["Archives of Insect Biochemistry and Physiology, Volume 122, Issue 4, August 2026. ", "HcEcR regulates chitin biosynthesis via 20E signaling rather than direct transcriptional activation in Hyphantria cunea larvae\n\n\n\n\n\nABSTRACT\nThe ecdysone receptor (EcR) is a key subunit of the heterodimeric EcR‐USP (ultraspiracle) receptor, whose role in insect growth and development has been well established. In contrast, its role in regulating chitin biosynthesis, particularly at transcriptional level received less attention. Here, we identified the EcR from Hyphantria cunea and investigated its roles in larval growth and development, particularly in chitin biosynthesis, via dsRNA‐mediated HcEcR knockdown. Bioassays showed that silencing of HcEcR significantly increased larval mortality and caused developmental defects in H. cunea. Meanwhile, HcEcR knockdown significantly downregulated the expression of genes in the chitin biosynthetic pathway. The yeast one‐hybrid (Y1H) assay indicated that HcEcR alone did not interact with the HcCHSA or HcHK2 promoters, suggesting that, independent of HcUSP, HcEcR might not transcriptionally regulate chitin biosynthesis pathway genes. Additionally, HcEcR knockdown significantly suppressed the expression of 20E‐induced response genes, but dramatically increased glucose, trehalose and glycogen levels in H. cunea larvae. Overall, HcEcR was pivotal for the growth, molting and chitin biosynthesis in H. cunea larvae. Meanwhile, HcEcR knockdown suppressed epidermal chitin synthesis probably by downregulating 20E signaling in H. cunea larvae. Furthermore, inhibiting chitin synthesis might reduce substrate consumption for chitin synthesis, thereby resulting in an increase of carbohydrate levels in the larvae. Moreover, HcEcR alone might not transcriptionally regulate chitin biosynthetic pathway genes. The findings provide novel insights into the physiological role of EcR in chitin biosynthesis and identify potential molecular targets for pest control."]