Localized maternal factors are required for zebrafish germ cell formation.

Maternally supplied factors in fertilized eggs play essential roles in the establishment of primordial germ cells. In zebrafish, cytoplasm at the distal ends of the first and second cleavage furrows has been assumed to contain germ lineage determinants, since maternal transcripts of germ lineage-specific genes are localized to ends of the cleavage furrows. To investigate whether these parts of cytoplasm are required for germ cell formation, we removed all four regions of the cytoplasm by glass capillary at the 4-cell stage. Histological analysis revealed that the ablation of cytoplasm at the ends of the cleavage planes resulted in a severe reduction in the number of germ cells. In addition, the expression of germ lineage markers was eliminated by cytoplasmic ablation. These results demonstrated that cytoplasm at the distal ends of cleavage furrows is essential for germ cell formation. We also found novel localization patterns for zDazl and brul mRNAs along the cleavage planes. Our findings provide the first direct evidence that localized cytoplasmic factors are indispensable for germ cell establishment in zebrafish.