Data-Independent Acquisition-Based Quantitative Proteomics Analysis of Fertile Red Eggs and Spermatozoa in Hermatypic Coral Galaxea fascicularis: Revealing Key Proteins Related to Gamete Maturation and Fertilization

Sexually propagated scleractinian corals are in high demand for coral reef restoration. However, for threatened reef-building corals, many of the molecular mechanisms related to their reproduction remain largely unknown, which forms a major bottleneck in the large-scale cultivation of sexually reproducing corals. In this study, we analyzed the proteomic signatures of red eggs and spermatozoa from the ecologically significant coral Galaxea fascicularis, using a data-independent acquisition mass spectrometry (DIA-MS) method. A total of 7741 and 7279 proteins from mature red eggs and spermatozoa were identified, respectively. Among these proteins, 596 proteins were spermatozoa-specific and 1056 were egg-specific. Additionally, a total of 4413 differentially abundant proteins (DAPs) were identified, among which 3121 proteins were up-regulated in red eggs and 1292 proteins were up-regulated in spermatozoa. Furthermore, anenrichment analyses showed that DAPs identified in red eggs were mainly involved in the progesterone-mediated oocyte maturation pathway and lectin pathway; and DAPs detected in spermatozoa were mainly involved in the insulin secretion pathway and metabolic pathways for the generation of energy. This result will contribute to the discovery of the intrinsic regulation pathway of gamete maturation and fertilization. Furthermore, at least 57 proteins associated with gamete maturation and reproduction were identified, including the red fluorescent protein (RFP), vitellogenin proteins (VG), the egg protein (EP), the testis-specific serine/threonine-protein kinase family (TSSKs), and the EF-hand Ca2+-binding protein family (EFHC1 and EFHC2). Particularly, the third yolk protein EUPHY was reported for the first time in G. fascicularis. In conclusion, this study unveiled groundbreaking molecular insights into coral sexual reproduction, paving the way for more effective conservation and sustainable development of coral reef ecosystems