A novel role for ATP2B in ascidians: Ascidian-specific mutations in ATP2B contribute to sperm chemotaxis.

Sperm chemotaxis, in which sperms are attracted to conspecific eggs via species-specific attractants, plays an important role in fertilization. This phenomenon has been observed in various animals and species-specific sperm attractants have been reported in some species. However, the mechanisms involved in the reception and recognition of the species-specific attractant by the sperms is poorly studied. Previously, we found that the plasma membrane-type Ca2+ /ATPase (PMCA) is the receptor for the sperm-activating and -attracting factor (SAAF) in the ascidian Ciona intestinalis. To determine the role of PMCA in species-specific sperm chemotaxis, we identified the amino acid sequences of PMCAs derived from six Phlebobranchia species. The testis-specific splice variant of PMCA was found to be present in all the species investigated and the ascidian-specific sequence was detected near the 3'-terminus. Moreover, dN/dS analysis revealed that the extracellular loops 1, 2, and 4 in ascidian PMCA underwent a positive selection. These findings suggest that PMCA recognizes the species-specific structure of SAAF at the extracellular loops 1, 2, and 4, and its testis-specific C-terminal region is involved in the activation and chemotaxis of ascidian sperms.

Kulikovia alborostrata and Kulikovia fulva comb. nov. (Nemertea: Heteronemertea) are Sister Species with Prezygotic Isolating Barriers.

The heteronemertean Kulikovia alborostrata (Takakura, 1898) was originally described as Lineus alborostratus based on material from Misaki, Japan. Although this species was regarded as consisting of two color variants, purple and brown-yellow, the identity of these variants has never been examined based on topotypes. In this study, we performed a multi-locus phylogeny reconstruction, species delimitation analyses, and cross-fertilization experiments to examine the species status of Takakura's original taxon concept consisting of these color variants. Our results suggest that the purple type is identical to Lineus alborostratus Takakura, 1898 auct. (currently Kulikovia alborostrata), whereas the brown-yellow type is conspecific with Lineus fulvus Iwata, 1954, originally established from Hokkaido. These two species appear to have a sister-taxon relationship and are reproductively isolated from each other by prezygotic mechanisms involving gamete incompatibility, minimally separated with 2.8% (16S rRNA) and 14.4% (COI) uncorrected p-distances. We propose that the purple type be considered as representing the true identity of the nominal species Lineus alborostratus (currently assigned to the genus Kulikovia) to maintain the common usage of the name. Although Takakura's type material is not extant, we consider that neotypification is unnecessary in this case because no taxonomic/nomenclatural confusion persists. We also propose to transfer Lineus fulvus to yield Kulikovia fulva comb. nov.

A Population Without Females: Males of Baseodiscus delineatus (Nemertea: Heteronemertea) Reproduce Asexually by Fragmentation.

The valenciniid heteronemertean Baseodiscus delineatus (Delle Chiaje, 1825) was originally described from Naples, Italy, and shows a circumglobal distribution from tropical to temperate seas in both hemispheres. To investigate its reproductive biology, we performed intermittent year-round sampling from 2014 to 2018 in Misaki on the Pacific coast of Honshu, Japan. Of the 40 specimens obtained, 11 were males, while 29 were immature. No female specimen was confirmed during the sampling period. We additionally observed that B. delineatus is fissiparous. A male individual in captivity reproduced asexually by spontaneous posterior fragmentation, an ability that is described here for the first time among Valenciniidae. Autotomy occurred every 2-10 days, with each of the tail fragments having undergone anterior regeneration, which took about 24-36 days before completion of internal organs, such as ocelli, cerebral organs, brain, alimentary canal, rhynchocoel, and proboscis. A review of the literature suggests that the species was absent in the Misaki region 120 years ago. We assume a recent settlement of a male founder, which has putatively reproduced asexually to yield a clonal, unisexual population in Misaki.

Ca2+ efflux via plasma membrane Ca2+-ATPase mediates chemotaxis in ascidian sperm.

When a spermatozoon shows chemotactic behavior, transient [Ca2+]i increases in the spermatozoon are induced by an attractant gradient. The [Ca2+]i increase triggers a series of stereotypic responses of flagellar waveforms that comprise turning and straight-swimming. However, the molecular mechanism of [Ca2+]i modulation controlled by the attractants is not well defined. Here, we examined receptive mechanisms for the sperm attractant, SAAF, in the ascidian, Ciona intestinalis, and identified a plasma membrane Ca2+-ATPase (PMCA) as a SAAF-binding protein. PMCA is localized in sperm flagella membranes and seems to interact with SAAF through basic amino acids located in the second and third extracellular loops. ATPase activity of PMCA was enhanced by SAAF, and PMCA inhibitors, 5(6)-Carboxyeosin diacetate and Caloxin 2A1, inhibited chemotactic behavior of the sperm. Furthermore, Caloxin 2A1 seemed to inhibit efflux of [Ca2+]i in the sperm, and SAAF seemed to competitively reduce the effect of Caloxin 2A1. On the other hand, chemotactic behavior of the sperm was disordered not only at low-Ca2+, but also at high-Ca2+ conditions. Thus, PMCA is a potent candidate for the SAAF receptor, and direct control of Ca2+ efflux via PMCA is a fundamental mechanism to mediate chemotactic behavior in the ascidian spermatozoa.