Annelid functional genomics reveal the origins of bilaterian life cycles

Abstract

Indirect development with an intermediate larva exists in all major animal lineages, and thus larvae are central to most scenarios for animal evolution. Yet how larvae evolved remains disputed. Here we show that changes in the timing of trunk formation underpin the diversification of larvae and bilaterian life cycles. Combining chromosome-scale genome sequencing with transcriptomic and epigenomic profiling in the slow-evolving oweniid Owenia fusiformis, we found that different genes and genomic regulatory elements control the development of its feeding larva and adult stage. First, O. fusiformis embryos develop into an enlarged anterior domain that forms larval tissues and the adult head, as posterior growth and trunk patterning is deferred to pre-metamorphic stages. These traits also occur in the so-called "head larvae" of other bilaterians, with whom O. fusiformis larva shows extensive transcriptomic similarities. Conversely, animals with non-feeding larvae and gradual metamorphoses, such as the annelid Capitella teleta, start trunk differentiation during embryogenesis, like direct developers. Together, our findings suggest that the ancestral temporal decoupling of head and trunk formation, as retained in extant "head larvae", allowed larval evolution in Bilateria, questioning prevailing scenarios that propose either co-option or innovation of gene regulatory programmes to explain larva and adult origins.

ERK1/2 is an ancestral organising signal in spiral cleavage

“…4d; Supplementary Table 6), including a variety of transcription factors that are markers of particular cells and tissue types (e.g., six3/6, gsc, cdx, AP2, foxQ2 ), required for mesoderm development (e.g., twist, hand2, foxH ) and neurogenesis (e.g., POU4, irxA ), Wnt ligands ( wnt1, wntA and wnt4) , TGF-β modulators ( noggin and BAMBI ), and Notch signalling components ( delta and notch-like ). Stage-specific RNA-seq data covering twelve developmental time-points 33 , from the unfertilized oocyte to the mature larva, confirmed that the expression of all candidate genes upregulates at the time of or just after 4d specification and di-P-ERK1/2 enrichment in this cell (Fig. 4d).…”

“…For GO mapping, the GO terms of differentially expressed genes associated with each comparison were extracted from O. fusiformis reference genome annotation 33 (available at the repository https://github.com/ChemaMD/OweniaGenome). GO enrichment analyses were implemented using the GOseq R package, correcting for gene length bias 62 , and analysing up-regulated and down-regulated genes independently.…”

“…To identify transcription factor motifs in chromatin accessible regions around the cdx and delta loci, we use two already available replicated Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) libraries 33 generated from >50,000 cells from 850-900 coeloblastula at 5 hpf. Briefly, cells were dissociated and lysed in ice cold ATAC lysis buffer (10 mM Tris-HCl pH 7.5, 10 mM NaCl, 3 mM MgCl 2 , 0.1% (v/v) IGEPAL, 0.1% (v/v) Tween-20, 0.01% (v/v) Digitonin) with the gentle use of a pestle and incubated on ice for 3 min.…”

“…After signal development with a Tyramide Signal Amplification kit (Akoya Biosciences, #NEL742001KT), these samples were washed several times in PTx and treated for secondary antibody incubation as described above. To characterise in silico gene expression dynamics during development, we used an available stage specific RNA-seq dataset 33 , representing the average expression of the two replicates with the package pheatmap v.1.0.12 available in R, were colour intensity shows the z-score value for each gene.…”

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

“…To investigate the expression dynamics of the Fox genes in O. fusiformis and relate each of these genes to major morphogenetic events during the life cycle of this annelid, we used available stage-specific RNA-seq data covering 14 developmental time points, from the unfertilized oocyte to the juvenile stage (Liang et al 2022). In O. fusiformis , the temporal expression dynamics of the Fox genes seem to correlate with their assignment to clade I and clade II, because most clade II Fox genes are expressed maternally and during the early cleavage stages (i.e., up to the 8-cell stage), while clade I Fox genes tend to show short peaks of expression at single developmental stages, from the 32-cell stage onwards (Figure 3A).…”