A Scube2-Shh feedback loop links morphogen release to morphogen signaling to enable scale invariant patterning of the ventral neural tube

Zach Collins, Kana Ishimatsu, Tony Tsai, Sean Megason

Preprint posted on 13 November 2018

Size matters: a size-dependent factor that scales sonic hedgehog morphogen gradient in early DV patterning.

Selected by Teresa Rayon

Categories: developmental biology


How do morphogen gradients adapt to size differences? Within populations, individuals vary in size, yet their proportions are similar. This process is known as scale-invariant patterning, and it is a crucial process in development [1,2]. A well-characterized example of this mechanism is the patterning of the neural tube, where it has been shown that embryonic proportions are constant in mouse embryos of different sizes, as well as in embryos of different species. In this preprint, Collins et al. investigate the mechanism by which sonic hedgehog (Shh) morphogen gradient scales in the ventral neural tube in zebrafish embryos of different sizes.

To generate embryos of varying sizes, they surgically remove some cells in the blastula stage (prior to neuroectoderm formation); this rendered embryos of smaller sizes but with constant proportions. Quantifications of a Shh target demonstrate that the response is scaled following embryo reduction.

The authors focus their attention on Scube2, a lipid-binding protein required for Shh release non-cell-autonomously, which is expressed in the dorsal and intermediate neural tube. They demonstrate that Scube2 is a diffusible factor distributed throughout the embryo that is secreted from dorsal cells, and it is repressed by Shh signaling.

The authors next wanted to test whether Scube2 expression would scale like Shh reporter genes in sized-reduced embryos. When they measure Scube2 expression in size-reduced embryos, they find that the reduction in Scube2 levels is not scaled: the levels are 50% reduced at DV positions of maximal expression compared to controls, a pattern not found for al scaling invariant genes formerly tested. In addition, when Scube2 is overexpressed in size-reduced embryos, Shh target response is of the same amplitude in embryos of all sizes. This implicates that control of scube2 is responsible for adjusting the Shh signalling gradient in a decreased tissue.


Figure 1. Scube2 expression is size-dependent and required for pattern scaling. Reproduced from Figure 6 of the preprint

Why I chose the paper:

 Embryo development is strikingly robust, and it can cope with variations in size or morphological alterations [3], however the mechanisms that allow embryos to adapt are poorly understood. Collins et al. identify a size-dependent factor that allows the scaling of Shh morphogen gradient in embryos of reduced sizes, adjusting proportionally to the embryonic axis.

The first thing I liked about this work is the surgical method developed in The Megason group to change embryo size and look at size-dependent scaling. They generate perfectly viable and scaled embryos by removing a big percentage of cells at the blastula stage without the need of genetically altering the embryos, which then allows them to perturb different genes.

Their molecular characterization with fish mutants and transgenic reporters is reminiscent of an expander-repressor model [1]. In this model, the morphogen inhibits the expression of an “expander” molecule (Scube2 ), which functions to increase the gradient, holding back morphogen levels at a specific position. What I like about this model is that it does not rely on morphogen diffusion or degradation – that are common molecular properties of proteins regardless of the embryo size – but it relies on the feedback between “expander” and “inhibitor” to continuously adjust the gradient globally. It will be good to see how an expander-repressor model adjusts to their findings and what new predictions we can infer from the model.

Further reading:

Almuedo-Castillo, M., Bläßle, A., Mörsdorf, D., Marcon, L., Soh, G. H., Rogers, K. W., … Müller, P. (2018). Scale-invariant patterning by size-dependent inhibition of Nodal signalling. Nature Cell Biology, 20(9), 1032–1042.

Ben-Zvi, D., & Barkai, N. (2010). Scaling of morphogen gradients by an expansion-repression integral feedback control. Proceedings of the National Academy of Sciences of the United States of America, 107(15), 6924–9.

Umulis, D. M., & Othmer, H. G. (2013). Mechanisms of scaling in pattern formation. Development (Cambridge, England), 140(24), 4830–43.                                         

Questions to the authors:

  1. The neural tube is highly similar in fish and mammals even though there are some differences in the formation of the neural tube. Do the authors think that the expander-repressor model through Scube2 will operate similarly in mammals?
  2. The size reduction of the embryos occurs early in development, before any Shh has been secreted. Do the authors think their identified mechanism can operate after injury, once the morphogen gradient is ongoing?
  3. Invariant scaling achieves the same proportions of embryos that vary in size. Do the authors know if the scaling mechanisms have the same tempo in embryos of different sizes?


  1. Shilo, B.-Z., & Barkai, N. (2017). Developmental Cell Perspective Buffering Global Variability of Morphogen Gradients. Developmental Cell, 40, 429–438.
  2. Garric, L., & Bakkers, J. (2018). Shaping up with morphogen gradients. Nature Cell Biology, 20(9), 998–999.
  3. Lawrence, P. A., & Levine, M. (2006). Mosaic and regulative development: two faces of one coin. Current Biology, 16(7), R236–R239.

Tags: morphogen gradient, neural tube, sonic hedgehog, zebrafish

Posted on: 3 December 2018


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Author's response

Sean Megason shared

Q1. The neural tube is highly similar in fish and mammals even though there are some differences in the formation of the neural tube. Do the authors think that the expander-repressor model through Scube2 will operate similarly in mammals?

By best guess is yes at the big picture level, but there are several potential differences that it would be great if someone would dig into. The first is scaling itself. In mammals, you can make smaller embryos by separating blastomeres at the 2 or 4 cell stage. These go on to make smaller blastocysts but after implantation there is an interesting process of size adjustment in which the smaller embryos catch up in size. It would be interesting to study how smaller embryos know what size to catch-up to and if this catch-up process overlaps with neural patterning and if so what is the role of Scube2. In the paper, we focus on neural patterning in size-reduced embryos because it is a nice experimental system, but we think that this feedback mechanism has broader implications. For example, the neural tube changes in size along the A-P axis and heterozygotes for Sonic hedgehog pattern fine. The scube2 feedback system may be more important for robustness in these contexts than for whole animal size variation. The second is the relationship of patterning and morphogenesis in the mammalian neural tube. Despite a lot of work on neural tube patterning in mouse, we know very little about what cell movements and dynamic gene expression changes within cells are occurring during patterning. Some nice recent work from Phillip Keller’s lab (McDole et al, Cell, 2018) has laid a foundation to address this. It will be nice to see how patterning dynamics at the single-cell level compare across species.

Q2. The size reduction of the embryos occurs early in development before any Shh has been secreted. Do the authors think their identified mechanism can operate after injury, once the morphogen gradient is ongoing?

There are time scales to both the specification of different cell types by different Shh concentrations, and the adjustment of the Shh gradient by Scube2. Our best guess is that both of these are on the 1-2hr scale in zebrafish and these processes may overlap in time. So if the Shh gradient was perturbed right in the midst of patterning, the system described in the paper might not have time to fully readjust. However, it is possible that there are conceptually and potentially mechanistically related feedback systems that ensure proper domain size as the embryo grows after the initial patterning which could fix the pattern.

Q3. Invariant scaling achieves the same proportions of embryos that vary in size. Do the authors know if the scaling mechanisms have the same tempo in embryos of different sizes?

Our previous work on somites (Ishimatsu, Development 2018) showed that somite patterning does happen at the same tempo in smaller embryos. However, for the neural tube, we would expect that smaller neural tubes should pattern faster unless there is a separation in time scales between gradient formation and interpretation. We don’t have clear data on this but our best guess is that there is not–these timescales are on the same order and thus fate specification happens during a dynamically changing rather than steady-state morphogen profile. If indeed, neural patterning happens at a different tempo in smaller embryos whereas the somites are the same, it raises the interesting question of how the rate of development is coordinated across the embryo

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