@phdthesis{19763,
  abstract     = {Pattern formation in developing organs is controlled by morphogens. These signalling
molecules form concentration gradients across tissues, thereby providing positional
information that instructs the pattern of cell differentiation. Morphogen gradients are highly
dynamic in space and time. Many factors such as morphogen production, spreading,
degradation, cellular rearrangements and others could contribute to changes in the gradient
shape, yet how the spatiotemporal signalling dynamics arise in many systems is still unclear.
We studied the dynamics of morphogen signalling and tissue patterning in the developing
vertebrate neural tube. In this system, neural crest, roof plate and distinct dorsal progenitor
subtypes are specified in a spatially and temporally ordered manner in response to dorsal-toventral gradients of BMP and WNT signalling activity. How the BMP and WNT gradients are
established and interpreted to ensure ordered cell specification is poorly understood.
To address this question, we developed a 2D embryonic stem cell differentiation system that
captures key features of dorsal neural tube development. In this system, differentiated
colonies display remarkable self-organised pattern formation in response to uniformly
applied BMP ligand. We established a method of differentiating the colonies using
microfabricated stencils, which allowed us to control the initial size and shape of colonies
without confining cell migration and colony growth. This led to highly reproducible pattern
formation that facilitates quantification.
Using this approach, we observed striking two-phase temporal dynamics of BMP signalling in
our colonies: a BMP gradient rapidly forms from the periphery to the centre of colonies,
subsequently disappears and is re-established again in the second phase. By combining our
quantitative data with a data-driven theoretical model, we uncovered a temporal relay
mechanism that underlies this biphasic BMP signalling dynamics. The first signalling phase is
controlled by fast tissue-autonomous negative feedback that restricts the duration of the
initial response to BMP. The early BMP activity gradient moreover controls the spatial
organisation of the cell type pattern: the absence of a first phase results in disordered cell
type pattern. The second phase is controlled by slow positive regulation of BMP signalling by
the transcription factor LMX1A, a key regulator of roof plate identity. WNT promotes the
second phase of BMP signalling via positive feedback on LMX1A.
Altogether, the mechanism that we uncovered ensures the coupling of sequential
developmental events, making pattern formation spatially and temporally organised.
Furthermore, this mechanism allows the BMP signalling pathway to be reused in different
contexts – first for the establishment of the neural plate border, and subsequently for dorsal
neural progenitor patterning. Our study supports a general developmental principle in which
multiple morphogens interact with transcriptional networks resulting in complex
spatiotemporal signalling dynamics that ultimately drive organised pattern formation.},
  author       = {Rus, Stefanie},
  issn         = {2663-337X},
  pages        = {129},
  publisher    = {Institute of Science and Technology Austria},
  title        = {{Dynamics of morphogen signalling and cell fate decisions in the dorsal neural tube}},
  doi          = {10.15479/AT-ISTA-19763},
  year         = {2025},
}

@article{18807,
  abstract     = {Developing tissues interpret dynamic changes in morphogen activity to generate cell type diversity. To quantitatively study bone morphogenetic protein (BMP) signaling dynamics in the mouse neural tube, we developed an embryonic stem cell differentiation system tailored for growing tissues. Differentiating cells form striking self-organized patterns of dorsal neural tube cell types driven by sequential phases of BMP signaling that are observed both in vitro and in vivo. Data-driven biophysical modeling showed that these dynamics result from coupling fast negative feedback with slow positive regulation of signaling by the specification of an endogenous BMP source. Thus, in contrast to relays that propagate morphogen signaling in space, we identify a BMP signaling relay that operates in time. This mechanism allows for a rapid initial concentration-sensitive response that is robustly terminated, thereby regulating balanced sequential cell type generation. Our study provides an experimental and theoretical framework to understand how signaling dynamics are exploited in developing tissues.},
  author       = {Rus, Stefanie and Brückner, David and Minchington, Thomas and Greunz, Martina and Merrin, Jack and Hannezo, Edouard B and Kicheva, Anna},
  issn         = {1534-5807},
  journal      = {Developmental Cell},
  number       = {4},
  pages        = {567--580},
  publisher    = {Elsevier},
  title        = {{Self-organized pattern formation in the developing mouse neural tube by a temporal relay of BMP signaling}},
  doi          = {10.1016/j.devcel.2024.10.024},
  volume       = {60},
  year         = {2025},
}

@article{17148,
  abstract     = {During neural tube (NT) development, the notochord induces an organizer, the floorplate, which secretes Sonic Hedgehog (SHH) to pattern neural progenitors. Conversely, NT organoids (NTOs) from embryonic stem cells (ESCs) spontaneously form floorplates without the notochord, demonstrating that stem cells can self-organize without embryonic inducers. Here, we investigated floorplate self-organization in clonal mouse NTOs. Expression of the floorplate marker FOXA2 was initially spatially scattered before resolving into multiple clusters, which underwent competition and sorting, resulting in a stable “winning” floorplate. We identified that BMP signaling governed long-range cluster competition. FOXA2+ clusters expressed BMP4, suppressing FOXA2 in receiving cells while simultaneously expressing the BMP-inhibitor NOGGIN, promoting cluster persistence. Noggin mutation perturbed floorplate formation in NTOs and in the NT in vivo at mid/hindbrain regions, demonstrating how the floorplate can form autonomously without the notochord. Identifying the pathways governing organizer self-organization is critical for harnessing the developmental plasticity of stem cells in tissue engineering.},
  author       = {Krammer, Teresa and Stuart, Hannah T. and Gromberg, Elena and Ishihara, Keisuke and Cislo, Dillon and Melchionda, Manuela and Becerril Perez, Fernando and Wang, Jingkui and Costantini, Elena and Rus, Stefanie and Arbanas, Laura and Hörmann, Alexandra and Neumüller, Ralph A. and Elvassore, Nicola and Siggia, Eric and Briscoe, James and Kicheva, Anna and Tanaka, Elly M.},
  issn         = {1878-1551},
  journal      = {Developmental Cell},
  number       = {15},
  pages        = {1940--1953.e10},
  publisher    = {Elsevier},
  title        = {{Mouse neural tube organoids self-organize floorplate through BMP-mediated cluster competition}},
  doi          = {10.1016/j.devcel.2024.04.021},
  volume       = {59},
  year         = {2024},
}

@article{18601,
  abstract     = {Geometrically controlled stem cell differentiation promotes reproducible pattern formation. Here, we present a protocol to fabricate elastomeric stencils for patterned stem cell differentiation. We describe procedures for using photolithography to produce molds, followed by molding polydimethylsiloxane (PDMS) to obtain stencils with through holes. We then provide instructions for culturing cells on stencils and, finally, removing stencils to allow colony growth and cell migration. This approach yields reproducible two-dimensional organoids tailored for quantitative studies of growth and pattern formation.
For complete details on the use and execution of this protocol, please refer to Lehr et al.1},
  author       = {Rus, Stefanie and Merrin, Jack and Kulig, Monika Aleksandra and Minchington, Thomas and Kicheva, Anna},
  issn         = {2666-1667},
  journal      = {STAR Protocols},
  number       = {4},
  publisher    = {Elsevier},
  title        = {{Protocol for fabricating elastomeric stencils for patterned stem cell differentiation}},
  doi          = {10.1016/j.xpro.2024.103187},
  volume       = {5},
  year         = {2024},
}

@article{13136,
  abstract     = {Despite its fundamental importance for development, the question of how organs achieve their correct size and shape is poorly understood. This complex process requires coordination between the generation of cell mass and the morphogenetic mechanisms that sculpt tissues. These processes are regulated by morphogen signalling pathways and mechanical forces. Yet, in many systems, it is unclear how biochemical and mechanical signalling are quantitatively interpreted to determine the behaviours of individual cells and how they contribute to growth and morphogenesis at the tissue scale. In this review, we discuss the development of the vertebrate neural tube and somites as an example of the state of knowledge, as well as the challenges in understanding the mechanisms of tissue size control in vertebrate organogenesis. We highlight how the recent advances in stem cell differentiation and organoid approaches can be harnessed to provide new insights into this question.},
  author       = {Minchington, Thomas and Rus, Stefanie and Kicheva, Anna},
  issn         = {2452-3100},
  journal      = {Current Opinion in Systems Biology},
  publisher    = {Elsevier},
  title        = {{Control of tissue dimensions in the developing neural tube and somites}},
  doi          = {10.1016/j.coisb.2023.100459},
  volume       = {35},
  year         = {2023},
}