Asperous coordinates regeneration timing by regulating damage-induced WNT signaling

Tissue regeneration requires precise control of signaling pathways to direct proliferation, differentiation, and patterning. While early responses to injury are well characterized, how differentiation is coordinated during later stages remains unclear. Here, we identify Asperous (Aspr), an epidermal growth factor (EGF)-repeat protein, as a regeneration-specific regulator in Drosophila wing discs. Aspr is dispensable for wing development but is strongly induced within 24 h post-injury. Maintaining aspr expression inhibits differentiation and alters reparative growth, while loss impairs regeneration. Structural prediction and localization analyses show Aspr is an extracellular protein secreted and associated with extracellular vesicles (EVs), where it co-localizes with the Wingless/Wnt (WNT) ligand Wingless (Wg). We find that Aspr regulates post-injury but not developmental Wg signaling, potentially by influencing its secretion or availability via EVs. These findings suggest Aspr regulates WNT activity to ensure proper timing of cell fate specification during regeneration, revealing a mechanism by which signaling dynamics are temporally controlled during tissue repair.