Ashley Marie Maynard

Last Name

Maynard

First Name

Ashley Marie

ORCID

0000-0002-3588-3080

Show all metadata

Search Results

Publications 1 - 8 of 8

Single-cell analyses of axolotl telencephalon organization, neurogenesis, and regeneration
Lust, Katharina; Maynard, Ashley Marie; Gomes, Tomas; et al. (2022)
Science
Salamanders are tetrapod models to study brain organization and regeneration; however, the identity and evolutionary conservation of brain cell types are largely unknown. We delineated the cell populations in the axolotl telencephalon during homeostasis and regeneration using single-cell genomic profiling. We identified glutamatergic neurons with similarities to amniote neurons of hippocampus, dorsal and lateral cortex, and conserved γ-aminobutyric acid-releasing (GABAergic) neuron classes. We inferred transcriptional dynamics and gene regulatory relationships of postembryonic, region-specific neurogenesis and unraveled conserved differentiation signatures. After brain injury, ependymoglia activate an injury-specific state before reestablishing lost neuron populations and axonal connections. Together, our analyses yield insights into the organization, evolution, and regeneration of a tetrapod nervous system.
Molecular Landscape of Neurogenesis in the Vertebrate Brain: Insights from Human Organoids and Axolotls
Maynard, Ashley Marie (2023)
The focal point of this thesis is neurogenesis, a fundamental process responsible for the generation of new neurons in the nervous system. Our exploration delves into the intricate aspects of neurogenesis occurring during brain development, in the post-embryonic stage, and throughout regeneration. To achieve this comprehensive investigation, we utilized two distinct models: human brain organoids derived from induced pluripotent stem cells (iPSCs) and the axolotl animal model. Leveraging a diverse toolbox of techniques, this research uncovers valuable insights into lineage dynamics, gene regulatory relationships, cellular states, and spatial positioning, all of which play crucial roles in the intricate process of neurogenesis. To gain a deeper understanding of developmental neurogenesis, we developed iTracer, a novel lineage tracing tool that combines reporter barcodes with inducible CRISPR-Cas9 scarring, enabling direct measurement of lineage relationships using single-cell transcriptomic analyses. Through the application of iTracer in brain organoid development, we uncovered a distinct time window of fate restriction and intriguing variations in the neurogenic dynamics among progenitor neuron families. Taking our investigation even further, we enhanced the capabilities of iTracer by integrating it with sequencing-based spatial transcriptomics, resulting in the creation of spatial iTracer, which allowed us to reveal regional clonality in the developing brain organoid, shedding light on the spatial organization of neurogenic processes. Furthermore, our quest for comprehensive insights led to the development of iTracer-perturb, which facilitates CRISPR-based gene perturbation while simultaneously recording lineage information. The use of iTracer-perturb suggests potential implications on metabolism and neural development delay related to TSC2 gene perturbations. Single-cell genomic profiling is employed to comprehensively identify cell populations in the axolotl telencephalon during both homeostatic conditions and regeneration. The study uncovers glutamatergic neurons that exhibit similarities to amniote neurons found in the hippocampus, dorsal, and lateral cortex. Furthermore, the presence of conserved γ-aminobutyric acid-releasing (GABAergic) neuron classes is identified. Transcriptional dynamics and gene regulatory relationships during post-embryonic neurogenesis provides critical insights into conserved differentiation signatures within specific brain regions during homeostatic conditions. Notably, after brain injury, ependymoglia, which we identify as bonafide stem cells of the axolotl brain, transition into an injury-specific state before contributing to the reestablishment of lost neuron populations and their associated axonal connections, showcasing the remarkable regenerative potential of axolotl brains. In summary, the findings from this work have significant implications for our understanding of neurogenesis in both human and axolotl models. The iTracer lineage tracing tool developed in Chapter 2 provides a powerful method for investigating lineage dynamics and can be broadly applied in other iPSC derived culture systems. Additionally, the single-cell genomic profiling in the axolotl telencephalon sheds light on the organization and evolution of a tetrapod nervous system, offering valuable insights into the cellular heterogeneity and differentiation processes. The robustness of injury-induced neurogenesis in axolotls, which stands in contrast to the limited regenerative capacity of neurons in mammalian brains, raises intriguing questions about the microenvironment and regulatory factors, which will be subject to future research.
Human melanocyte development and melanoma dedifferentiation at single-cell resolution
Belote, Rachel L.; Le, Daniel; Maynard, Ashley Marie; et al. (2021)
Nature Cell Biology
In humans, epidermal melanocytes are responsible for skin pigmentation, defence against ultraviolet radiation and the deadliest common skin cancer, melanoma. Although there is substantial overlap in melanocyte development pathways between different model organisms, species-dependent differences are frequent and the conservation of these processes in human skin remains unresolved. Here, we used a single-cell enrichment and RNA-sequencing pipeline to study human epidermal melanocytes directly from the skin, capturing transcriptomes across different anatomical sites, developmental age, sexes and multiple skin tones. We uncovered subpopulations of melanocytes that exhibit anatomical site-specific enrichment that occurs during gestation and persists through adulthood. The transcriptional signature of the volar-enriched subpopulation is retained in acral melanomas. Furthermore, we identified human melanocyte differentiation transcriptional programs that are distinct from gene signatures generated from model systems. Finally, we used these programs to define patterns of dedifferentiation that are predictive of melanoma prognosis and response to immune checkpoint inhibitor therapy.
Lineage recording in human cerebral organoids
He, Zhisong; Maynard, Ashley Marie; Jain, Akanksha; et al. (2022)
Nature Methods
Induced pluripotent stem cell (iPSC)-derived organoids provide models to study human organ development. Single-cell transcriptomics enable highly resolved descriptions of cell states within these systems; however, approaches are needed to directly measure lineage relationships. Here we establish iTracer, a lineage recorder that combines reporter barcodes with inducible CRISPR–Cas9 scarring and is compatible with single-cell and spatial transcriptomics. We apply iTracer to explore clonality and lineage dynamics during cerebral organoid development and identify a time window of fate restriction as well as variation in neurogenic dynamics between progenitor neuron families. We also establish long-term four-dimensional light-sheet microscopy for spatial lineage recording in cerebral organoids and confirm regional clonality in the developing neuroepithelium. We incorporate gene perturbation (iTracer-perturb) and assess the effect of mosaic TSC2 mutations on cerebral organoid development. Our data shed light on how lineages and fates are established during cerebral organoid formation. More broadly, our techniques can be adapted in any iPSC-derived culture system to dissect lineage alterations during normal or perturbed development.
Single-cell genomic profiling to study regeneration
Maynard, Ashley Marie; Soretić, Mateja; Treutlein, Barbara (2024)
Current Opinion in Genetics & Development
Regenerative capacities and strategies vary dramatically across animals, as well as between cell types, organs, and with age. In recent years, high-throughput single-cell transcriptomics and other single-cell profiling technologies have been applied to many animal models to gain an understanding of the cellular and molecular mechanisms underlying regeneration. Here, we review recent single-cell studies of regeneration in diverse contexts and summarize key concepts that have emerged. The immense regenerative capacity of some invertebrates, exemplified by planarians, is driven mainly by the differentiation of abundant adult pluripotent stem cells, whereas in many other cases, regeneration involves the reactivation of embryonic or developmental gene-regulatory networks in differentiated cell types. However, regeneration also differs from development in many ways, including the use of regeneration-specific cell types and gene regulatory networks.
Ageing compromises mouse thymus function and remodels epithelial cell differentiation
Baran-Gale, Jeanette; Morgan, Michael D.; Maio, Stefano; et al. (2020)
eLife
Ageing is characterised by cellular senescence, leading to imbalanced tissue maintenance, cell death and compromised organ function. This is first observed in the thymus, the primary lymphoid organ that generates and selects T cells. However, the molecular and cellular mechanisms underpinning these ageing processes remain unclear. Here, we show that mouse ageing leads to less efficient T cell selection, decreased self-antigen representation and increased T cell receptor repertoire diversity. Using a combination of single-cell RNA-seq and lineage-tracing, we find that progenitor cells are the principal targets of ageing, whereas the function of individual mature thymic epithelial cells is compromised only modestly. Specifically, an early-life precursor cell population, retained in the mouse cortex postnatally, is virtually extinguished at puberty. Concomitantly, a medullary precursor cell quiesces, thereby impairing maintenance of the medullary epithelium. Thus, ageing disrupts thymic progenitor differentiation and impairs the core immunological functions of the thymus.
Reconstructing cell interactions and state trajectories in pancreatic cancer stromal tumoroids
Okuda, Ryo; Gjeta, Bruno; Popovic, Doris; et al. (2022)
bioRxiv
Interlineage communication within a cancer microenvironment can augment cancer cell behaviour and impact response to therapy. Patient-derived cancer organoids provide an opportunity to explore cancer cell biology, however it is a major challenge to generate a complex cancer microenvironment in vitro. Here, we established a stromal tumoroid culture system modeling pancreatic ductal adenocarcinoma (PDAC) that reconstitutes multilineage interactions between cancer, endothelial, and fibroblast cells and recapitulates several aspects of primary tumors. Whole-mount immunohistochemistry on cleared tumoroids reveals organized vessel, desmoplastic fibroblast, and glandular cancer cell phenotypes that emerge over time. Time-course scRNA-seq measurements show that tumoroid formation activates fibroblasts, altering the extracellular matrix (ECM) composition and inducing cancer cell signal-response signatures and metabolic state change. Comparison between tumoroids with normal or cancer associated fibroblasts (CAFs) reveals different ECM compositions, as well as differential effects on cancer cell behaviors and metabolism. We identify Syndecan 1 (SDC1) and Peroxisome proliferator-activated receptor gamma (PPARG) as receptor and metabolic nodes involved in cancer cell response to CAF signals, and blocking SDC1 disrupts cancer cell growth within the tumoroid. Tumoroids from multiple PDAC patients revealed co-existence of subpopulations associated with classical and basal phenotypes, and CAF-induced migration behaviors emerged in certain patient tumoroids. Comparisons between patient tumoroids revealed a multigene migration signature that develops over time reflecting a stress response mechanism that correlates with worse clinical outcome. Altogether, stromal tumoroids can be used to explore dynamic and reciprocal interactions between cancer, CAF and endothelial cell states, and our data provides new inroads into the discovery of personalized pancreatic cancer therapies.
Lineage recording reveals dynamics of cerebral organoid regionalization
He, Zhisong; Gerber, Tobias; Maynard, Ashley Marie; et al. (2020)
bioRxiv
Diverse regions develop within cerebral organoids generated from human induced pluripotent stem cells (iPSCs), however it has been a challenge to understand the lineage dynamics associated with brain regionalization. Here we establish an inducible lineage recording system that couples reporter barcodes, inducible CRISPR/Cas9 scarring, and single-cell transcriptomics to analyze lineage relationships during cerebral organoid development. We infer fate-mapped whole organoid phylogenies over a scarring time course, and reconstruct progenitor-neuron lineage trees within microdissected cerebral organoid regions. We observe increased fate restriction over time, and find that iPSC clones used to initiate organoids tend to accumulate in distinct brain regions. We use lineage-coupled spatial transcriptomics to resolve lineage locations as well as confirm clonal enrichment in distinctly patterned brain regions. Using long term 4-D light sheet microscopy to temporally track nuclei in developing cerebral organoids, we link brain region clone enrichment to positions in the neuroectoderm, followed by local proliferation with limited migration during neuroepithelial formation. Our data sheds light on how lineages are established during brain organoid regionalization, and our techniques can be adapted in any iPSC-derived cell culture system to dissect lineage alterations during perturbation or in patient-specific models of disease.

Publications 1 - 8 of 8

Ashley Marie Maynard

Last Name

First Name

ORCID

Organisational unit

Filters

Search Results