Maternal and zygotic expression of a nanos-class gene in the leech Helobdella robusta: primordial germ cells arise from segmental mesoderm

The nanos-class gene of the leech Helobdella robusta (Hro-nos) is present as a maternal transcript whose levels decay during cleavage; HRO-NOS protein is more abundant in the D quadrant cells relative to the A, B, and C quadrants; and HRO-NOS is more abundant in the ectodermal precursor cell (DNOPQ) than in its sister mesodermal precursor (DM) (Pilon and Weisblat, 1997). Here, using in situ hybridization, we show that Hro-nos mRNA is broadly distributed throughout the zygote, is concentrated in both animal and vegetal teloplasm during stage I and is at higher levels in DNOPQ than in DM at stage 4b. Hro-nos expression increases after stage 7, as judged by in situ hybridization, developmental RT-PCR, and western blots; this increase must therefore represent later zygotic expression. Of particular interest, during stages 9 and 10, each of 11 mid-body segments (M8-M18) has a pair of Hro-nos positive 'spots' comprising of one or two large cells each. These spots later disappear in an anteroposterior progression. We find that these Hro-nos-expressing cells are of mesodermal origin, arising in a segmentally iterated manner from the M lineage, and correspond to cells previously proposed as primordial germ cells (PGCs; Burger, 1891; Weisblat and Shankland, 1985). These results support the proposal that nanos-class genes functioned in the specification of germline cells in the ancestral bilaterian and possibly in a separate process related to embryonic polarity in the ancestral protostome. Key Words: annelid; leech; Helobdella; germline; nanos.