Evolution and development of segmented body plan revealed by engrailed and wnt1 gene expression in the annelid Alitta virens

Segmentation is one of the most striking features of bilaterians, and understanding its mechanisms provides insights into the evolution of body plans. In annelids, segmentation occurs at different developmental stages through a variety of processes, yet the molecular pathways remain underexplored. Aiming to compare segmentation patterns in ontogeny and phylogeny, we analysed the expression of Avi-en (homologous to engrailed ) and Avi-wnt1 in the nereidid polychaete Alitta virens . Using in situ hybridization, immunofluorescence, and cell proliferation assays, we mapped the spatiotemporal expression of these genes across embryonic, larval, and postlarval stages. We found that Avi-en was expressed in solid lateral domains early in the unsegmented protrochophore stage and progressed through a metameric pattern, while Avi-wnt1 expression appeared later, also aligning with segmental boundaries. At the nectochaete stage, the posterior domain of Avi-en expression in the growth zone expanded and splitted into two due to increased cell proliferation. The postlarval segment primordium then developed progressively, culminating in the activation of Avi-wnt1 at the posterior border. According to available published data, the revealed pattern of gradual segment formation is unique to nereidids. The observed divergence in gene expression and cell proliferation across annelids suggests that segmentation in bilaterians did not arise from a common ancestral mechanism. Our study enhances future progress in understanding the evolution of body patterning by providing a foundation for future comparisons.