- Email: [email protected]
Spinal Cords Built to Scale
Developmental Cell
Previews F-actin in the zippering cells (Hashimoto et al., 2015). Ogura and Sasakura (2016) show this to be blocked when epidermal cell-cycle timing is manipulated, but the causal link between cell-cycle timing and F-actin accumulation remains unclear. One intriguing possibility is that the epidermis may be actively pushing dorsomedially to bring the neural folds together and that control of the epidermal cell cycle is essential for maintaining the balance of mechanical forces in the ectoderm. The small, simple Ciona embryo will be a powerful model to investigate the molecular details of how S-phase length is spatially patterned and the mechanistic details of how patterns of cell division in
the epidermis influence neural tube closure. The ability to easily image the full anterior-to-posterior length of a neurulating chordate embryo with subcellular detail made it more feasible to first detect this unusual cell-cycle compensation phenomenon in Ciona, but it will be interesting and important to determine whether similar mechanisms are used in other taxa.
Duncan, T., and Su, T.T. (2004). Curr. Biol. 14, R305–R307. Edgar, B.A., Lehman, D.A., and O’Farrell, P.H. (1994). Development 120, 3131–3143. Farrell, J.A., Shermoen, A.W., Yuan, K., and O’Farrell, P.H. (2012). Genes Dev. 26, 714–725. Hashimoto, H., Robin, F.B., Sherrard, K.M., and Munro, E.M. (2015). Dev. Cell 32, 241–255. Ogura, Y., and Sasakura, Y. (2016). Dev. Cell 37, this issue, 148–161.
REFERENCES
Ogura, Y., Sakaue-Sawano, A., Nakagawa, M., Satoh, N., Miyawaki, A., and Sasakura, Y. (2011). Development 138, 577–587.
Bouldin, C.M., and Kimelman, D. (2014). Cell Cycle 13, 2165–2171.
Stolfi, A., and Christiaen, L. (2012). Genetics 192, 55–66.
Dumollard, R., Hebras, C., Besnardeau, L., and McDougall, A. (2013). Dev. Biol. 384, 331–342.
Veeman, M., and Reeves, W. (2015). Genesis 53, 143–159.
Spinal Cords Built to Scale Benjamin L. Allen1,* 1Department of Cell and Developmental Biology, University of Michigan, Ann Arbor, MI 48109, USA *Correspondence: [email protected] http://dx.doi.org/10.1016/j.devcel.2016.04.007
An outstanding question in embryogenesis is how different-sized animals maintain similarly proportioned body plans. In this issue of Developmental Cell, Uygur et al. (2016) tackle this issue in the context of neural tube patterning, discovering that differential sensitivity to the Hedgehog pathway may provide one answer to this fundamental issue. One of the fundamental questions in developmental biology is: what regulates size? Cell size, organ size, and embryo size can all be dynamically regulated both within and across species; however, the mechanisms that control these processes remain largely unexplored, particularly in the context of the whole embryo, where the maintenance of similarly proportioned body plans in the face of significant differences in organism size is referred to as scaling. Recent studies have addressed scaling in the context of early, BMP-dependent dorsoventral patterning in the developing Xenopus embryo (Inomata et al., 2013), as well as via the Bicoid morphogen gradient in Drosophila embryos (Wu et al., 2015). This question has also been addressed during dorsoventral patterning of the vertebrate neural tube, where recent work suggested that altered differentia108 Developmental Cell 37, April 18, 2016
tion in neural cell progenitors is essential for regulating scaling in this tissue (Kicheva et al., 2014). Despite these studies, the fundamental mechanisms that control scaling in related organisms of different sizes remain unclear. In this issue of Developmental Cell, Uygur et al. (2016) investigate the role of the Hedgehog signaling pathway in scaling of the neural tube across different avian species, specifically comparing spinal cord patterning in zebra finch and chicken embryos, two species of significantly different embryonic and adult size. The authors initially analyzed the expression of three transcription factors (NKX2.2, OLIG2, and NKX6.1) that play essential roles in mediating cell fate specification in the neural tube and are themselves direct transcriptional targets of the HH pathway (Oosterveen et al., 2012; Peterson et al., 2012). Strikingly, patterning occurs faster
and across a smaller tissue field in the zebra finch embryo than in the chicken embryo. One might expect that the faster patterning in the zebra finch embryos could be due to relatively higher levels of HH ligand production; however, Sonic HH (SHH) levels are actually lower in the zebra finch neural tube compared to the chicken neural tube. These data suggested that zebra finch neuroepithelial cells are more sensitive to SHH levels than their chicken counterparts. To test this hypothesis, the authors obtained transgenic lines of both species that ubiquitously express GFP. From these animals, the authors created chimeric neural tubes, transplanting GFP-positive donor tissue into GFP-negative host tissue within neural tubes of embryos from the opposite species. Consistent with their hypothesis, the zebra finch neuroepithelial cells are more sensitive to endogenous SHH ligand than
Developmental Cell
Previews the corresponding chicken neuroepithelia. To confirm and extend these data, the authors also utilized naive intermediate neural explants isolated from both zebra finch and chicken embryos in combination with recombinant SHH ligand to demonstrate that the zebra finch neuroepithelium activates higher-level HH pathway targets (e.g., NKX2.2) in response to lower levels of HH ligand. In a final set of experiments, in ovo electroporations were performed with a HH-responsive GFP reporter to demonstrate a greater range of HH pathway activity across the dorsoventral axis in the zebra finch neural tube compared to the chicken neural tube. The use of careful comparative biology combined with classic developmental biology approaches thus identifies a novel role for morphogen signaling in scaling of the embryonic neural tube. So, while differential SHH sensitivity appears to play a role in scaling, an important question remains: what regulates the differential responsiveness to SHH in the zebra finch neural tube? The authors initially tackled this question using a pharmacologic tool, SAG, a smoothened agonist that activates HH signaling downstream of ligand. Surprisingly, the data suggested that the altered HH response is downstream of Smoothened; in fact, subsequent experiments demonstrated that zebra finch embryos express lower levels of Gli3, one of the key transcriptional effectors of the HH pathway. Importantly, GLI3 acts largely as a transcriptional repressor in the developing neural tube, so that reduced repressor levels in the zebra finch neural tube may poise this tissue to activate HH targets faster,
and in response to lower levels of ligand, than in the chicken neural tube, which possesses significantly higher levels of Gli3 expression. Why do neuroepithelial cells in the zebra finch neural tube need to be more sensitive to HH signaling than their chicken counterparts? Perhaps one explanation is that zebra finch embryos develop on a significantly faster timescale (14–16 days) than chicken embryos (20–21 days). Thus, the accelerated patterning observed in the zebra finch embryo may be dictated by this shortened gestation length. An intriguing experiment would be to compare neural patterning in avian species of similar size with different gestation periods. Alternatively, comparison of spinal cord development in different chicken breeds with similar gestation lengths but different sizes (e.g., bantam chickens) may yield significant insight. Further, as noted by the authors, SHH ligands do not operate in a vacuum; instead, complex interplays with other key signaling pathways (e.g., Wnt, BMP, Notch, FGF, RA, etc.) all combine to properly pattern the spinal cord (reviewed in Gouti et al., 2015). Thus, it will be essential to examine how the contributions of these key pathways may also be altered across different species to achieve proper tissue scaling. In a similar vein, establishing whether interactions between pathways classically associated with patterning (e.g., SHH) and those associated with growth control (e.g., Hippo) also contribute to scaling will be important future directions. In fact, recent work suggests that, in the Drosophila wing disc, Hh
signaling regulates wing size through JNK-mediated inhibition of the Hippo pathway (Willsey et al., 2016). Overall, the concept of scaling remains one of the most intriguing fundamental questions in all of biology; the work of Uygur et al. (2016) highlights the value of combining innovative comparative biology approaches with classic developmental biology techniques and modern molecular tools to provide significant insight into this process during neural patterning. Similarly integrated and innovative approaches will be required to address scaling in other tissues across different species. REFERENCES Gouti, M., Metzis, V., and Briscoe, J. (2015). Trends Genet. 31, 282–289. Inomata, H., Shibata, T., Haraguchi, T., and Sasai, Y. (2013). Cell 153, 1296–1311. Kicheva, A., Bollenbach, T., Ribeiro, A., Valle, H.P., Lovell-Badge, R., Episkopou, V., and Briscoe, J. (2014). Science 345, 1254927. Oosterveen, T., Kurdija, S., Alekseenko, Z., Uhde, C.W., Bergsland, M., Sandberg, M., Andersson, E., Dias, J.M., Muhr, J., and Ericson, J. (2012). Dev. Cell 23, 1006–1019. Peterson, K.A., Nishi, Y., Ma, W., Vedenko, A., Shokri, L., Zhang, X., McFarlane, M., Baizabal, J.M., Junker, J.P., van Oudenaarden, A., et al. (2012). Genes Dev. 26, 2802–2816. Uygur, A., Young, J., Huycke, T.R., Koska, M., Briscoe, J., and Tabin, C.J. (2016). Dev. Cell 37, this issue, 127–135. Willsey, H.R., Zheng, X., Carlos Pastor-Pareja, J., Willsey, A.J., Beachy, P.A., and Xu, T. (2016). eLife 5, http://dx.doi.org/10.7554/eLife.11491. Wu, H., Manu, Jiao, R., and Ma, J. (2015). Nat. Commun. 6, 10031.
Developmental Cell 37, April 18, 2016 109
Previews F-actin in the zippering cells (Hashimoto et al., 2015). Ogura and Sasakura (2016) show this to be blocked when epidermal cell-cycle timing is manipulated, but the causal link between cell-cycle timing and F-actin accumulation remains unclear. One intriguing possibility is that the epidermis may be actively pushing dorsomedially to bring the neural folds together and that control of the epidermal cell cycle is essential for maintaining the balance of mechanical forces in the ectoderm. The small, simple Ciona embryo will be a powerful model to investigate the molecular details of how S-phase length is spatially patterned and the mechanistic details of how patterns of cell division in
the epidermis influence neural tube closure. The ability to easily image the full anterior-to-posterior length of a neurulating chordate embryo with subcellular detail made it more feasible to first detect this unusual cell-cycle compensation phenomenon in Ciona, but it will be interesting and important to determine whether similar mechanisms are used in other taxa.
Duncan, T., and Su, T.T. (2004). Curr. Biol. 14, R305–R307. Edgar, B.A., Lehman, D.A., and O’Farrell, P.H. (1994). Development 120, 3131–3143. Farrell, J.A., Shermoen, A.W., Yuan, K., and O’Farrell, P.H. (2012). Genes Dev. 26, 714–725. Hashimoto, H., Robin, F.B., Sherrard, K.M., and Munro, E.M. (2015). Dev. Cell 32, 241–255. Ogura, Y., and Sasakura, Y. (2016). Dev. Cell 37, this issue, 148–161.
REFERENCES
Ogura, Y., Sakaue-Sawano, A., Nakagawa, M., Satoh, N., Miyawaki, A., and Sasakura, Y. (2011). Development 138, 577–587.
Bouldin, C.M., and Kimelman, D. (2014). Cell Cycle 13, 2165–2171.
Stolfi, A., and Christiaen, L. (2012). Genetics 192, 55–66.
Dumollard, R., Hebras, C., Besnardeau, L., and McDougall, A. (2013). Dev. Biol. 384, 331–342.
Veeman, M., and Reeves, W. (2015). Genesis 53, 143–159.
Spinal Cords Built to Scale Benjamin L. Allen1,* 1Department of Cell and Developmental Biology, University of Michigan, Ann Arbor, MI 48109, USA *Correspondence: [email protected] http://dx.doi.org/10.1016/j.devcel.2016.04.007
An outstanding question in embryogenesis is how different-sized animals maintain similarly proportioned body plans. In this issue of Developmental Cell, Uygur et al. (2016) tackle this issue in the context of neural tube patterning, discovering that differential sensitivity to the Hedgehog pathway may provide one answer to this fundamental issue. One of the fundamental questions in developmental biology is: what regulates size? Cell size, organ size, and embryo size can all be dynamically regulated both within and across species; however, the mechanisms that control these processes remain largely unexplored, particularly in the context of the whole embryo, where the maintenance of similarly proportioned body plans in the face of significant differences in organism size is referred to as scaling. Recent studies have addressed scaling in the context of early, BMP-dependent dorsoventral patterning in the developing Xenopus embryo (Inomata et al., 2013), as well as via the Bicoid morphogen gradient in Drosophila embryos (Wu et al., 2015). This question has also been addressed during dorsoventral patterning of the vertebrate neural tube, where recent work suggested that altered differentia108 Developmental Cell 37, April 18, 2016
tion in neural cell progenitors is essential for regulating scaling in this tissue (Kicheva et al., 2014). Despite these studies, the fundamental mechanisms that control scaling in related organisms of different sizes remain unclear. In this issue of Developmental Cell, Uygur et al. (2016) investigate the role of the Hedgehog signaling pathway in scaling of the neural tube across different avian species, specifically comparing spinal cord patterning in zebra finch and chicken embryos, two species of significantly different embryonic and adult size. The authors initially analyzed the expression of three transcription factors (NKX2.2, OLIG2, and NKX6.1) that play essential roles in mediating cell fate specification in the neural tube and are themselves direct transcriptional targets of the HH pathway (Oosterveen et al., 2012; Peterson et al., 2012). Strikingly, patterning occurs faster
and across a smaller tissue field in the zebra finch embryo than in the chicken embryo. One might expect that the faster patterning in the zebra finch embryos could be due to relatively higher levels of HH ligand production; however, Sonic HH (SHH) levels are actually lower in the zebra finch neural tube compared to the chicken neural tube. These data suggested that zebra finch neuroepithelial cells are more sensitive to SHH levels than their chicken counterparts. To test this hypothesis, the authors obtained transgenic lines of both species that ubiquitously express GFP. From these animals, the authors created chimeric neural tubes, transplanting GFP-positive donor tissue into GFP-negative host tissue within neural tubes of embryos from the opposite species. Consistent with their hypothesis, the zebra finch neuroepithelial cells are more sensitive to endogenous SHH ligand than
Developmental Cell
Previews the corresponding chicken neuroepithelia. To confirm and extend these data, the authors also utilized naive intermediate neural explants isolated from both zebra finch and chicken embryos in combination with recombinant SHH ligand to demonstrate that the zebra finch neuroepithelium activates higher-level HH pathway targets (e.g., NKX2.2) in response to lower levels of HH ligand. In a final set of experiments, in ovo electroporations were performed with a HH-responsive GFP reporter to demonstrate a greater range of HH pathway activity across the dorsoventral axis in the zebra finch neural tube compared to the chicken neural tube. The use of careful comparative biology combined with classic developmental biology approaches thus identifies a novel role for morphogen signaling in scaling of the embryonic neural tube. So, while differential SHH sensitivity appears to play a role in scaling, an important question remains: what regulates the differential responsiveness to SHH in the zebra finch neural tube? The authors initially tackled this question using a pharmacologic tool, SAG, a smoothened agonist that activates HH signaling downstream of ligand. Surprisingly, the data suggested that the altered HH response is downstream of Smoothened; in fact, subsequent experiments demonstrated that zebra finch embryos express lower levels of Gli3, one of the key transcriptional effectors of the HH pathway. Importantly, GLI3 acts largely as a transcriptional repressor in the developing neural tube, so that reduced repressor levels in the zebra finch neural tube may poise this tissue to activate HH targets faster,
and in response to lower levels of ligand, than in the chicken neural tube, which possesses significantly higher levels of Gli3 expression. Why do neuroepithelial cells in the zebra finch neural tube need to be more sensitive to HH signaling than their chicken counterparts? Perhaps one explanation is that zebra finch embryos develop on a significantly faster timescale (14–16 days) than chicken embryos (20–21 days). Thus, the accelerated patterning observed in the zebra finch embryo may be dictated by this shortened gestation length. An intriguing experiment would be to compare neural patterning in avian species of similar size with different gestation periods. Alternatively, comparison of spinal cord development in different chicken breeds with similar gestation lengths but different sizes (e.g., bantam chickens) may yield significant insight. Further, as noted by the authors, SHH ligands do not operate in a vacuum; instead, complex interplays with other key signaling pathways (e.g., Wnt, BMP, Notch, FGF, RA, etc.) all combine to properly pattern the spinal cord (reviewed in Gouti et al., 2015). Thus, it will be essential to examine how the contributions of these key pathways may also be altered across different species to achieve proper tissue scaling. In a similar vein, establishing whether interactions between pathways classically associated with patterning (e.g., SHH) and those associated with growth control (e.g., Hippo) also contribute to scaling will be important future directions. In fact, recent work suggests that, in the Drosophila wing disc, Hh
signaling regulates wing size through JNK-mediated inhibition of the Hippo pathway (Willsey et al., 2016). Overall, the concept of scaling remains one of the most intriguing fundamental questions in all of biology; the work of Uygur et al. (2016) highlights the value of combining innovative comparative biology approaches with classic developmental biology techniques and modern molecular tools to provide significant insight into this process during neural patterning. Similarly integrated and innovative approaches will be required to address scaling in other tissues across different species. REFERENCES Gouti, M., Metzis, V., and Briscoe, J. (2015). Trends Genet. 31, 282–289. Inomata, H., Shibata, T., Haraguchi, T., and Sasai, Y. (2013). Cell 153, 1296–1311. Kicheva, A., Bollenbach, T., Ribeiro, A., Valle, H.P., Lovell-Badge, R., Episkopou, V., and Briscoe, J. (2014). Science 345, 1254927. Oosterveen, T., Kurdija, S., Alekseenko, Z., Uhde, C.W., Bergsland, M., Sandberg, M., Andersson, E., Dias, J.M., Muhr, J., and Ericson, J. (2012). Dev. Cell 23, 1006–1019. Peterson, K.A., Nishi, Y., Ma, W., Vedenko, A., Shokri, L., Zhang, X., McFarlane, M., Baizabal, J.M., Junker, J.P., van Oudenaarden, A., et al. (2012). Genes Dev. 26, 2802–2816. Uygur, A., Young, J., Huycke, T.R., Koska, M., Briscoe, J., and Tabin, C.J. (2016). Dev. Cell 37, this issue, 127–135. Willsey, H.R., Zheng, X., Carlos Pastor-Pareja, J., Willsey, A.J., Beachy, P.A., and Xu, T. (2016). eLife 5, http://dx.doi.org/10.7554/eLife.11491. Wu, H., Manu, Jiao, R., and Ma, J. (2015). Nat. Commun. 6, 10031.
Developmental Cell 37, April 18, 2016 109