A transposon-introduced G-quadruplex motif is selectively retained and constrained to downregulate CYP321A1

Insects utilize xenobiotic compounds to up- and downregulate cytochrome P450 monooxygenases (P450s) involved in detoxification of toxic xenobiotics including phytochemicals and pesticides. G-quadruplexes (G4)-forming DNA motifs are enriched in the promoter regions of transcription factors and function as cis-acting elements to regulate these genes. Whether and how P450s gain and keep G4 DNA motifs to regulate their expression still remain unexplored. Here, we show that CYP321A1, a xenobiotic-metabolizing P450 from Helicoverpa zea, a polyphagous insect of economic importance, has acquired and preserved a G4 DNA motif by selectively retaining a transposon known as HzIS1-3 that carries this G4 DNA motif in its promoter region. The HzIS1-3 G4 DNA motif acts as a silencer to suppress the constitutive and induced expression of CYP321A1 by plant allelochemicals flavone and xanthotoxin through folding into an intramolecular parallel or hybrid-1 conformation in the absence or presence of K⁺. The G4 ligand N-methylmesoporphyrin IX (NMM) strengthens the silencing effect of HzIS1-3 G4 DNA motif by switching its structure from hybrid-1 to hybrid-2. The enrichment of transposons in P450s and other environment-adaptation genes implies that selective retention of G4 DNA motif-carrying transposons may be the main evolutionary route for these genes to obtain G4 DNA motifs.