Biology subjects

Leisegang, M. S.

Publications and source records attributed to Leisegang, M. S..

11 recordsLinked to original sources

bioRxiv2025

Haematopoietic loss of KDM6A impairs cardiac recovery in heartfailure via epigenetic reprogramming of myeloid cells

Clonal haematopoiesis (CH) is recognized as a potent independent risk factor for cardiovascular disease (CVD). While mutations in common CH-associated genes, such as DNMT3A and TET2, have been extensively studied, the pathological roles of other CH mutations remain poorly understood. Among these is KDM6A (UTX), an X-linked histone demethylase recently found to be commonly mutated in patients with heart failure. The mechanistic implications of KDM6A mutations in cardiac dysfunction remain largely unknown. Here, using multi-omics profiling and functional characterisation of murine models and patient-derived data, we demonstrate that haematopoietic loss of KDM6A substantially impairs cardiac recovery following myocardial infarction (MI). KDM6A deficiency enhances systemic and cardiac inflammation, characterized by augmented myeloid cell infiltration into the infarcted murine heart. Single-cell chromatin accessibility and single-cell RNA sequencing analyses revealed profound epigenetic and transcriptional reprogramming in KDM6A-deficient myeloid cells, notably CCR2 recruited macrophages and neutrophils. These cells exhibited heightened inflammatory (Il1b, Nlpr3, Saa3) and chemotactic signatures (Ccr2, Mif, Cxcl12), increased activation of inflammatory transcription factor networks (AP-1, C/EBP), disrupted chromatin architecture, and enhanced glycolytic activity. Clinically, patients with heart failure harbouring KDM6A-driven CH exhibited increased pro-inflammatory monocyte signatures (CCR2, NLPR3, NFKB1, FOS, JUN, IL6R, IL32), underscoring the translational relevance. Integrative analyses further predicted pathogenic crosstalk between KDM6A-mutated monocytes and cardiac resident cells and was experimentally validated by demonstrating that KDM6A-silenced macrophages drive cardiomyocyte hypertrophy and cardiac fibroblast activation. Our findings establish a critical mechanistic link between KDM6A-driven CH, immune dysregulation, and worsened cardiac outcomes post-MI, highlighting novel avenues for personalized therapeutic strategies in heart failure.

cell biology↗

bioRxiv2024

Concerted loss of micro-RNAs uncovers multi-level restructuring of gene regulation in rodents

BackgroundThe regulatory network that coordinates gene expression ultimately determines the phenotype of an organism. Micro-RNAs (miRNAs) are post-transcriptional regulators involved in key biological processes. Lineage-specific losses of multiple miRNA families are rare, and reported cases of multiple miRNA losses coincide with significant changes in gene regulation resulting in body plan modifications. Recently, 15 mammalian miRNA families were found to be missing in the Eumuroidea, the rodent lineage that includes the model organisms mouse and rat. However, the impact of their absence on the gene regulatory networks in this lineage remains unknown. ResultsThe in silico characterization of all 15 miRNAs revealed that their absence is best explained by multiple independent losses. Analyzing their target genes in humans reveals a significant enrichment of GO- terms linked to cellular and developmental processes. Overexpressing two of the co-absent miRNAs, Mir-197 and Mir-769, in human and mouse inducible pluripotent stem cells (iPSCs) resulted in significantly perturbed expression patterns in both species. In silico target site prediction revealed a significant enrichment of direct targets exclusively in the down-regulated genes. Four genes were down-regulated in both mouse and human and maximum parsimony suggests that the corresponding miRNA target sites were already present in the last common ancestor of mammals. The response of these genes to miRNA overexpression in mice, therefore, unveils remnants of the ancient gene regulatory network that have persisted until today. The evolutionary age of these regulatory connections provides initial evidence that the miRNA losses in Eumuroidea must have had consequences for the regulation of gene expression. Notably, we show that the miRNA loss in the Eumuroidea coincides with the largest lineage-specific loss of transcription factors within the mammals. ConclusionsThe impact of miRNA losses has thus far been investigated on a gene-by-gene basis. Our findings indicate that cooperative effects between miRNAs should be considered when assessing the impact of miRNA loss. We provide evidence that the Eumuroidea have modified their gene regulatory networks on two levels, transcriptionally and post-transcriptionally. It will now be interesting to precisely chart the differences in gene regulation and assess their combined impact on the suitability of mice and rats as model systems for human disease.

genetics↗

bioRxiv2024

Detection of statistically robust interactions from diverse RNA-DNA ligation data

Chromatin-localized RNAs play diverse roles in gene regulation and nuclear architecture. Mapping genome-wide RNA-DNA interactions is possible using a variety of molecular methods, including using bridging oligonucleotides to ligate RNA and DNA in proximity. While molecular methods have progressed, a robust computational method for calling biologically meaningful RNA-DNA interactions from these data is lacking. Herein, we present RADIAnT, a reads-to-interactions pipeline for analyzing RNA-DNA ligation data. RADIAnT calls interactions against a dataset-specific, unified background which considers RNA binding site-TSS distance and genomic region bias. By scaling the background by RNA abundance, RADIAnT is sensitive enough to detect specific interactions of lowly expressed transcripts, while remaining specific enough to discount false positive interactions of highly abundant RNAs. RADIAnT outperforms previously proposed methods in the accurate recall of genome-wide Malat1-DNA interactions, and in a use case, was utilized to identify dynamic chromatin-associated RNAs in the physiologically- and pathologically-relevant process of endothelial-to-mesenchymal transition.

bioinformatics↗

bioRxiv2024

GeneCOCOA: Detecting context-specific functions of individual genes using co-expression data

Extraction of meaningful biological insight from gene expression profiling often focuses on the identification of statistically enriched terms or pathways. These methods typically use gene sets as input data, and subsequently return overrepresented terms along with associated statistics describing their enrichment. This approach does not cater to analyses focused on a single gene-of-interest, particularly when the gene lacks prior functional characterization. To address this, we formulated GeneCOCOA, a method which utilizes context-specific gene co-expression and curated functional gene sets, but focuses on a user-supplied gene-of-interest. The co-expression between the gene-of-interest and subsets of genes from functional groups (e.g. pathways, GO terms) is derived using linear regression, and resulting root-mean-square error values are compared against background values obtained from randomly selected genes. The resulting p values provide a statistical ranking of functional gene sets from any collection, along with their associated terms, based on their co-expression with the gene of interest in a manner specific to the context and experiment. GeneCOCOA thereby provides biological insight into both gene function, and putative regulatory mechanisms by which the expression of the gene-of-interest is controlled. Despite its relative simplicity, GeneCOCOA outperforms similar methods in the accurate recall of known gene-disease associations. GeneCOCOA is formulated as an R package for ease-of-use, available at https://github.com/si-ze/geneCOCOA. Author summaryUnderstanding the biological functions of different genes and their respective products is a key element of modern biological research. While one can examine the relative abundance of a gene product in transcriptomics data, this alone does not provide any clue to the biological relevance of the gene. Using a type of analysis called co-expression, it is possible to identify other genes which have similar patterns of regulation to a gene-of-interest, but again, this cannot tell you what a gene does. Genes whose function has previously been studied are often assembled into groups (e.g. pathways, ontologies), which can be used to annotate gene sets of interest. However, if a gene has not yet been characterized, it will not appear in these gene set enrichment analyses. Here, we propose a new method - GeneCOCOA - which uses co-expression of a single gene with genes in functional groups to identify which functional group a gene is most similar too, resulting in a putative function for the gene, even if it has not been studied before. We tested GeneCOCOA by using it to find gene-disease links which have already been scientifically studied, and showed that GeneCOCOA can do this more effectively than other available methods.

bioinformatics↗

bioRxiv2024

Long non-coding RNAs direct the SWI/SNF complex to cell-specific enhancers

The coordination of chromatin remodeling is essential for DNA accessibility and gene expression control1. The highly conserved and ubiquitously expressed SWItch/Sucrose Non-Fermentable (SWI/SNF) chromatin remodeling complex plays a central role in cell type- and context-dependent gene expression2. Despite the absence of a defined DNA recognition motif, SWI/SNF binds lineage specific enhancers genome-wide where it actively maintains open chromatin state2-5. It does so while retaining the ability to respond dynamically to cellular signals4. However, the mechanisms that guide SWI/SNF to specific genomic targets have remained elusive. Here we demonstrate that trans-acting long non-coding RNAs (lncRNAs) direct the SWI/SNF complex to cell type-specific enhancers. SWI/SNF preferentially binds lncRNAs and these predominantly bind DNA targets in trans. Together they localize to enhancers, many of which are cell type-specific. Knockdown of SWI/SNF- and enhancer-bound lncRNAs causes the genome-wide redistribution of SWI/SNF away from enhancers and a concomitant differential expression of spatially connected target genes. These lncRNA-SWI/SNF-enhancer networks support an enhancer hub model of SWI/SNF genomic targeting. Our findings reveal a competitive recruitment of SWI/SNF by lncRNAs which provide a specific and dynamic layer of control in chromatin accessibility and gene expression.

molecular biology↗

bioRxiv2024

The aging-induced long non-coding RNA MIRIAL controls endothelial cell and mitochondrial function

AimsVascular aging is characterized by the progressive deterioration of endothelial function. Long non-coding RNAs (lncRNAs) are critical regulators of gene expression and protein function. However, their involvement in aging-related dysregulation of endothelial cell function remains largely unknown. Here, we aim to characterize the aging-regulated lncRNA MIRIAL in endothelial cells. Methods + ResultsWe identified Mirial as an aging-induced lncRNA in RNA-sequencing data of mouse cardiac endothelial cells. In human umbilical vein endothelial cells (HUVECs), gapmer-mediated knockdown of MIRIAL led to decreases in proliferation, migration and basal angiogenic sprouting. Additionally, MIRIAL knockdown led to increased mitochondrial mass, spare respiratory capacity, and vascular endothelial growth factor (VEGF)-stimulated sprouting. Mechanistically, we demonstrate that MIRIAL forms an RNA{middle dot}DNA:DNA triple helix (triplex) with a regulatory region of the quiescence-promoting Forkhead Box O1 (FOXO1) gene, thus inducing its expression. The formation of this triplex involves an Alu element within the MIRIAL transcript, representing a previously undescribed mechanism of action for a lncRNA. Further, we generated a global Mirial knockout mouse line of. Angiogenic sprouting of aortic rings from Mirial knockout mice was reduced under basal conditions, but increased after VEGF administration, validating the in vitro angiogenic phenotype. Importantly, cardiac contractile function after acute myocardial infarction is severely reduced in Mirial knockout mice, as compared to wild-type littermates. ConclusionsThe lncRNA MIRIAL is an aging-induced regulator of endothelial quiescence and metabolism. Translational PerspectiveLncRNAs often exhibit cell-type or tissue-specific expression and regulation, rendering them potentially druggable targets requiring lower doses and having fewer side effects compared to protein targets. Our current research highlights, that loss of Mirial correlates with adverse outcomes post-acute myocardial infarction in a murine model. Dysregulation of MIRIAL in various human pathological conditions, such as ischemic heart disease, abdominal aortic aneurysm, cancer, and aging, indicates its potential as a diagnostic marker. Mechanistically, MIRIAL regulates endothelial quiescence by modulating FOXO1 expression, suggesting it as a promising therapeutic target to counteract the age-related decline in endothelial cell function.

cell biology↗

bioRxiv2022

Circular RNA circPLOD2 regulates pericyte function by targeting the transcription factor KLF4

Circular RNAs (circRNAs) are generated by back-splicing and control cellular signaling and phenotypes. Pericytes stabilize the capillary structure and play an important role in the formation and maintenance of new blood vessels. Here, we characterized hypoxia-regulated circRNAs in human pericytes and showed that circPLOD2 is induced by hypoxia and regulates pericyte function. Silencing of circPLOD2 increased pericyte proliferation, endothelial-pericyte interaction and tube formation. Transcriptional profiling of circPLOD2-depleted cells and epigenomic analyses revealed widespread changes in gene expression and identified the circPLOD2-dependent regulation of the transcription factor KLF4 as a key effector of these changes. Importantly, overexpression of KLF4 was sufficient to reverse the effects on pericyte proliferation and endothelial-pericyte interactions observed after circPLOD2 depletion. Together, these data revealed a novel function of circPLOD2 in the control of pericyte proliferation and capillary formation and showed that circPLOD2-mediated regulation of KLF4 significantly contributes to the transcriptional response to hypoxia. HighlightsO_LIcircPLOD2 is upregulated in hypoxia in human vascular pericytes C_LIO_LISelective depletion of circPLOD2, but not linear PLOD2 mRNA, changes pericyte migration and endothelial-pericyte interaction C_LIO_LIcircPLOD2 depletion triggers widespread changes in gene expression that are mirrored in the transcriptional hypoxia response C_LIO_LIEpigenomic analyses pinpoint the transcription factor KLF4 as a central player in circPLOD2-mediated expression changes C_LIO_LIKLF4 overexpression is sufficient to rescue the changes in pericyte function caused by circPLOD2 depletion C_LI

molecular biology↗

bioRxiv2022

The lncRNA landscape of cardiac resident macrophages and identification of Schlafenlnc as a regulator of macrophage migratory function

Cardiac resident macrophages (crMPs) were recently shown to exert pivotal functions in cardiac homeostasis and disease, but the underlying molecular mechanisms are largely unclear. Long non-coding RNAs (lncRNAs) are increasingly recognized as important regulatory molecules in a number of cell types, but neither the identity nor the molecular mechanisms of lncRNAs in crMPs are known. Here, we have employed deep RNA-seq and single cell RNA sequencing to resolve the crMP lncRNA landscape from healthy and diseased murine myocardium. CrMPs express previously unknown and highly cell type-specific lncRNAs, among which one lncRNA, termed Schlafenlnc, was particularly abundant and enriched in crMPs. We found Schlafenlnc to be necessary for migration-associated gene expression in macrophages in vitro and in vivo and essential for their adhesion and migration. Collectively, our data provide a basis to the systematic characterization of lncRNAs in crMPs and establish Schlafenlnc as a critical regulator of macrophage migratory functions.

cell biology↗

bioRxiv2022

A novel role for cystathionine gamma lyase in the control of p53: impact on endothelial senescence and metabolic reprograming

AimsAdvanced age is unequivocally linked to the development of cardiovascular disease, however, the mechanisms leading to loss of endothelial cell regenerative capacity during aging remain poorly understood. Here we aimed to investigate novel mechanisms involved in endothelial cell senescence, that impact on endothelial cell transcription and the vascular repair response upon injury Methods and resultsRNA sequencing of a unique collection of native endothelial cells from young and aged individuals, showed that aging (20 vs. 80 years) is characterized by p53- mediated reprogramming to promote the expression of senescence-associate genes. Molecular analysis revelead that p53 accumulated and acetylated in the nucleus of aged human endothelial cells to suppress glycolysis. Metabolic flux analysis identified an associated reduction in glucose uptake and ATP availability that inhibited the assembly of the telomerase complex, which was essential for proliferation. Nuclear translocation of p53 in aged endothelial cells was attributed to the loss of the vasoprotective enzyme, cystathionine {gamma}-lyase (CSE), which physically anchored p53 in the cytosol. In mice, loss of endothelial cell CSE activated p53 and arrested vascular repair upon injury, while the AAV9 mediated re-expression of an active CSE mutant retained p53 in the cytosol, maintained endothelial glucose metabolism and proliferation, and prevented endothelial cell senescence. Adenoviral overexpression of CSE in human native aged endothelial cells maintained low p53 activity and re-activated telomerase to revert endothelial cell senescence. ConclusionOur data identified the interaction between CSE and p53 as a promising target to preserve vascular regeneration during aging. Key QuestionTo identify the mechanisms that regulate endothelial cell senescence under native conditions and their impact on vascular repair in aging. Key FindingLack of a physical interaction between CSE and p53 metabolically reprogrammes endothelial cells to reduce telomerase activity and halt endothelial cell regeneration. Take home messageInterventions to increase CSE expression represent a novel therapy against p53-induced endothelial cell cycle arrest and senescense Translational perspectiveEndothelial rejuvenation strategies could serve as promising therapies against age-related cardiovascular diseases. By investigating human native endothelial cells from young and aged individuals, we identified that the age-related nuclear accumulation of p53 reprograms endothelial cell metabolism, regulates telomerase activity and inhibits endothelial cell regeneration. Nuclear localization of p53 resulted from a loss of its interaction with the cysteine catabolizing enzyme cystathionine {gamma}-lyase in the cytoplasm. Enhancing the physical interaction of p53 with CSE by gene therapy could revert endothelial cell senescence and activate endothelial reparative responses.

cell biology↗

bioRxiv2022

The endothelial-specific LINC00607 mediates endothelial angiogenic function

Long non-coding RNAs (lncRNAs) can act as regulatory RNAs which, by altering the expression of target genes, impact on the cellular phenotype and cardiovascular disease development. Endothelial lncRNAs and their vascular functions are largely undefined. Deep RNA-Seq and FANTOM5 CAGE analysis revealed the lncRNA LINC00607 to be highly enriched in human endothelial cells. LINC00607 was induced in response to hypoxia, arteriosclerosis regression in non-human primates and also in response to propranolol used to induce regression of human arteriovenous malformations. siRNA knockdown or CRISPR/Cas9 knockout of LINC00607 attenuated VEGF-A-induced angiogenic sprouting. LINC00607 knockout in endothelial cells also integrated less into newly formed vascular networks in an in vivo assay in SCID mice. Overexpression of LINC00607 in CRISPR knockout cells restored normal endothelial function. RNA- and ATAC-Seq after LINC00607 knockout revealed changes in the transcription of endothelial gene sets linked to the endothelial phenotype and in chromatin accessibility around ERG-binding sites. Mechanistically, LINC00607 interacted with the SWI/SNF chromatin remodeling protein BRG1. CRISPR/Cas9-mediated knockout of BRG1 in HUVEC followed by CUT&RUN revealed that BRG1 is required to secure a stable chromatin state, mainly on ERG-binding sites. In conclusion, LINC00607 is an endothelial-enriched lncRNA that maintains ERG target gene transcription by interacting with the chromatin remodeler BRG1.

molecular biology↗

bioRxiv2021

HIF1α-AS1 is a DNA:DNA:RNA triplex-forming lncRNA interacting with the HUSH complex

DNA:DNA:RNA triplexes that are formed through Hoogsteen base-pairing have been observed in vitro, but the extent to which these interactions occur in cells and how they impact cellular functions remains elusive. Using a combination of bioinformatic techniques, RNA/DNA pulldown and biophysical studies, we set out to identify functionally important DNA:DNA:RNA triplex-forming long non-coding RNAs (lncRNA) in human endothelial cells. The lncRNA HIF1-AS1 was retrieved as a top hit. Endogenous HIF1-AS1 reduced the expression of numerous genes, including EPH Receptor A2 and Adrenomedullin through DNA:DNA:RNA triplex formation by acting as an adapter for the repressive human silencing hub complex (HUSH). Moreover, the oxygen-sensitive HIF1-AS1 was down-regulated in pulmonary hypertension and loss-of-function approaches not only resulted in gene de-repression but also enhanced angiogenic capacity. As exemplified here with HIF1-AS1, DNA:DNA:RNA triplex formation is a functionally important mechanism of trans-acting gene expression control.

molecular biology↗