Transmembrane proteins such as G-protein coupled receptors (GPCRs) mediate molecular signal transduction across lipid bilayers via stimulus-triggered conformational changes. Drawing inspiration from these biomolecular mechanisms, we engineered a single-stranded amphiphilic DNA hairpin bearing dual cholesterol anchors that axially inserts into lipid membranes and undergoes toehold-mediated structural reconfiguration upon target nucleic acid hybridization. This conformational change facilitates transbilayer signaling while preserving membrane integrity. Systematic optimization of hairpin stem length, cholesterol orientation, and bilayer fluidity enabled robust and stable membrane incorporation in synthetic vesicles and live mammalian cells. All-atom molecular dynamics simulations demonstrate that cholesterol functionalization stabilizes the hairpin conformation within membrane environments, with target sequence specificity and lipid milieu governing the dynamic behavior of the sensor. Quantitative analyses revealed high-efficiency signal transduction, reaching 80%–90% responsiveness within 90–120 min, tightly linked to sequence-dependent target recognition and selective signal propagation from defined cell populations. Integration with hybridization chain reaction (HCR) and proximal split-initiator induced HCR amplified the signal up to ten-fold, enabling lysis-free detection of low-abundance intracellular RNA. The observation of dual-mode signal transduction mechanisms provides mechanistic insight into DNA branch migration events across membrane during information transfer. This modular and chemically programmable framework establishes a new paradigm for reconstituting GPCR-like molecular signaling with nucleic acids, unlocking broad utility for live-cell diagnostics, synthetic biology, and targeted therapeutics.
Sasmal et al. (Sun,) studied this question.