FECH negatively regulates protoporphyrin IX accumulation and shell coloration in the dwarf surf clam Mulinia lateralis

Abstract

Shell color in mollusks, shaped by pigment synthesis and deposition, offers a readily observable natural trait. The dwarf surf clam Mulinia lateralis serves as an ideal bivalve model but has a uniform white shell, which limits its utility for genetic studies. In this study, we aimed to explore the feasibility of constructing a shell color‑based endogenous reporter by inducing pigment accumulation. Pigment feeding experiments showed that exogenous pigments, especially hemin, can efficiently accumulate in tissues (p < 0.05) and induce visible changes in shell color. By screening the porphyrin metabolic pathways responsible for heme biosynthesis, we identified five key regulatory genes (PBGD, UROS, UROD, TSPO, FECH). Multiple sequence alignment and phylogenetic analysis confirmed that these genes are evolutionarily conserved across species, implying similar physiological functions. Spatiotemporal expression analysis revealed that FECH is highly expressed from the trochophore to the spat stage and in adult tissue, suggesting a key role in shell pigment deposition. RNA interference (RNAi)‑mediated knockdown of MlFECH significantly suppressed its expression in the gill, mantle, and digestive gland tissues, leading to an obvious decrease in heme content and a significant accumulation of ELISA-detected protoporphyrin IX (PPIX) (p < 0.05). Consequently, the shell color of M. lateralis changed from white to brownish‑yellow, and the visceral mass turned reddish‑brown. These results support a role for FECH in regulating PPIX accumulation, and provide evidence that increased ELISA‑detected PPIX levels are associated with shell color formation. Thus, the FECH/PPIX axis represents a promising proof-of-concept candidate with potential for developing an endogenous, non‑invasive visible reporter system. With further systematic optimization and technical engineering, this system could potentially enable tracking gene expression or visualizing gene editing events in M. lateralis. This work provides a foundation for using shell color as a simple and economical reporter in bivalves, particularly in species where stable gene editing remains challenging.