High-Dimensional CRISPR-SERS Interactomics Tracks the Topological Evolution of Plant Viral Pathogenesis and Therapeutic Response

Abstract

Precision agriculture is currently limited by a reliance on viral load quantification, a static metric that obscures the dynamic molecular arms race between invading pathogens and host immunity. To overcome this limitation, we present a multidimensional surface-enhanced Raman spectroscopy (SERS) platform designed to map the real-time topology of the host–virus interactome. Overcoming the spectral constraints of conventional assays, we engineered a de novo library of isomeric Raman reporters via rational positional and electronic tuning, enabling high-density spectral coding. Furthermore, by integrating CRISPR-dCas9 as an isothermal recognition module, we ensure multiplexed signals accurately reflect the stoichiometric integrity of viral and host gene expression due to the elimination of thermodynamic biases of traditional DNA hybridization. Applying this platform to the Nicotiana benthamiana–Potato virus Y (PVY) pathosystem, we dissect the longitudinal evolution of infection under ningnanmycin (NNM) treatment. Beyond enabling presymptomatic diagnosis at Day 1, our interactome analysis reveals that chronic infection stabilizes into a regulatory triangle clamped by the Vpg–PUB4 interface, whereas therapeutic relapse manifests as a chaotic, Vpg-centralized network. This work establishes interactome topology as a superior diagnostic metric to viral load, providing a blueprint for identifying drug resistance mechanisms and targeting the specific protein-protein interactions that drive viral persistence.