The pericardium of Oikopleura dioica (Tunicata, Appendicularia) contains two distinct cell types and is rotated by 90 degrees to the left

The planktonic Oikopleura dioica belongs to Tunicata, the probable sister taxon to Craniota, and might show plesiomorphic characters, conserved from the common lineage of Tunicata and Craniota. In O. dioica a pericardium in a position similar to other chordates but also to the heart and pericardium of craniates is found. Surprisingly, little is known about the ultrastructure of the pericardium in O. dioica. Here, we show based on electron microscopy that the pericardium is completely lined by a single layer of 16 epithelial cells: 6 epithelial myocardial cells on the left side of the pericardium and 10 peritoneal cells constituting the right side. One of the peritoneal cells, situated at the ventral border between peritoneal cells and myocardial cells has an extension that anchors the pericardium to the basal lamina beneath the latero-ventral epidermis. The primary body cavity of O. dioica appears quite uniformly clear in electron microscopic aspect but several sheets, resembling the basal lamina of the pericardium cross the larger spaces of the body cavity and connect to the pericardial basal lamina. This is the first detailed description of two distinct cell types in the epithelial lining of the pericardium of O. dioica. In comparison with other chordates, we conclude that two cell types can be reconstructed for the last common ancestor of Chordata at least. The position of the pericardium at the intersection of trunk and tail in combination with the basal-lamina like sheets spanning the hemocoel is probably of importance for the function of the circulation of the hemocoelic fluid. Similar to the tail, the axis of the pericardium is shifted through 90 degrees to the left as compared to the main body axis of the trunk and we infer that this shift is an apomorphic character of Appendicularia.

Massive Gene Loss and Function Shuffling in Appendicularians Stretch the Boundaries of Chordate Wnt Family Evolution

Gene loss is a pervasive source of genetic variation that influences species evolvability, biodiversity and the innovation of evolutionary adaptations. To better understand the evolutionary patterns and impact of gene loss, here we investigate as a case study the evolution of the wingless (Wnt) family in the appendicularian tunicate Oikopleura dioica, an emergent EvoDevo model characterized by its proneness to lose genes among chordates. Genome survey and phylogenetic analyses reveal that only four of the thirteen Wnt subfamilies have survived in O. dioica—Wnt5, Wnt10, Wnt11, and Wnt16,—representing the minimal Wnt repertoire described in chordates. While the loss of Wnt4 and Wnt8 likely occurred in the last common ancestor of tunicates, representing therefore a synapomorphy of this subphylum, the rest of losses occurred during the evolution of appendicularians. This work provides the first complete Wnt developmental expression atlas in a tunicate and the first insights into the evolution of Wnt developmental functions in appendicularians. Our work highlights three main evolutionary patterns of gene loss: (1) conservation of ancestral Wnt expression domains not affected by gene losses; (2) function shuffling among Wnt paralogs accompanied by gene losses; and (3) extinction of Wnt expression in certain embryonic directly correlated with gene losses. Overall our work reveals that in contrast to “conservative” pattern of evolution of cephalochordates and vertebrates, O. dioica shows an even more radical “liberal” evolutionary pattern than that described ascidian tunicates, stretching the boundaries of the malleability of Wnt family evolution in chordates.

Telomere-to-telomere assembly of the genome of an individual Oikopleura dioica from Okinawa using Nanopore-based sequencing

Background The larvacean Oikopleura dioica is an abundant tunicate plankton with the smallest (65–70 Mbp) non-parasitic, non-extremophile animal genome identified to date. Currently, there are two genomes available for the Bergen (OdB3) and Osaka (OSKA2016) O. dioica laboratory strains. Both assemblies have full genome coverage and high sequence accuracy. However, a chromosome-scale assembly has not yet been achieved. Results Here, we present a chromosome-scale genome assembly (OKI2018_I69) of the Okinawan O. dioica produced using long-read Nanopore and short-read Illumina sequencing data from a single male, combined with Hi-C chromosomal conformation capture data for scaffolding. The OKI2018_I69 assembly has a total length of 64.3 Mbp distributed among 19 scaffolds. 99% of the assembly is contained within five megabase-scale scaffolds. We found telomeres on both ends of the two largest scaffolds, which represent assemblies of two fully contiguous autosomal chromosomes. Each of the other three large scaffolds have telomeres at one end only and we propose that they correspond to sex chromosomes split into a pseudo-autosomal region and X-specific or Y-specific regions. Indeed, these five scaffolds mostly correspond to equivalent linkage groups in OdB3, suggesting overall agreement in chromosomal organization between the two populations. At a more detailed level, the OKI2018_I69 assembly possesses similar genomic features in gene content and repetitive elements reported for OdB3. The Hi-C map suggests few reciprocal interactions between chromosome arms. At the sequence level, multiple genomic features such as GC content and repetitive elements are distributed differently along the short and long arms of the same chromosome. Conclusions We show that a hybrid approach of integrating multiple sequencing technologies with chromosome conformation information results in an accurate de novo chromosome-scale assembly of O. dioica’s highly polymorphic genome. This genome assembly opens up the possibility of cross-genome comparison between O. dioica populations, as well as of studies of chromosomal evolution in this lineage.

3D reconstruction of structures of hatched larva and young juvenile of the larvacean Oikopleura dioica using SBF-SEM

The larvacean Oikopleura dioica is a planktonic chordate and an emerging model organism with a short life cycle of 5 days that belongs toTunicata (Urochordata), the sister clade of vertebrates. It is characterized by the rapid development of a tadpole-shaped body. Organ formation in the trunk proceeds within 7 h after the hatching of the tailbud larvae at 3 h after fertilization (hpf) and is completed at 10 hpf, giving rise to fully functional juveniles as miniature adult form. Serial block face scanning electron microscopy was used to acquire ~ 2000 serial transverse section images of a 3 hpf larva and a 10 hpf juvenile to characterize the structures and cellular composition of the trunk and organs using 3D images and movies. Germ cells were found to fuse and establish a central syncytial cell in the gonad as early as 10 hpf. Larval development gave rise to functional organs after several rounds of cell division through trunk morphogenesis. The feature would make O. dioica ideal for analyzing cellular behaviors during morphogenetic processes using live imaging. The detailed descriptions of the larvae and juveniles provided in this study can be utilized as the start and end points of organ morphogenesis in this rapidly developing organism.