Comparative transcriptomic profiling reveals metabolic and regulatory features of Drosophila melanogaster S2 cells relative to newly hatched larval tissues.

Abstract

Background: Drosophila melanogaster S2 cells are widely used as an in vitro model system and have undergone extensive adaptation during long-term culture. Understanding how their transcriptional programs differ from in vivo tissues is essential for interpreting their biological characteristics and experimental utility.

Objective: This study aimed to characterize transcriptomic differences between Drosophila S2 cells and newly hatched larval tissues, with a focus on identifying metabolic, regulatory, and proliferative features associated with the long-term maintenance of S2 cells in vitro.

Methods: RNA sequencing was performed on S2 cells and newly hatched larvae. Differentially expressed genes (DEGs) were identified using edgeR (FDR < 0.05, |log2FC| ≥ 1). Gene Ontology (GO) and KEGG enrichment analyses were used to investigate functional changes. Protein-protein interaction (PPI) networks were constructed based on STRING data to identify hub genes, and selected genes were validated using quantitative real-time PCR (qRT-PCR).

Results: A total of 5,937 DEGs were detected between S2 cells and larval tissues. S2 cells displayed pronounced upregulation of genes linked to amino acid metabolism, lipid biosynthesis, cell cycle progression, protein turnover, and RNA interference pathways, whereas genes associated with development and differentiation were broadly downregulated. PPI analysis highlighted 10 hub genes-including P5CS, GluProRS, ND-ACP, Ubi-p63E, and Dcr-2-that represent central nodes in metabolic regulation, protein homeostasis, transcriptional control, and stress response. These features collectively reflect a transcriptional state shaped by long-term in vitro adaptation.

Conclusions: This comparative analysis provides a comprehensive overview of transcriptomic and regulatory differences between S2 cells and in vivo larval tissues. The results clarify key molecular characteristics of S2 cells and offer a useful reference for their application in functional genomics, metabolism research, and cell-based assays.