Two Muscle-Specific and Direct Transcriptional Targets of DAF-16/FOXO Activated by Reduced Insulin/IGF-1 Signaling

Abstract

C. elegansinsulin/insulin-like growth factor 1 signaling (IIS) affects diverse physiological processes through the DAF-16/FOXO transcription factor. Despite its presence in all somatic cells, DAF-16's physiological effects, such as modulation of dauer formation, synapse maturation, axon regeneration, and adult longevity, exhibit prevalent tissue-specificity as well as tissue crosstalk. This implies that tissue-specific DAF-16 transcriptional programs contribute to the functional diversity of IIS. To further examine this possibility, we sought to identify tissue-specific and direct transcriptional targets of DAF-16 in muscle cells. Following FACS-sorting to enrich mature muscle cells from young adult animals, we compared the muscle transcriptomes under high and low IIS signaling conditions, with and without DAF-16. We further analyzed and compared the DAF-16 docking sites in muscle and intestine cells from published datasets. These analyses revealed 14 potential muscle-specific DAF-16 transcriptional targets, among which we validated two that are strongly and specifically activated by DAF-16 in muscles: a secreted protein C54F6.5 and a calcium-binding protein CEX-1/Calexcitin. Both genes exhibit DAF-16-independent non-muscle expression, explaining their low rank or absence from the current DAF-16 target lists generated by multiple independent whole-animal microarray or mRNA-sequencing analyses. These results support the notion of tissue-specific DAF-16 transcriptional programs and highlight the importance of verifying FOXO targets in a cell-type-specific manner.