Sufficient Numbers of Early Germ Cells Are Essential for Female Sex Development in Zebrafish

Abstract

The sex determination for zebrafish is controlled by a combination of genetic and environmental factors. The determination of sex in zebrafish has been suggested to rely on a mechanism that is affected by germ cell-derived signals. To begin our current study, a simplified and efficient germ cell-specific promoter of the dead end (dnd) gene was identified. Utilizing the metrodinazole (MTZ)/ bacterial nitroreductase (NTR) system for inducible germ cell ablation, several stable Tg (dnd:NTR-EGFP-3'UTR) and Tg (dnd:NTR-EGFP+3'UTR) zebrafish lines were then generated with the identified promoter. A thorough comparison of the expression patterns and tissue distributions of endogenous dnd and ntr-egfp transcripts in vivo revealed that the identified 2032-bp zebrafish dnd promoter can recapitulate dnd expression faithfully in stable transgenic zebrafish. The correlation between the levels of the germ cell-derived signals and requirement for maintaining the female fate has been also explored with different durations of the MTZ treatments. Our results revealed the decreasing ratios of female presented in the treated transgenic group are fairly associated with the reducing levels of the early germ cell-derived signals. After the juvenile transgenic fish treated with 5 mM MTZ for 20 days, all MTZ-treated transgenic fish exclusively developed into males with subfertilities. Taken together, our results identified here a simplified and efficient dnd promoter, and provide clear evidence indicating that it was not the presence but the sufficiency of signals derived from germ cells that is essential for female sex development in zebrafish. Our model also provides a unique system for sex control in zebrafish studies.

Hybrid Sterility in Fish Caused by Mitotic Arrest of Primordial Germ Cells

“…zebrafish, ablation of PGCs by knockdown or knockout studies for the dead end (dnd) gene promotes the masculinization of genetically female fish (Ciruna et al 2002;Slanchev et al 2005;Dai et al 2015;Li et al 2017). Further, during the embryonic stage of zebrafish, a certain number of PGCs must be present in the genital ridges for the subsequent ovarian development in females to successfully proceed (Tzung et al 2015).…”

Generation of all-male-like sterile zebrafish by eliminating primordial germ cells at early development

“…Therefore, the fish would be sterile if their PGCs are eliminated in the embryos at early development. Actulaly, in the absence of PGCs, the somatic cells of zebrafish and medaka ( Oryzias latipes ) gonads tend to spontaneously differentiate into male characteristics, while gonadal differentiation with female characteristics requires PGCs 15 , 27 – 31 , although the underlying mechanisms remain to be elucidated. Consistenly, Wong and Collodi reported that overexpressing stromal-derived factor 1a (Sdf1a), providing the directional cue that guides the migration of PGCs to the gonadal tissue in a gradient, disrupted the normal PGC migration pattern, resulting that the embryos developed into sterile males 9 .…”

“…However, 5 mM is not optimal amount to kill all the germ cells in zebrafish embryos because Dai et al . administrated 5 mM MTZ to treat transgenic zebrafish Tg( dnd :NTR-EGFP + 3′UTR) for 20 days from 18 dpf and found that all MTZ-treated transgenic fish exclusively developed into males with subfertilities 15 .…”

“…Dai et al . employed dnd promoter to construct NTR transgenic fish and demonstrated that MTZ treatment could induce male transformation and severely impair reproduction capability 15 .…”

Zebrafishdazlregulates cystogenesis upstream of the meiotic transition and germline stem cell specification and independent of meiotic checkpoints

“…Timing of entry into meiosis and OLC abundance are thought to contribute to sexual fate determination, such that more GCs promote female fate (Ye et al, 2019). Conversely, low numbers of germ cells and OLCs drive male development (Dai et al, 2015;Orban et al, 2009;Tzung et al, 2015). Despite these correlations highlighting the importance of proper development of OLCs, little is known about the actual mechanisms and cell biology regulating GC numbers and meiotic entry in zebrafish.…”