Engineering topics

Osterwalder, Marco

Publications and source records attributed to Osterwalder, Marco.

4 records

DOE OSTI2025

Uncovering hidden enhancers through unbiased in vivo testing

Chromatin signatures are widely used to identify tissue-specific in vivo enhancers, but their sensitivity and specificity remains unclear. Here we show that many developmental enhancers remain undetectable using currently available chromatin data. In an initial comparison of over 1200 developmental enhancers with tissue-matched chromatin data, 14% (n = 285) lacked canonical enhancer-associated chromatin signatures. To further assess the prevalence of enhancers missed by chromatin profiling approaches, we used a high-throughput transgenic enhancer assay to screen the regulatory landscapes of two key developmental genes at 5 kb resolution, spanning 1.3 Mb of mouse sequence in total. We observed that 23 of 88 (26%) in vivo enhancers discovered by this approach lacked enhancer-associated chromatin signatures in the respective tissue. Our findings suggest the existence of tens of thousands of enhancers that remain undiscovered by currently available chromatin data, underscoring the continued need for expanding resources for enhancer discovery.

Epigenomics↗

DOE OSTI2024

A gene desert required for regulatory control of pleiotropic Shox2 expression and embryonic survival

Approximately a quarter of the human genome consists of gene deserts, large regions devoid of genes often located adjacent to developmental genes and thought to contribute to their regulation. However, defining the regulatory functions embedded within these deserts is challenging due to their large size. Here, we explore the cis-regulatory architecture of a gene desert flanking the Shox2 gene, which encodes a transcription factor indispensable for proximal limb, craniofacial, and cardiac pacemaker development. We identify the gene desert as a regulatory hub containing more than 15 distinct enhancers recapitulating anatomical subdomains of Shox2 expression. Ablation of the gene desert leads to embryonic lethality due to Shox2 depletion in the cardiac sinus venosus, caused in part by the loss of a specific distal enhancer. The gene desert is also required for stylopod morphogenesis, mediated via distributed proximal limb enhancers. In summary, our study establishes a multi-layered role of the Shox2 gene desert in orchestrating pleiotropic developmental expression through modular arrangement and coordinated dynamics of tissue-specific enhancers.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2024

Dynamic enhancer landscapes in human craniofacial development

The genetic basis of human facial variation and craniofacial birth defects remains poorly understood. Distant-acting transcriptional enhancers control the fine-tuned spatiotemporal expression of genes during critical stages of craniofacial development. However, a lack of accurate maps of the genomic locations and cell type-resolved activities of craniofacial enhancers prevents their systematic exploration in human genetics studies. Here, we combine histone modification, chromatin accessibility, and gene expression profiling of human craniofacial development with single-cell analyses of the developing mouse face to define the regulatory landscape of facial development at tissue- and single cell-resolution. We provide temporal activity profiles for 14,000 human developmental craniofacial enhancers. We find that 56% of human craniofacial enhancers share chromatin accessibility in the mouse and we provide cell population- and embryonic stage-resolved predictions of their in vivo activity. Taken together, our data provide an expansive resource for genetic and developmental studies of human craniofacial development.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2023

Identification of ancestral gnathostome Gli3 enhancers with activity in mammals

Abnormal expression of the transcriptional regulator and hedgehog (Hh) signaling pathway effector Gli3 is known to trigger congenital disease, most frequently affecting the central nervous system (CNS) and the limbs. Accurate delineation of the genomic cis-regulatory landscape controlling Gli3 transcription during embryonic development is critical for the interpretation of noncoding variants associated with congenital defects. Here, we employed a comparative genomic analysis on fish species with a slow rate of molecular evolution to identify seven previously unknown conserved noncoding elements (CNEs) in Gli3 intronic intervals (CNE15–21). Transgenic assays in zebrafish revealed that most of these elements drive activities in Gli3 expressing tissues, predominantly the fins, CNS, and the heart. Intersection of these CNEs with human disease associated SNPs identified CNE15 as a putative mammalian craniofacial enhancer, with conserved activity in vertebrates and potentially affected by mutation associated with human craniofacial morphology. Finally, comparative functional dissection of an appendage-specific CNE conserved in slowly evolving fish (elephant shark), but not in teleost (CNE14/hs1586) indicates co-option of limb specificity from other tissues prior to the divergence of amniotes and lobe-finned fish. In conclusion, these results uncover a novel subset of intronic Gli3 enhancers that arose in the common ancestor of gnathostomes and whose sequence components were likely gradually modified in other species during the process of evolutionary diversification.

59 BASIC BIOLOGICAL SCIENCES↗