Regulatory cis- and trans-elements of mitochondrial D-loop-driven reporter genes in budding tunicates.

To unveil the underlying mechanism of mitochondrial gene regulation associated with ageing and budding in the tunicate Polyandrocarpa misakiensis, mitochondrial non-coding-region (NCR)-containing reporter genes were constructed. PmNCR2.3K/GFP was expressed spatiotemporally in a pattern quite similar to mitochondrial 16SrRNA.

Histone methylation codes involved in stemness, multipotency, and senescence in budding tunicates.

We examined the dynamics of nuclear histone H3 trimethylation related to cell differentiation and aging in a budding tunicate, Polyandrocarpa misakiensis. Throughout zooidal life, multipotent epithelial and coelomic cell nuclei showed strong trimethylation signals at H3 lysine27 (H3K27me3), consistent with the results of western blotting. Epidermal H3K27me3 repeatedly appeared in protruding buds and disappeared in senescent adult zooids.

Senescence-associated superoxide dismutase influences mitochondrial gene expression in budding tunicates.

A recent study has shown that in the budding tunicate Polyandrocarpa misakiensis, the mitochondrial respiratory chain (MRC) dramatically attenuates the gene activity during senescence. In this study, we examined the possible involvement of superoxide dismutase (SOD) in the attenuation of gene expression of cytochrome c oxidase subunit 1 (COX1) in aged zooids. By RT-PCR and in situ hybridization, Cu/Zn-SOD (SOD1) was found to be expressed in most cells and tissues of buds and juvenile zooids but showed a conspicuous decline in senescent adult zooids, except in the gonad tissue in which the cytoplasm of juvenile oocytes was stained heavily.

Molecular anatomy of tunicate senescence: reversible function of mitochondrial and nuclear genes associated with budding cycles.

Zooids of the asexual strain of Polyandrocarpa misakiensis have a lifespan of 4-5 months; before dying, they produce many buds, enabling continuation of the strain. This study was designed to investigate the nature of gene inactivation and reactivation during this continuous process of senescence and budding. During senescence, the zooidal epidermis showed acid β-galactosidase activity, lost proliferating cell nuclear antigen immunoreactivity and became ultrastructurally worn, indicating that the epidermis is a major tissue affected by the ageing process.

RACK1 regulates mesenchymal cell recruitment during sexual and asexual reproduction of budding tunicates.

A homolog of receptor for activated protein kinase C1 (RACK1) was cloned from the budding tunicate Polyandrocarpa misakiensis. By RT-PCR and in situ hybridization analyses, PmRACK1 showed biphasic gene expression during asexual and sexual reproduction. In developing buds, the signal was exclusively observed in the multipotent atrial epithelium and undifferentiated mesenchymal cells that contributed to morphogenesis by the mesenchymal-epithelial transition (MET).

Role of Vasa, Piwi, and Myc-expressing coelomic cells in gonad regeneration of the colonial tunicate, Botryllus primigenus.

In the colonial tunicate, Botryllus primigenus Oka, gonads consist of indifferent germline precursor cells, the primordial testis and ovary, and mature gonads, of which the immature gonad components can be reconstructed de novo in vascular buds that arise from the common vascular system, although the mechanism is uncertain. In this study, we investigated how and what kinds of cells regenerated the gonad components. We found that few Vasa-positive cells in the hemocoel entered the growing vascular bud, where their number increased, and finally developed exclusively into female germ cells.

Germline cell formation and gonad regeneration in solitary and colonial ascidians.

The morphology of ascidian gonad is very similar among species. The testis consists of variable number of testicular follicles; the ovary consists of ovarian tubes that are thickened forming the germinal epithelium with stem cells for female germ cells with the exception of botryllid ascidians. Peculiar accessory cells that would be germline in origin accompany the oocytes.

Piwi-expressing hemoblasts serve as germline stem cells during postembryonic germ cell specification in colonial ascidian, Botryllus primigenus.

Animals that propagate asexually are exciting models to investigate the cellular system, which produces germline cells constitutively throughout life. The present research investigated whether piwi was a germline-specific marker in the colonial ascidian Botryllus primigenus. An approximately 2.

Hemoblasts in colonial tunicates: are they stem cells or tissue-restricted progenitor cells?

Colonial tunicates have hemoblasts, which are undifferentiated coelomic cells that play a key role in tissue renewal during reproduction and regeneration. Some hemoblasts differentiate into somatic lineage cells such as endodermal multipotent epithelial, cardiac and body-wall muscle, and blood cells. There is no well established evidence that somatic hemoblasts are stem cells.

The role of Nanos homologue in gametogenesis and blastogenesis with special reference to male germ cell formation in the colonial ascidian, Botryllus primigenus.

We examined the structure, expression, and possible functions of a nanos homologue during gametogenesis and blastogenesis in the colonial ascidian Botryllus primigenus. An approximately 1.3-kb-long cDNA was cloned; it was termed BpNos since the deduced amino acid sequence (288 aa) contained 2 Nanos-like CCHC zinc finger motifs.

Cell proliferation dynamics of somatic and germline tissues during zooidal life span in the colonial tunicate Botryllus primigenus.

Botryllus primigenus is a colonial tunicate in which three successive generations develop synchronously. To identify proliferation centers and possible adult stem cells during asexual reproduction, somatic and germline cells were labeled with 5-bromo-2'-deoxyuridine (BrdU). In the youngest generation, multipotent epithelial cells exhibited an average labeling index (LI) of 30% 24 hr after BrdU injection.

Multipotent epithelial cells in the process of regeneration and asexual reproduction in colonial tunicates.

The cellular and molecular features of multipotent epithelial cells during regeneration and asexual reproduction in colonial tunicates are described in the present study. The epicardium has been regarded as the endodermal tissue-forming epithelium in the order Enterogona, because only body fragments having the epicardium exhibit the regenerative potential. Epicardial cells in Polycitor proliferus have two peculiar features; they always accompany coelomic undifferentiated cells, and they contain various kinds of organelles in the cytoplasm.

Involvement of vasa homolog in germline recruitment from coelomic stem cells in budding tunicates.

We investigated the mechanism by which germline cells are recruited in every asexual reproductive cycle of the budding tunicate Polyandrocarpa misakiensis using a vasa homolog (PmVas) as the germline-specific probe. A presumptive gonad of Polyandrocarpa arose as a loose cell aggregate in the ventral hemocoel of a 1-week-old developing zooid. It developed into a compact clump of cells and then separated into two lobes, each differentiating into the ovary and the testis.

In vitro culture of mesenchymal lineage cells established from the colonial tunicate Botryllus primigenus.

Body trunks were isolated from juvenile zooids of the Japanese colonial tunicate Botryllus primigenus and cultured in vitro to establish tissue-specific cell lines. Epidermal cells from some explants spread and formed a flat sheet consisting of vacuolated cells. They then dissociated into single cells, and their growth stopped within two weeks.

Postembryonic epigenesis of Vasa-positive germ cells from aggregated hemoblasts in the colonial ascidian, Botryllus primigenus.

We investigated whether Vasa was a germline-specific marker in the colonial ascidian Botryllus primigenus, and whether it was inducible epigenetically in the adult life span. We cloned a Botryllus Vasa homologue (BpVas). The deduced open reading frame encoded 687 amino acid residues.