ERK1/2 is an ancestral organising signal in spiral cleavage

Abstract

Embryonic organisers are signalling centres that instruct the establishment of body plans during animal embryogenesis, thus underpinning animal morphological diversity. In spiral cleavage - a stereotypic developmental programme ancestral to 14, nearly half, of the animal phyla (e.g., molluscs, annelids and flatworms), a cell known as the D-quadrant organiser defines cell fates and the body axes. ERK1/2 specifies the embryonic organiser in molluscs, yet how this signalling cascade exerts organising activity and whether this role is conserved in other spiral cleaving groups is unclear. Here, we demonstrate that ERK1/2 promotes the specification and inductive activity of the D-quadrant organiser in Owenia fusiformis, an early-branching annelid exhibiting ancestral developmental traits. In this species, active di-phosphorylated ERK1/2 mediated by FGF receptor activity localises to the 4d micromere, establishing the bilateral symmetry and specifying the hindgut and trunk mesodermal progenitor. Accordingly, impairing FGFR and ERK1/2 activity, as well as cell communication results in embryos developing anteroventrally radialised. Differential transcriptomic profiling shows the ParaHox cdx and the Notch ligand delta as FGFR/ERK1/2 downstream targets in 4d, further revealing that 4d specification instructs the expression of mesodermal and posterodorsal genes in neighbouring cells, putatively via the Notch pathway. The instructing role of ERK1/2 in the D-quadrant organiser is thus shared between O. fusiformis and molluscs, representing an ancestral trait of spiral cleavage. Altogether, our study begins to dissect the gene network promoting axial patterning and posterior growth in spiral cleavage, revealing extensive mechanistic diversification in body plan specification despite overall conservation of cleavage patterns in Spiralia.

Annelid functional genomics reveal the origins of bilaterian life cycles

“…This expression resembles that of molluscs 34,35 and brachiopods 28 , providing further evidence that conserved developmental mechanisms underpin larval formation in O. fusiformis [26][27][28] . However, differently from non-feeding annelid trochophores 6 , anterior ectodermal genes [26][27][28] pattern most larval tissues in O. fusiformis (Fig. 2f), whereas posterior genes are restricted to few circum-anal cells, in line with the vastly enlarged pre-oral region of the mitraria that forms early on in the adult head 21 .…”

“…Tables 2-7). Therefore, O. fusiformis genome contains a more complete gene repertoire that those reported for other annelid lineages, which together with its phylogenetic position and conserved early embryogenesis 25,26 makes it a key lineage to reconstruct the evolution of Annelida and Lophotrochozoa generally. Chromosomal linkage of orthologous genes between deuterostomes and protostomes supports the presence of 22 ancestral linkage groups in bilaterians 29,30 .…”

“…In particular, the idiosyncratic oweniid larva, commonly referred to as "mitraria" 21 , has an enlarged pre-oral region and a single tuft of posterior chaetae, as well as a pair of nephridia and a long monociliated ciliary band alike those of phylogenetically distant larvae of deuterostome lineages, such as echinoderms and hemichordates [22][23][24] . Despite these distinctive larval traits, oweniids exhibit many developmental characters considered ancestral to Annelida, and even Lophotrochozoa as a whole 25,26 , as well as similarities in larval molecular patterns with other trochophore and bilaterian larvae 24,25,27,28 . Consequently, whether the mitraria larva is the result of divergent or convergent evolution is unclear, which makes the comparative analysis of this lineage-restricted larva an excellent case study to investigate how larval traits evolve, and thereby formulate and assess scenarios on the origin of animal life cycles.…”

The Fox gene repertoire in the annelid Owenia fusiformis reveals multiple expansions of the foxQ2 class in Spiralia

“…For instance, foxJ3 , foxM1 , foxN2/3-b and foxN1/4-a and foxN1/4-b show maternal expression, which is maintained until the 8-cell stage, suggesting that they might be potential regulators of early cleavage and/or cell fate specification in this species. The genes foxG1 and foxP have a narrow peak of expression at the time of the specification of the embryonic organiser in O. fusiformis (Seudre et al 2022), and could therefore be involved in the establishment of the embryonic polarity and body plan in this annelid. Finally, some Fox genes are restricted to either the larva ( foxK2 ) or the juvenile ( foxB, foxL2 and foxQ1 ), yet their potential role at those stages remains unknown.…”

“…Riboprobes were synthesized with the T7 enzyme following manufacturer’s recommendations (Ambion’s MEGAscript kit, #AM1334) and stored in hybridization buffer at a concentration of 50 ng.μl-1 at -20°C. Single colorimetric in situ hybridization of embryos and mitraria larvae were performed following an established protocol (Martín-Durán et al 2016; Carrillo-Baltodano et al 2021; Seudre et al 2022).…”