We use and develop methods in single cell genomics to study cell fate decisions in health and disease. We integrate these approaches with light microscopy and tools from zebrafish developmental biology. Our main model system is the zebrafish, where we investigate embryonic development as well as adult organ regeneration. In embryos we focus on understanding how variability and plasticity of cell fate decisions are linked to developmental robustness. In adult fish we study mechanisms of organ regeneration in heart, brain and pancreas. Our work combines biophysics, systems biology and developmental biology.

Method development

Single-cell genomics, and in particular single-cell RNA-sequencing have emerged as powerful methods to dissect cellular complexity and dynamics. By analyzing all genes in thousands of cells, cellular heterogeneity and transitions that are missed in bulk analysis and in traditional approaches based on marker genes, can be resolved. Recent methodological efforts focus on combined measurement of multiple modalities and on computational integration of different data types such as open chromatin, transcriptome, and protein measurements. While these approaches have been instrumental for achieving unified and multimodal definitions of cellular identity, two additional layers of information are needed to understand how cells interact with each other: space and time. Method development in our focuses on inclusion of spatiotemporal information in single-cell RNA-seq data.

  • Junker et al., Cell, 2014: tomo-seq, a method for spatial transcriptomics in 3D
  • Spanjaard et al., Nat Biotech, 2018: Simultaneous lineage tracing and cell-type identification using CRISPR scars.
  • Holler et al., Nat Commun, 2021: Spatio-temporal mRNA tracking in the early zebrafish embryo – spatial transcriptomics with sub-single cell resolution and RNA labeling in vivo.
  • Neuschulz et al., in revision: A single-cell RNA labeling strategy for measuring stress response upon tissue dissociation.

Embryonic development

Embryonic development is a study in contrasts between variation and stability. On the one hand, generation of complex spatial patterns proceeds with striking precision: the different tissues and organs are generally formed at the correct position, at the right time, and with a defined size. On the other hand, regulation should also not be too rigid, since embryos need to flexibly adjust to environmental perturbations and correct errors caused by noisy gene expression. We study the interplay between variation and stability by using the zebrafish as a model system.

Organ regeneration

Zebrafish can efficiently regenerate several organs that in humans have only limited regenerative capacity, such as the heart, brain and pancreas. Understanding regenerative processes is of great importance for basic as well as translational research. Interestingly, regeneration in these organs proceeds via different paradigms: From the heart, where cardiomyocyte pools are replenished by dedifferentiation and proliferation; to the brain, where neural stem cells generate new neurons; and the pancreas, where ablation of beta cells triggers transdifferentiation of alpha and delta cells into beta cells. We address organ regeneration as an emergent phenomenon that is based on the collective action of multiple activated cell states. By combining strategies for spatio-temporal single-cell RNA-seq with functional perturbation experiments, we aim to understand cell-cell communication and gene regulation during regeneration of adult organs.

  • Hu et al., Nat Genet, 2022: Identification of an activated fibroblast state with a pro-regenerative function in the zebrafish heart: single-cell RNA-seq, CRISPR lineage tracing, microscopy, and target cell ablation.
  • Mitic et al., in preparation: Dissecting the spatiotemporal diversity of radial glia states in the adult zebrafish brain.
  • Mintcheva et al., in preparation: Identification of the sentinel cells that first respond to heart injury based on in vivo RNA labeling.

Additional systems

In collaboration with other labs, we seek to apply our methods and concepts to other systems beyond zebrafish development and regeneration. This includes work in zebrafish cancer models, mice, organoids, and patient samples.

  • Fresmann et al., in preparation: Neuroblastoma cell state plasticity and its link to microenvironment and disease progression.
  • Olivares-Chauvet et al., in preparation: Optimized CRISPR lineage tracing systems in mice and organoids.
cartoon methods
Selected experimental methods. A. Tomo-seq method for spatially resolved transcriptomics (Junker et al., Cell, 2014). Three individual zebrafish embryos are sectioned along the three main body axes. By sequencing the mRNA from each slice we obtain three spatially-resolved datasets in 1D. We use computational approaches to then reconstruct the original 3D image. B. Massively parallel single-cell lineage tracing (Spanjaard et al., Nature Biotechnology, 2018). With the help of CRISPR/Cas9 technology, we introduce molecular barcodes into single cells at around the 1000 cell stage. Barcoded cells are indicated as circles with different colors. Barcodes are transmitted to the progeny of the labeled cells, and can be read out by single-cell RNA-seq of dissociated fish.

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