Figure 6

Buc-VBM and zfVasa-BBM inhibit germ cell specification in vivo. (A) Cartoon representation of the lateral view shown in panels (B–D) of 15–18-somite stage zebrafish larvae, with the animal to the left. Imaging area boxed in red. (B) Live un-injected embryos with fluorescent germ cells (green cells (white arrow); transgenic for Buc-GFP). (C) Embryos injected with Buc-VBM and (D) zfVasa-BBM. (E) Quantification of the germ cell number. Embryos injected with either Buc-VBM (6 ± 1.0, n =18) or zfVasa-BBM (6.3 ± 1.1, n = 16) showed a reduction of germ cells compared to the un-injected embryos (12 ± 1.6, n = 15). Additionally, we did not observe a significant statistical difference between Buc-VBM and zfVasa-BBM. The data presented are averaged from three independent experiments. The Y-axis represents the average germ cell count, and the X-axis represents the injected constructs Buc-VBM and zfVasa-BBM. Error bars represent the standard deviation of the mean. (F) Hypothetical model of the Buc-zfVasa interaction. Buc (green line) binds directly to zfVasa (black line) through zfVasa-BBM (orange) and Buc-VBM (pink). During the interaction, disordered Buc-VBM adopts an α-helical structure. Conversely, Buc-VBM stimulates the ATPase activity of zfVasa (blue arrow), which is further enhanced when RNA (red line) binds to zfVasa. The helicase activity probably triggers an unknown downstream RNA processing specifying primordial germ cell development. Scale bar 50 µm. Test statistics: Student’s t-test, ** = 0.01. ns = nonsignificant.