Positional Memory in Vertebrate Regeneration: A Century's Insights from the Salamander Limb.

Salamanders, such as axolotls and newts, can regenerate complex tissues including entire limbs. But what mechanisms ensure that an amputated limb regenerates a limb, and not a tail or unpatterned tissue? An important concept in regeneration is positional memory-the notion that adult cells "remember" spatial identities assigned to them during embryogenesis (e.g., "head" or "hand") and use this information to restore the correct body parts after injury. Although positional memory is well documented at a phenomenological level, the underlying cellular and molecular bases are just beginning to be decoded. Herein, we review how major principles in positional memory were established in the salamander limb model, enabling the discovery of positional memory-encoding molecules, and advancing insights into their pattern-forming logic during regeneration. We explore findings in other amphibians, fish, reptiles, and mammals and speculate on conserved aspects of positional memory. We consider the possibility that manipulating positional memory in human cells could represent one route toward improved tissue repair or engineering of patterned tissues for therapeutic purposes.

Spatiotemporal control of cell cycle acceleration during axolotl spinal cord regeneration.

Axolotls are uniquely able to resolve spinal cord injuries, but little is known about the mechanisms underlying spinal cord regeneration. We previously found that tail amputation leads to reactivation of a developmental-like program in spinal cord ependymal cells (Rodrigo Albors et al., 2015), characterized by a high-proliferation zone emerging 4 days post-amputation (Rost et al., 2016). What underlies this spatiotemporal pattern of cell proliferation, however, remained unknown. Here, we use modeling, tightly linked to experimental data, to demonstrate that this regenerative response is consistent with a signal that recruits ependymal cells during ~85 hours after amputation within ~830 µm of the injury. We adapted Fluorescent Ubiquitination-based Cell Cycle Indicator (FUCCI) technology to axolotls (AxFUCCI) to visualize cell cycles in vivo. AxFUCCI axolotls confirmed the predicted appearance time and size of the injury-induced recruitment zone and revealed cell cycle synchrony between ependymal cells. Our modeling and imaging move us closer to understanding bona fide spinal cord regeneration.

The giant axolotl genome uncovers the evolution, scaling, and transcriptional control of complex gene loci.

Vertebrates harbor recognizably orthologous gene complements but vary 100-fold in genome size. How chromosomal organization scales with genome expansion is unclear, and how acute changes in gene regulation, as during axolotl limb regeneration, occur in the context of a vast genome has remained a riddle. Here, we describe the chromosome-scale assembly of the giant, 32 Gb axolotl genome. Hi-C contact data revealed the scaling properties of interphase and mitotic chromosome organization. Analysis of the assembly yielded understanding of the evolution of large, syntenic multigene clusters, including the Major Histocompatibility Complex (MHC) and the functional regulatory landscape of the Fibroblast Growth Factor 8 (Axfgf8) region. The axolotl serves as a primary model for studying successful regeneration.

Quiescent Neural Stem Cells for Brain Repair and Regeneration: Lessons from Model Systems.

Neural stem cells (NSCs) are multipotent progenitors that are responsible for producing all of the neurons and macroglia in the nervous system. In adult mammals, NSCs reside predominantly in a mitotically dormant, quiescent state, but they can proliferate in response to environmental inputs such as feeding or exercise. It is hoped that quiescent NSCs could be activated therapeutically to contribute towards repair in humans. This will require an understanding of quiescent NSC heterogeneities and regulation during normal physiology and following brain injury. Non-mammalian vertebrates (zebrafish and salamanders) and invertebrates (Drosophila) offer insights into brain repair and quiescence regulation that are difficult to obtain using rodent models alone. We review conceptual progress from these various models, a first step towards harnessing quiescent NSCs for therapeutic purposes.

Dorsal-Ventral Differences in Neural Stem Cell Quiescence Are Induced by p57KIP2/Dacapo.

Quiescent neural stem cells (NSCs) in the adult brain are regenerative cells that could be activated therapeutically to repair damage. It is becoming apparent that quiescent NSCs exhibit heterogeneity in their propensity for activation and in the progeny that they generate. We discovered recently that NSCs undergo quiescence in either G0 or G2 in the Drosophila brain, challenging the notion that all quiescent stem cells are G0 arrested. We found that G2-quiescent NSCs become activated prior to G0 NSCs. Here, we show that the cyclin-dependent kinase inhibitor Dacapo (Dap; ortholog of p57KIP2) determines whether NSCs enter G0 or G2 quiescence during embryogenesis. We demonstrate that the dorsal patterning factor, Muscle segment homeobox (Msh; ortholog of MSX1/2/3) binds directly to the Dap locus and induces Dap expression in dorsal NSCs, resulting in G0 arrest, while more ventral NSCs undergo G2 quiescence. Our results reveal region-specific regulation of stem cell quiescence.

A newly discovered neural stem cell population is generated by the optic lobe neuroepithelium during embryogenesis in Drosophila melanogaster.

Neural stem cells must balance symmetric and asymmetric cell divisions to generate a functioning brain of the correct size. In both the developing Drosophila visual system and mammalian cerebral cortex, symmetrically dividing neuroepithelial cells transform gradually into asymmetrically dividing progenitors that generate neurons and glia. As a result, it has been widely accepted that stem cells in these tissues switch from a symmetric, expansive phase of cell divisions to a later neurogenic phase of cell divisions. In the Drosophila optic lobe, this switch is thought to occur during larval development. However, we have found that neuroepithelial cells start to produce neuroblasts during embryonic development, demonstrating a much earlier role for neuroblasts in the developing visual system. These neuroblasts undergo neurogenic divisions, enter quiescence and are retained post-embryonically, together with neuroepithelial cells. Later in development, neuroepithelial cells undergo further cell divisions before transforming into larval neuroblasts. Our results demonstrate that the optic lobe neuroepithelium gives rise to neurons and glia over 60 h earlier than was thought previously.

The vasculature as a neural stem cell niche.

Neural stem cells (NSCs) are multipotent, self-renewing progenitors that generate progeny that differentiate into neurons and glia. NSCs in the adult mammalian brain are generally quiescent. Environmental stimuli such as learning or exercise can activate quiescent NSCs, inducing them to proliferate and produce new neurons and glia. How are these behaviours coordinated? The neurovasculature, the circulatory system of the brain, is a key component of the NSC microenvironment, or 'niche'. Instructive signals from the neurovasculature direct NSC quiescence, proliferation, self-renewal and differentiation. During ageing, a breakdown in the niche accompanies NSC dysfunction and cognitive decline. There is much interest in reversing these changes and enhancing NSC activity by targeting the neurovasculature therapeutically. Here we discuss principles of neurovasculature-NSC crosstalk, and the implications for the design of NSC-based therapies. We also consider the emerging contributions to this field of the model organism Drosophila melanogaster.

Freedom of expression: cell-type-specific gene profiling.

Cell fate and behavior are results of differential gene regulation, making techniques to profile gene expression in specific cell types highly desirable. Many methods now enable investigation at the DNA, RNA and protein level. This review introduces the most recent and popular techniques, and discusses key issues influencing the choice between these such as ease, cost and applicability of information gained. Interdisciplinary collaborations will no doubt contribute further advances, including not just in single cell type but single-cell expression profiling.