Biology subjects

Kretzmer, H.

Publications and source records attributed to Kretzmer, H..

13 recordsLinked to original sources

bioRxiv2026

Epigenetic regulators partition the genome by evolutionary and developmental constraint

Many epigenetic regulators play essential roles in animal development, but precisely how they function across different cell types without clear sequence preferences remains a foundational challenge. Here, we employed single-cell RNA sequencing to survey the mutant phenotypes of 30 essential regulatory pathways across over 450 gastrulation stage mouse embryo replicates, including essential factors associated with repressive chromatin states, RNA surveillance/processing, and other genomic roles. We then use this integrated manifold to partition mutant responses into a developmentally-resolved interaction network, revealing a hierarchy of heterochromatin-associated interactions that act orthogonally to Polycomb Repressive Complex-mediated control of embryo patterning. Compared to the embryo proper, we find that the developing placenta and yolk sac can accommodate broad swings in repetitive element expression, a resilience in keeping with the relative evolutionary recency and phenotypic plasticity of these tissues. Dense temporal sampling of Human Silencing Hub (HUSH) mutant embryogenesis further supports the notion that the gastrulating embryo is particularly constrained and depends on multiple layers of insulation, including the need to suppress evolutionarily young genetic elements to stably license highly conserved, phylotypic programs. In combination, our work highlights the power of single cell-resolved developmental genetics to contextualize the roles of epigenetic regulators, which restrict genomic action to support robust cellular identities and embryological forms.

developmental biology↗

bioRxiv2026

Genome-wide multi-layered epigenomic profiling across human aging

Aging is characterized by highly reproducible alterations across multiple layers of the epigenetic landscape, including DNA methylation, chromatin accessibility, and histone modifications. However, it remains unclear to what extent these age-associated changes are interconnected and coordinated coherently. To investigate the genome-wide distribution and interplay of age-associated epigenetic alterations, we generated whole-genome bisulfite sequencing (WGBS) data from blood samples of 120 healthy donors. Integration with ATAC-seq data revealed no clear relationship between age-related changes in DNA methylation and chromatin accessibility. We further examined the association of these alterations with age-dependent changes in CTCF occupancy and histone modifications, including H3K27ac, H3K27me3, H3K4me1, H3K4me3, and H3K9me3, but observed very little corresponding changes in chromatin states. Collectively, our integrative genome-wide analysis revealed only limited association between age-associated epigenetic alterations in DNA methylation, chromatin accessibility, and histone modifications, arguing against a broadly coordinated remodeling of the aging epigenome.

genomics↗

bioRxiv2026

Multimodal single-cell analysis uncovers transcription factor networks underlying T-cell aging

Aging of the immune system is associated with chronic inflammation and impaired immune function, yet the regulatory mechanisms underlying these changes remain incompletely understood. Here, we generated paired single-cell transcriptomic and chromatin accessibility profiles from peripheral blood mononuclear cells of young and old healthy donors to characterize immune aging at single-cell resolution. Using an integrative computational framework for multi-omic single-cell analysis, we detected pronounced age-associated changes in T cells, including loss of naive CD8+ T cells and expansion of differentiated memory and effector populations. Aging was accompanied by increased inflammatory signaling and reduced oxidative phosphorylation programs. Enhancer-based gene regulatory network analyses identified a reduced role of TCF7 and increased activity of inflammatory regulators, including FOSL2, in aged T cells. Integration with genetic association and eQTL datasets further supported the functional relevance of age-associated regulatory regions and their target genes.

bioinformatics↗

bioRxiv2026

Nucleolus transformation and Pol1 activity pattern dual transcriptomes of neuronal subtype identity

Gene expression directs neuronal fate during development with multiple RNA polymerases in the nucleus. mRNA transcribed by Pol2 ultimately depends on the rRNA transcribed by Pol1 in the nucleolus, which generates a ribosome pool to control mRNA translation. How Pol1 activity is synchronized with the Pol2 transcriptome to direct gene expression and neuronal fate is unknown. By creating a high-resolution map of nucleoli in the developing neocortex, we discover that the nucleolus is restructured in neuronal lineages with distinct Pol1 activity. With a new method for rRNA and mRNA dual-transcriptome sequencing in single-cells, we reveal that rRNA transcription is neuronal subtype and stage specific. Rather than a general feature of neural progenitor differentiation, rRNA production peaks transiently in post-mitotic subcortical projection neurons, which have atypical nucleoli containing a single, large cavity. Pol1 inactivation redirects these neurons towards a distinct fate that expresses a hybrid of subcortical and intracortical projection neuron markers. Thus, nucleolar architecture and Pol1 activity pattern dual rRNA-mRNA transcriptome signatures of neuronal subtype identity.

neuroscience↗

bioRxiv2025

Human brain organoids record the passage of time over multiple years in culture

The human brain develops and matures over an exceptionally prolonged period of time that spans nearly two decades of life. Processes that govern species-specific aspects of human postnatal brain development are difficult to study in animal models. While human brain organoids offer a promising in vitro model, they have thus far been shown to largely mimic early stages of brain development. Here, we developed human brain organoids for an unprecedented 5 years in culture, optimizing growth conditions able to extend excitatory neuron viability beyond previously-known limits. Using module scores of maturation-associated genes derived from a time course of endogenous human brain maturation, we show that brain organoids transcriptionally age with cell type-specificity through these many years in culture. Whole-genome methylation profiling reveals that the predicted epigenomic age of organoids sampled between 3 months and 5 years correlates precisely with time spent in vitro, and parallels epigenomic aging in vivo. Notably, we show that in chimeric organoids generated by mixing neural progenitors derived from "old" organoids with progenitors from "young" organoids, old progenitors rapidly produce late neuronal fates, skipping the production of earlier neuronal progeny that are instead produced by their young counterparts in the same co-cultures. The data indicate that human brain organoids can mature and record the passage of time over many years in culture. Progenitors that age in organoids retain a memory of the time spent in culture reflected in their ability to execute age-appropriate, late developmental programs.

neuroscience↗

bioRxiv2025

An advanced head-to-tail mouse embryo model with hypoxia-mediated neural patterning

The developing mammalian embryo is guided by the continuously changing signals that it receives from maternal tissues and its microenvironment. The dynamic cell-cell and cell-environment interactions that together shape the embryo largely remained unexplorable until the advance of stem cell-based embryo models. These revealed the self-organizing properties of cells in response to endogenous and exogenous cues. Among the latter, restricted oxygen (hypoxia) emerged as a critical microenvironmental regulator that influences cell type diversification in multicellular systems. Here we built a modular ESC-based head-to-tail model of mouse embryogenesis by developing an antero-posterior (AP) assembly strategy under hypoxia. These structures called HAP-gastruloids feature stage-appropriate anterior neural tissues that recapitulate the morphological organization and transcriptional identity of fore- and midbrain including spatial organizer regions such as the midbrain-hindbrain boundary. These anterior tissues develop in synchrony with posterior tissues such as the spinal cord, somites, and gut endoderm derivatives, ultimately yielding a unified structure. We show via genetic, environmental, and pharmacological perturbations that timed hypoxia is essential to boost anterior neural cell identities and their patterning through HIF1a and in part by modulating TGF{beta} activity. These results underline the key beneficial role of hypoxia in early development and offer a uniquely modular system to investigate antero-posterior phenotypes for basic discovery and translation.

developmental biology↗

bioRxiv2024

Pipeline Olympics: continuable benchmarking of computational workflows for DNA methylation sequencing data against an experimental gold-standard

DNA methylation is a widely studied epigenetic mark and a powerful biomarker of cell type, age, environmental exposures, and disease. Whole-genome sequencing following selective conversion of unmethylated cytosines into thymines via bisulfite treatment or enzymatic methods remains the reference method for DNA methylation profiling genome-wide. While numerous software tools facilitate processing of DNA methylation sequencing reads, a comprehensive benchmarking study has been lacking thus far. In this study, we systematically compared complete computational workflows for processing DNA methylation sequencing data using a dedicated benchmarking dataset generated with five genome-wide profiling protocols. As an evaluation reference, we employed highly quantitative locus-specific measurements from our preceding benchmark of targeted DNA methylation assays. Based on this experimental gold-standard assessment and several comprehensive metrics, we identified workflows that consistently demonstrated superior performance and revealed major workflow development trends. To facilitate the sustainability of our benchmark, we implemented an interactive workflow execution and data presentation platform, adaptable to user-defined criteria and seamlessly expandable to future software.

bioinformatics↗

bioRxiv2024

The Greenland shark (Somniosus microcephalus) genome provides insights into extremelongevity

The Greenland shark (Somniosus microcephalus) is the longest-lived vertebrate known, with an estimated lifespan of [~] 400 years. Here, we present a chromosome-level assembly of the 6.45 Gb Greenland shark, rendering it one of the largest non-tetrapod genomes sequenced so far. Expansion of the genome is mostly accounted for by a substantial expansion of transposable elements. Using public shark genomes as a comparison, we found that genes specifically duplicated in the Greenland shark form a functionally connected network enriched for DNA repair function. Furthermore, we identified a unique insertion in the conserved C-terminal region of the key tumor suppressor p53. We also provide a public browser to explore its genome.

genomics↗

bioRxiv2022

Discovery and characterization of LNCSOX17 as an essential regulator in human endoderm formation

Long non-coding RNAs (lncRNAs) have emerged as fundamental regulators in various biological processes, including embryonic development and cellular differentiation. Despite much progress over the past decade, the genome-wide annotation of lncRNAs remains incomplete and many known non-coding loci are still poorly characterized. Here, we report the discovery of a previously not annotated lncRNA that is transcribed upstream of the SOX17 gene and located within the same topologically associating domain. We termed it LNCSOX17 and show that it is induced following SOX17 activation but its expression is more tightly restricted to early definitive endoderm. Loss of LNCSOX17 affects crucial functions independent of SOX17 and leads to an aberrant endodermal transcriptome, signaling pathway deregulation and epithelial to mesenchymal transition defects. Consequently, cells lacking the lncRNA cannot further differentiate into more mature endodermal cell types. We identified and characterized LNCSOX17 as an essential new actor in early human endoderm, thereby further expanding the list of functionally important non-coding regulators.

genomics↗

bioRxiv2020

Dissecting the DNA binding landscape and gene regulatory network of p63 and p53

The transcription factor (TF) p53 is the best-known tumor suppressor, but its ancient sibling p63 ({Delta}Np63) is a master regulator of epidermis development and a key oncogenic driver in squamous cell carcinomas (SCC). Despite multiple gene expression studies becoming available in recent years, the limited overlap of reported p63-dependent genes has made it difficult to decipher the p63 gene regulatory network (GRN). In particular, analyses of p63 response elements differed substantially among the studies. To address this intricate data situation, we provide an integrated resource that enables assessing the p63-dependent regulation of any human gene of interest. Here, we use a novel iterative de novo motif search approach in conjunction with extensive publicly available ChIP-seq data to achieve a precise global distinction between p53 and p63 binding sites, recognition motifs, and potential co-factors. We integrate all these data with enhancer:gene associations to predict p63 target genes and identify those that are commonly de-regulated in SCC and, thus, may represent candidates for therapeutic interventions.

genomics↗

bioRxiv2020

Mouse embryonic stem cells self-organize into trunk-like structures with neural tube and somites

Post-implantation embryogenesis is a highly dynamic process comprising multiple lineage decisions and morphogenetic changes inaccessible to deep analysis in vivo. Mouse embryonic stem cells (mESCs) can form aggregates reflecting the post-occipital embryo (gastruloids), but lacking proper morphogenesis. Here we show that embedding of aggregates derived from mESCs in an extracellular matrix compound results in Trunk-Like-Structures (TLS) with a high level of organization comprising a neural tube and somites. Comparative single-cell RNA-seq analysis demonstrates that TLS execute gene-regulatory programs in an embryo-like order, and generate primordial germ cell like cells (PGCLCs). TLS lacking Tbx6 form ectopic neural tubes, mirroring the embryonic mutant phenotype. ESC-derived trunk-like structures thus constitute a novel powerful in vitro platform for investigating lineage decisions and morphogenetic processes shaping the post-implantation embryo. One sentence summaryA platform for generating trunk-like-structures with precursors of spinal cord, bone and muscle from stem cells in a dish

developmental biology↗

bioRxiv2018

Repeat expansion and methylation state analysis with nanopore sequencing

Expansions of short tandem repeats are genetic variants that have been implicated in neuropsychiatric and other disorders but their assessment remains challenging with current molecular methods. Here, we developed a Cas12a-based enrichment strategy for nanopore sequencing that, combined with a new algorithm for raw signal analysis, enables us to efficiently target, sequence and precisely quantify repeat numbers as well as their DNA methylation status. Taking advantage of these single molecule nanopore signals provides therefore unprecedented opportunities to study pathological repeat expansions.

genetics↗

bioRxiv2018

Molecular recording of mammalian embryogenesis

Understanding the emergence of complex multicellular organisms from single totipotent cells, or ontogenesis, represents a foundational question in biology. The study of mammalian development is particularly challenging due to the difficulty of monitoring embryos in utero, the variability of progenitor field sizes, and the indeterminate relationship between the generation of uncommitted progenitors and their progression to subsequent stages. Here, we present a flexible, high information, multi-channel molecular recorder with a single cell (sc) readout and apply it as an evolving lineage tracer to define a mouse cell fate map from fertilization through gastrulation. By combining lineage information with scRNA-seq profiles, we recapitulate canonical developmental relationships between different tissue types and reveal an unexpected transcriptional convergence of endodermal cells from extra-embryonic and embryonic origins, illustrating how lineage information complements scRNA-seq to define cell types. Finally, we apply our cell fate map to estimate the number of embryonic progenitor cells and the degree of asymmetric partitioning within the pluripotent epiblast during specification. Our approach enables massively parallel, high-resolution recording of lineage and other information in mammalian systems to facilitate a quantitative framework for describing developmental processes.

developmental biology↗