Transcriptomic Profiling of Wild-Type C. elegans Exhibiting High and Low Exopher Production

We performed RNA sequencing on age-synchronized wild-type C. elegans that produced either high numbers of exophers (>20 per animal) or low numbers of exophers (<2 per animal) on adulthood day 2. This approach mirrors strategies using natural transcriptional heterogeneity within isogenic populations to uncover molecular determinants of phenotypic variation. The transcriptomes of high- and low-exopher producers differed profoundly. In total, 609 genes were significantly upregulated and 665 genes were significantly downregulated in high-exopher animals (p adj < 0.05, |log2FC| > 1). These results demonstrate that exopher production is associated with distinct transcriptional states, indicating that variation in exopher output reflects meaningful underlying molecular differences within genetically identical populations. Overall design: This study investigates transcriptomic differences between wild-type age-synchronized C. elegans that produce high versus low numbers of exophers. Animals were synchronized and scored on adulthood day 2 based on the number of visible exophers per worm. Two phenotypic groups were defined: (1) high-exopher producers (more than 20 exophers per animal) and (2) low-exopher producers (fewer than 2 exophers per animal). These two groups represent the main experimental conditions and constitute the variables under investigation. For each condition, independent biological samples were generated from separate populations of worms classified into the high- or low-exopher group. The design therefore consists of comparative transcriptomic profiling of high- vs low-exopher wild-type animals. No additional genetic mutants or treatment conditions are included in this submission.