Systematic Identification of Microproteins during the Development of Drosophila melanogaster

Short open reading frame-encoded peptides (SEPs) are microproteins with less than 100 amino acids that play an essential role in the growth and development of organisms. There are plenty of short open reading frames in Drosophila melanogaster that potentially code polypeptides. We chose 11 time points during the life cycle of Drosophila to investigate microproteins, particularly those related to development. Finally, we identified a total of 410 microproteins, of which 27 were noncoding RNA-encoded proteins. Of the 410 microproteins, 74 were expressed in all stages from embryo to adults, whereas 300 microproteins were only found in one or two time points. Approximately, one-third of the microproteins were not reported previously and 44 were obtained from de novo sequencing, validated by synthetic peptides. These microproteins are related to the main bioprocesses of growth and development, such as multicellular organism reproduction, postmating behavior, and oviposition. Over half of the microproteins have predicted functional domains and are conserved across species, suggesting that these microproteins have critical functions in fly development. This work enriches the D. melanogaster proteome and provides a significant data resource for growth and development research.