (TE) landscape of the human hypoxic response across different human tissues, cell lines, and conditions. This directional TE response was reproduced in transformed cells and in primary immune cells isolated from blood and the physiologically oxygen-restricted tonsil. Single-cell profiling of peripheral blood mononuclear cells (PBMC) under hypoxia, pharmacological HIF stabilization, and interferon stimulation revealed a striking difference between the gene and retrotranscriptome responses. While gene responses were strongly cell-type dependent and in a bidirectional manner, TEs were overwhelmingly activated. This pattern extended to blood and tonsil immune cells, where ~70-90% of tested TE families were induced under hypoxia, with activated tonsil cells showing exclusively induced significant families, including THE1B, alongside increased LTR7 and HERVH. Integrating HIF1A ChIP-seq with transcriptional responses revealed that HIF1A does not engage repetitive DNA indiscriminately. Instead, its binding converged on LTR7, the promoter long terminal repeat of the HERVH endogenous retrovirus. Approximately 80% of HIF1A-bound LTR7 elements contained a canonical hypoxia-response element, and disruption of HIF1A DNA binding dramatically reduced the expression of occupied HERVH loci. CRISPR deletion of individual LTR7/HERVH loci altered the expression of distant and neighboring genes, demonstrating that hypoxia-responsive retroelements can participate directly in host gene regulation and contribute to overall physiology. Our findings reveal the repetitive genome as a previously underappreciated component of oxygen sensing. We propose that HIF1A recruits selected endogenous retroviral elements into the human hypoxic response, extending oxygen-dependent regulation beyond conventional gene promoters and providing an additional regulatory layer through which tissue oxygenation can shape immune-cell state and human physiology.

genomics↗

bioRxiv2026

Loss-of-function phenomics, ncORFs, and ambiguity of mutant phenotypes in Medicago truncatula

Non-canonical open reading frames (ncORFs) are an emerging area of research that is quickly gaining momentum. Many peptides and proteins missed in initial annotation efforts (ncProts) were subsequently shown to be crucial for a wide range of biological processes. The discovery of ncORFs continues to improve the accuracy of loss-of-function studies because they often occupy the same genomic spaces as annotated ORFs. While databases of mutant phenotypes linked to genomic loci exist in a few species, none of these databases integrate the information on ncORFs present in already characterized loci. In this study, we introduce a nearly comprehensive loss-of-function phenomics dataset of Medicago truncatula (673 loci characterized over the past 30 years), which was integrated as a new track into the genome browser of this organism. This dataset helped critically analyze the potential contribution of ncORFs to published phenotypes. We detected mass spectrometry (MS)-validated ncORFs in 10 characterized genes, including major regulators of development and symbiotic relationships. We also found conserved ncORFs in 113 characterized genes, including four genes with highly conserved ncORFs. In some studies, the contribution of these ncORFs can be ruled out, while in others it cannot. Using real examples, we systematized ambiguities associated with ncORFs. Furthermore, we highlighted little-known trans effects of insertional mutagenesis on splicing as contributors to that ambiguity. Finally, our meta-analysis of published phenotypes revealed that different protein classes have significantly different (unique) proportions of unconditional, conditional, and neutral phenotypes, potentially reflecting their relative functional importance. Significance statementThis study is the first to merge a nearly comprehensive inventory of loss-of-function studies in a eukaryotic organism with the information on novel MS-validated and conserved ncORFs.

genetics↗

bioRxiv2025

Transposable Element-Mediated Epigenomic Remodeling Drives Erythropoietin-Induced Neurogenesis in the Adult Hippocampus

Understanding the molecular mechanisms by which erythropoietin (EPO) acts as neurotrophic factor that enhances hippocampal function and learning is essential to harness its therapeutic potential. Here, we employ single-nucleus ATAC-seq and RNA-seq to map the epigenomic and transcriptional landscapes of adult mouse hippocampus under recombinant human EPO (rhEPO) treatment. We discover significant lineage-specific chromatin remodeling predominantly in newly formed and immature excitatory neurons, highlighting a robust EPO-driven neurogenic response as the first direct evidence that an extrinsic factor can induce adult hippocampal neurogenesis. Notably, many EPO-induced accessible regions overlap ancient transposable elements, particularly ancient LINEs and SINEs, that are bound by key neurogenic transcription factors such as NEUROD1/2, NEUROG2, FOXG1, and ASCL1 and are linked to nearby genes governing neuronal differentiation and synaptic plasticity. Our findings uncover a previously unrecognized transposon-mediated mechanism underlying EPO-induced neurogenesis, highlighting an underappreciated role for TE-derived sequences in this process, and establish a publicly available single-nucleus multiomic atlas as a resource for understanding cell-type-specific gene regulation and neuroplasticity in the adult brain.

neuroscience↗

bioRxiv2025

Erythropoietin alleviates intellectual disability and autism-like behavior of mice caused by Zbtb20 haploinsufficiency, a construct-valid model of Primrose syndrome

Among the known genetic causes of syndromic autism spectrum disorders (ASD) are transcription factor deficiencies. In this regard, haploinsufficiency of the zinc finger and broad complex, tramtrack, bric and brac domain-containing protein 20 (ZBTB20) leads to a prototypical clinical picture, referred to as Primrose syndrome, comprising severe ASD symptoms together with intellectual disability. Here, we present a comprehensive behavioral and phenotypical characterization of Zbtb20+/- mice, a construct valid model of this thus far untreatable human condition. Zbtb20+/- mice exhibit diminished sociability, reduced vocalization, distinct repetitive behaviors, impaired cognitive flexibility, hyperactivity and hypoalgesia. Magnetic resonance imaging reveals increased volumes of hippocampus, cerebellum, brain matter, and whole brain, confirmed by postmortem brain weight measurements. Due to our previous observation of enhanced ZBTB20 expression in CA1 pyramidal neurons upon recombinant human erythropoietin (rhEPO) injections, we anticipated a mitigating effect through rhEPO treatment of Zbtb20 deficiency/Primrose syndrome. Indeed, after three weeks of alternate-day rhEPO injections, a remarkable improvement in the behavioral phenotype was observed. Our results highlight rhEPO as a first promising treatment for Primrose syndrome.

neuroscience↗

bioRxiv2025

Forebrain-specific loss of erythropoietin provokes compensatory upregulation of different EPO receptors

The procognitive growth factor erythropoietin (EPO) and its canonical receptor, EPOR, have long been recognized to be expressed by most cell types in the brain. Cognitive domains, improved by injections of exogenous EPO or by endogenous, hypoxia-stimulated EPO, include important forebrain functions, namely attention, working memory, drive, and executive performance. To gain mechanistic insight into the involvement of forebrain-expressed EPO, we deleted EPO in mice using as specific cre-driver Emx1. Here, we report that these mutant mice act comparably to their wildtype littermates in a comprehensive behavioral test battery. Importantly, we find that the transcripts of both EPOR and a novel, brain-expressed EPO receptor, EphB4, respond to EPO deletion with compensatory upregulation. EphB4 expression in brain and its increase upon forebrain erasure of EPOR are confirmed by in situ hybridization and immunohistochemistry. The augmented expression of both EPOR and EphB4 and their regulatory intercorrelation may explain why EmxEPO mutants show an even superior performance in the most challenging working memory task. Using the previously published single-nuclei-RNA-seq dataset, we further confirm the suggested compensatory mechanism, wherein EPO loss or reduction drives elevated EPOR expression, adding another layer to the intricate regulation of EPO signaling in hippocampal pyramidal neurons. Collectively, these data may explain the lack of behavioral and negative cognitive consequences upon forebrain-wide EPO elimination.

neuroscience↗

bioRxiv2025

ShiftSCAN, a program that predicts potential alternative sources of mass spectrometry-derived peptides, improves the accuracy of studies on novel amino acid sequences

Mass spectrometry (MS) proteomics is currently the most powerful tool for identifying both annotated proteins and proteins translated from non-canonical open reading frames or unusual genetic events. With this method, numerous novel protein-coding loci have been discovered by searching for short fragments of hypothetical longer peptides and polypeptides. Apart from the validation of translation from mRNA transcripts, MS proteomics has been instrumental for the detection of peptides encoded by non-mRNA transcripts. A special application field of MS proteomics is studies on programmed ribosomal frameshifting (PRF), where the detection of chimeric peptides produced from two different reading frames is vital. Each novel chimeric peptide is thought to originate from a certain genetic locus. However, due to the short length of MS peptides, there is a possibility that MS-supported chimeric peptides are produced by additional (alternative) loci via PRF. This scenario evaded due attention because the contribution of non-canonical peptides and proteins to the functional diversity of proteomes is still thought to be minor. Recent studies have challenged this paradigm. To the best of our knowledge, our group was the first to include alternative chimeric sources into the analysis pipeline. This resulted in a much higher certainty about loci responsible for the production of unusual peptides. This certainty is crucial for the functional characterization of such loci. At the same time, our study revealed enormous diversity of potential alternative sources for a subset of MS-supported non-canonical peptides. Here, we present a highly flexible program that predicts alternative chimeric and non-chimeric sources of peptides detected by MS proteomics.

bioinformatics↗

bioRxiv2025

A universal pipeline MosaicProt enables large-scale modeling and detection of chimeric protein sequences for studies on programmed ribosomal frameshifting

Peptides and proteins produced by programmed ribosomal frameshifting (PRF) are well-known in viruses. In non-viral systems, only a few examples of such chimeric sequences have been documented until recently. Two new studies, one in humans and one in plants, showed that chimeric peptides are numerous and diverse. In humans, their discovery was possible due to focusing on sequences with naturally repeated codons. This way, many candidate sequences with mass spectrometry (MS) proteomics-based support for translation have been identified. In the plant study, our group discovered MS-validated chimeric peptides using a unique modeling algorithm, which is described and made available here. Our pipeline enables the identification of chimeric peptides in any organism for which transcript sequences and MS proteomic data are available. By design, our approach does not require prior knowledge about sequence similarity to already characterized PRF sites and can detect forward and backward frameshifts by 1 and 2 nucleotides. Thus, our pipeline opens a path for uncovering previously unknown PRF events across various transcript types, potentially broadening our understanding of proteome diversity.

bioinformatics↗

bioRxiv2024

Discovery of diverse chimeric peptides in a eukaryotic proteome sets the stage for the experimental proof of the mosaic translation hypothesis

The high complexity of eukaryotic organisms enabled their evolutionary success, which became possible due to the diversification of eukaryotic proteomes. Various mechanisms contributed to this process. Alternative splicing had the largest known impact among these mechanisms: tens or hundreds of protein isoforms produced from a single genetic locus. Earlier, we hypothesized that along with alternative splicing, a different but conceptually similar mechanism creates novel versions of existing proteins in all eukaryotes. However, this mechanism acts at the level of translation, where the novelty of an amino acid sequence is achieved via multiple programmed ribosomal frameshifting. This mechanism, which is termed mosaic translation, is very difficult to demonstrate even with the most up-to-date molecular tools. Thus, it remained unnoticed so far. Using only a portion of all mass spectrometry proteomic data generated from various organs of the model plant Medicago truncatula, we attempted the first step toward the experimental proof of this hypothesis. Our original in silico approach resulted in the discovery of two candidates for mosaic proteins (homologs of EF1 and RuBisCo) and 154 candidates for chimeric peptides. Chimeric peptides and polypeptides are produced in the course of one ribosomal frameshifting event and may correspond to parts of mosaic proteins. In addition, our analysis reveals the possibility of translation of chimeric peptides from five ribosomal RNA transcripts, ten long non-coding RNA transcripts, and one transfer RNA transcript. These findings are very novel and will be the basis for experimental validation in future studies. In this work, we present multiple lines of indirect evidence that support the validity of our in silico data.

biochemistry↗