The brittle star genome illuminates the genetic basis of animal appendage regeneration

Elise Parey, Olga Ortega-Martinez, Jérôme Delroisse, Laura Piovani, Anna Czarkwiani, David Dylus, Srishti Arya, Samuel Dupont, Michael Thorndyke, Tomas Larsson, Kerstin Johannesson, Katherine M. Buckley, Pedro Martinez, Paola Oliveri, Ferdinand Marlétaz

Posted on: 16 December 2023

Preprint posted on 8 November 2023

Fortifying our understanding of regeneration: @Ferdix, Elise Parey, and collaborators use the brittle star genome to reveal conserved gene expression dynamics across regenerating model systems

Selected by Isabella Cisneros


When you think of a starfish, the brittle star Amphiura filiformis is likely not the first echinoderm to come to mind. Composed of serpent-like arms and a disk-like center, brittle stars belong to a class called Ophiuroidea, sister to sea stars and one of five overall classes within the echinoderm phylum [1]. While its relative Patiria miniata, colloquially known as the bat star, is more frequently used as the representative for echinoderms in various types of studies, A. filiformis has recently been used for studies aiming to understand arm and skeletal regeneration as well as ecological studies [2, 3].

Despite its experimental potential, however, A. filiformis and Ophiuroidea at large have not been well represented in the development of genomic resources for echinoderms, which has so far favored representatives from the Asteroidea (sea stars), Holothuroidea (sea cucumbers), and Echinoidea (sea urchin) classes. Previous work has established genomes for other brittle stars, such as Ophioderma brevispinum, Ophionereis fasciata, and Ophiothrix spiculata [4, 5, 6]. However, none exists for A. filiformis, one of the main models currently used in regenerative biology studies. In order to address this gap, the authors sequenced and assembled the A. filiformis genome and used it to answer questions regarding echinoderm evolutionary history as well as conservation of regeneration dynamics across organisms outside of the echinoderms. In doing so, the authors establish an important tool for brittle star researchers and set the stage for future comparative evolutionary work.


Main Findings:

Brittle star genome reveals both broad inter-chromosomal rearrangements and smaller scale rearrangements in the Hox and Parahox clusters

The authors began by assembling the A. filiformis genome using high-coverage long nanopore reads, which they subsequently annotated and used to build curated lists for genes associated with major functions and signaling pathways. After doing so, they used previously developed chromosome-scale genomes for sea stars, sea cucumbers, and sea urchins to investigate chromosomal conservation and fusion. Synteny comparisons using these genomes revealed that the brittle star genome is significantly rearranged, with only three chromosomes having a one-to-one relationship with spiny sea star chromosomes. The authors also reconstructed linkage groups present in the ancestor to all echinoderms in order to assess derived and ancestral chromosomal arrangements. Their reconstruction predicted the presence of 23 ELGs (Elutherozoa linkage groups). Using these ELGs, they found that the brittle star genome had undergone 26 inter-chromosomal rearrangements, making it the most reshuffled echinoderm genome surveyed thus far. The authors also investigated the structure of the Hox and Parahox clusters in A. filiformis and found that they too exhibit genomic rearrangements. Interestingly, these rearrangements appear to parallel ones characterized in the sea urchin genome as well.

Gene duplications underlie brittle star larval skeleton and bioluminescence as well as regeneration-related gene family expansions

The authors next decided to look at two gene families known to have undergone gene duplications in echinoderms: phb/pmar1, which is connected to the development of an elaborate larval skeleton, and luciferase genes, which are connected to the brittle star’s bioluminescent abilities. They identified a total of 13 phb paralogs, which are themselves distant homologs of sea urchin pmar1, in A. filiformis, which they believe supported the development of larval skeletons in brittle stars. They also found nine luciferase-like gene copies in the brittle star and identified similar duplications in all echinoderm lineages except for the sea star, confirming previous claims that echinoderms possessed multiple copies of luciferase.

Following this analysis, the authors broadened their scope and further analyzed gene families that had undergone significant expansions or contractions. In the brittle star, these gene families were linked to immune-related processes, regeneration, and keratan sulfate metabolism. Diving deeper into the gene families associated with regeneration, the authors found that these were indeed expressed during A. filiformis arm regeneration. Additionally, they found that four of these gene families have roles related to coagulation and/or clotting in vertebrates; accordingly, duplications of these families may have contributed to the rapid and efficient wound closure necessary for regeneration in brittle stars.

Gene expression and gene dynamics are conserved across the regenerative models Ambystoma mexicanum and Parhyale hawaiensis

The authors decided to further probe transcriptional programs underlying arm regeneration in the brittle star by profiling gene expression during seven different stages of regeneration following amputation. They classified genes into nine major temporal clusters and, through functional enrichment analysis, identified three phases of arm regeneration (wound healing, proliferation, and tissue differentiation) they believed to be consistent with timelines in other animal systems. Additionally, genomic phylostratigraphy analysis revealed that the majority of brittle star regeneration is driven by ancient genes dating back to metazoan ancestors.

Given these two findings, the authors decided to use datasets from the regenerative models Ambystoma mexicanumand Parhyale hawaiensis to investigate temporal deployment of genes in a comparative framework. While the Parhyale dataset already had defined clusters, the authors had to define nine co-expression clusters recapitulating the three phases of regeneration in the axolotl dataset. They found that four of these clusters were shared between the three species. Additionally, the co-expression clusters that were most broadly conserved across the species primarily consisted of genes expressed during the proliferative stage of regeneration. Interestingly, these co-expression clusters were deployed within similar temporal sequences, with results showing that this order is more conserved within regeneration and developmental datasets across species than within them.


Why I Chose This Preprint:

I was really excited by this study for multiple reasons! First, I feel that the authors’ efforts to generate the brittle star genome fills an important phylogenetic gap that can help contextualize both echinoderm-specific trends as well as broader evolutionary questions. Additionally, I particularly enjoyed their comparative approach and their use of the axolotl and Parhyale regeneration datasets; I believe approaches like these will be very informative for comparisons of processes like regeneration, especially across model systems. This preprint will be very informative for both brittle star researchers and those interested in the evo-devo underlying regeneration, and I cannot wait to see how this research continues to develop.


Questions For The Authors:

  1. Given that this study contains an impressive amount of bioinformatics analysis, how are you planning on validating these results experimentally? Do you have any pertinent future directions in mind?
  2. While the amount of limb regeneration related datasets may be limited, do you have plans for expanding your comparisons to other regenerative models, perhaps outside of the context of regeneration?



[1] Cannon JT, Kocot KM, Waits DS, Weese DA, Swalla BJ, Santos SR, Halanych KM. 2014. Phylogenomic resolution of the hemichordate and echinoderm clade. Curr. Biol. [Internet] 24:2827–2832. Available from: doi:10.1016/j.cub.2014.10.016

[2] Czarkwiani  A, Ferrario  C, Dylus  DV, Sugni  M, Oliveri  P. 2016. Skeletal regeneration in the brittle star Amphiura filiformis. Front. Zool. [Internet] 13:18. Available from: doi:10.1186/s12983-016-0149-x

[3] Dupont  S, Thorndyke  MC. 2006. Growth or differentiation? Adaptive regeneration in the brittlestar Amphiura filiformis. J. Exp. Biol. [Internet] 209:3873–3881. Available from: doi:10.1242/jeb.02445

[4] Kudtarkar P, Cameron RA. Echinobase: an expanding resource for echinoderm genomic information. Database. 2017.

[5] Long KA, Nossa CW, Sewell MA, Putnam NH, Ryan JF. Low coverage sequencing of three echinoderm genomes: the brittle star Ophionereis fasciata, the sea star Patiriella regularis, and the sea cucumber Australostichopus mollis. GigaScience. 2016; 5(1):13742–016.

[6] Mashanov V, Akiona J, Khoury M, Ferrier J, Reid R, Machado DJ, Zueva O, Janies D. Active Notch signaling is required for arm regeneration in a brittle star. PLoS ONE. 2020; 15(5):0232981.

Tags: axolotl, brittle star, echinoderms, genomics, parhyale, regeneration


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