Unveiling the Role of Two Rhodopsin-like GPCR Genes in Insecticide-Resistant House Flies, Musca domestica

Insecticide resistance in insects, driven by the overexpression of P450 enzymes, presents a significant challenge due to the enhanced metabolic detoxification of insecticides. Although the transcriptional regulation of P450 genes is not yet fully understood, G-protein-coupled receptor (GPCR) genes have emerged as key regulators in this process. This study is the first to associate GPCR genes with insecticide resistance in <i>Musca domestica</i>. We identified two key rhodopsin-like GPCR genes, <i>ALHF_02706.g1581</i> and <i>ALHF_04422.g2918</i>, which were significantly overexpressed in the resistant ALHF strain compared to sensitive strains. Notably, both <i>ALHF_02706.g1581</i> and <i>ALHF_04422.g2918</i> were mapped to autosome 2, where critical but unidentified regulatory factors controlling resistance and P450 gene regulation are located. This supports our hypothesis that GPCRs function as trans-regulatory factors for P450-mediated resistance. Functional analysis using transgenic <i>Drosophila</i> demonstrated that overexpression of these rhodopsin-like GPCR genes increased permethrin resistance by approximately two-fold. Specifically, <i>ALHF_02706.g1581</i> overexpression significantly upregulated the <i>Drosophila</i> resistance-related P450 genes <i>CYP12D1</i>, <i>CYP6A2</i>, and <i>CYP6A8</i>, while <i>ALHF_04422.g2918</i> increased <i>CYP6G1</i> and <i>CYP6A2</i> expression, thereby enhancing insecticide detoxification in rhodopsin-like GPCR transgenic <i>Drosophila</i> lines. These findings suggest that these rhodopsin-like GPCR genes on autosome 2 may act as trans-regulatory factors for P450-mediated resistance, underscoring their critical role in insecticide detoxification and resistance development in <i>M. domestica</i>.