An insight into the development stages and proteome of the early-stage embryo in the horseshoe crab species T. gigas

The Indo-Pacific horseshoe crab, Tachypleus gigas is often referred to as a living fossil due to its 450 million years ancestry. It plays a critical ecological role in coastal ecosystems and offers unique insights into arthropod evolution. Despite being investigated for more than a century, its early embryonic development remains poorly understood. In this study, we identified and presented the first comprehensive proteomic characterization of the early-stage embryo of T. gigas from stage 2 to stage 5 using microscopy and mass spectrometry-based approaches, respectively. Peptides were analyzed in an EASY-nLC 1200 system coupled to an Orbitrap Fusion mass spectrometer, leading to the identification of 388 proteins. The dataset revealed enrichment in proteins associated with cellular growth, morphogenesis, cytoskeletal organization, and metabolic regulation. Functional annotation and pathway enrichment were performed using STRING v12.0, with Gene Ontology analysis highlighting key pathways related to energy metabolism, transcriptional control, and immune regulation. These findings provide new molecular perspectives on embryogenesis in T. gigas and offer a valuable reference point for future studies on chelicerate development, evolutionary biology, and species conservation.