Dataset Information

I-SceI Meganuclease-mediated transgenesis in the acorn worm, Saccoglossus kowalevskii.

ABSTRACT: Hemichordates are a phylum of marine invertebrate deuterostomes that are closely related to chordates, and represent one of the most promising models to provide insights into early deuterostome evolution. The genome of the hemichordate, Saccoglossus kowalevskii, reveals an extensive set of non-coding elements conserved across all three deuterostome phyla. Functional characterization and cross-phyla comparisons of these putative regulatory elements will enable a better understanding of enhancer evolution, and subsequently how changes in gene regulation give rise to morphological innovation. Here, we describe an efficient method of transgenesis for the characterization of non-coding elements in S. kowalevskii. We first test the capacity of an I-SceI transgenesis system to drive ubiquitous or regionalized gene expression, and to label specific cell types. Finally, we identified a minimal promoter that can be used to test the capacity of putative enhancers in S. kowalevskii. This work demonstrates that this I-SceI transgenesis technique, when coupled with an understanding of chromatin accessibility, can be a powerful tool for studying how evolutionary changes in gene regulatory mechanisms contributed to the diversification of body plans in deuterostomes.

SUBMITTER: Minor PJ

PROVIDER: S-EPMC6327965 | biostudies-literature | 2019 Jan

REPOSITORIES: biostudies-literature

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I-SceI Meganuclease-mediated transgenesis in the acorn worm, Saccoglossus kowalevskii.

Minor Paul J PJ Clarke D Nathaniel DN Andrade López José M JM Fritzenwanker Jens H JH Gray Jessica J Lowe Christopher J CJ

Developmental biology 20181106 1

Hemichordates are a phylum of marine invertebrate deuterostomes that are closely related to chordates, and represent one of the most promising models to provide insights into early deuterostome evolution. The genome of the hemichordate, Saccoglossus kowalevskii, reveals an extensive set of non-coding elements conserved across all three deuterostome phyla. Functional characterization and cross-phyla comparisons of these putative regulatory elements will enable a better understanding of enhancer e ...[more]

PMID: 30412702

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Project description:Hemichordates are an important group for investigating the evolution of bilaterian nervous systems. As the closest chordate outgroup with a bilaterally symmetric adult body plan, hemichordates are particularly informative for exploring the origins of chordates. Despite the importance of hemichordate neuroanatomy for testing hypotheses on deuterostome and chordate evolution, adult hemichordate nervous systems have not been comprehensively described using molecular techniques, and classic histological descriptions disagree on basic aspects of nervous system organization. A molecular description of hemichordate nervous system organization is important for both anatomical comparisons across phyla and for attempts to understand how conserved gene regulatory programs for ectodermal patterning relate to morphological evolution in deep time. Here, we describe the basic organization of the adult hemichordate Saccoglossus kowalevskii nervous system using immunofluorescence, in situ hybridization, and transgenic reporters to visualize neurons, neuropil, and key neuronal cell types. Consistent with previous descriptions, we found the S. kowalevskii nervous system consists of a pervasive nerve plexus concentrated in the anterior, along with nerve cords on both the dorsal and ventral side. Neuronal cell types exhibited clear anteroposterior and dorsoventral regionalization in multiple areas of the body. We observed spatially demarcated expression patterns for many genes involved in synthesis or transport of neurotransmitters and neuropeptides but did not observe clear distinctions between putatively centralized and decentralized portions of the nervous system. The plexus shows regionalized structure and is consistent with the proboscis base as a major site for information processing rather than the dorsal nerve cord. In the trunk, there is a clear division of cell types between the dorsal and ventral cords, suggesting differences in function. The absence of neural processes crossing the basement membrane into muscle and extensive axonal varicosities suggest that volume transmission may play an important role in neural function. These data now facilitate more informed neural comparisons between hemichordates and other groups, contributing to broader debates on the origins and evolution of bilaterian nervous systems.

Project description:IntroductionTraditionally, the origin of the third germ layer and its special formation of coelomic cavities by enterocoely is regarded to be an informative character in phylogenetic analyses. In early deuterostomes such as sea urchins, the mesoderm forms through a single evagination pinching off from the apical end of the archenteron which then gives off mesocoela and metacoela on each side. This echinoid-type coelom formation has conventionally been assumed to be ancestral for Deuterostomia. However, recent phylogenetic analyses show that Echinodermata hold a more derived position within Deuterostomia. In this regard a subgroup of Hemichordata, namely enteropneusts, seem to host promising candidates, because they are supposed to have retained many ancestral deuterostome features on the one hand, and furthermore share some characteristics with chordates on the other hand. In enteropneusts a wide range of different modes of coelom formation has been reported and in many cases authors of the original observations carefully detailed the limitations of their descriptions, while these doubts disappeared in subsequent reviews. In the present study, we investigated the development of all tissues in an enteropneust, Saccoglossus kowalevskii by using modern morphological techniques such as complete serial sectioning for LM and TEM, and 3D-reconstructions, in order to contribute new data to the elucidation of deuterostome evolution.ResultsOur data show that in the enteropneust S. kowalevskii all main coelomic cavities (single protocoel, paired mesocoela and metacoela) derive from the endoderm via enterocoely as separate evaginations, in contrast to the aforementioned echinoid-type. The anlagen of the first pair of gill slits emerge at the late kink stage (~96 h pf). From that time onwards, we documented a temporal left-first development of the gill slits and skeletal gill rods in S. kowalevskii until the 2 gill slit juvenile stage.ConclusionsThe condition of coelom formation from separate evaginations is recapitulated in the larva of amphioxus and can be observed in crinoid echinoderms in a similar way. Therefore, coelom formation from separated pouches, rather than from a single apical pouch with eventual subdivision is suggested as the ancestral type of coelom formation for Deuterostomia. Left-right asymmetries are also present in echinoderms (rudiment formation), cephalochordates (larval development), tunicates (gut coiling) and vertebrates (visceral organs), and it is known from other studies applying molecular genetic analyses that genes such as nodal, lefty and pitx are involved during development. We discuss our findings in S. kowalevskii in the light of morphological as well as molecular genetic data.

Dataset Information

I-SceI Meganuclease-mediated transgenesis in the acorn worm, Saccoglossus kowalevskii.

Publications

I-SceI Meganuclease-mediated transgenesis in the acorn worm, Saccoglossus kowalevskii.

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