This project explores the origins of genotype-phenotype (g/P) linkage in cell-free systems for bacteriophage assembly. Building on findings of Levrier et al. (2024), who demonstrated co-assembly of phage mutants in vitro can yield significant g/P coupling, contrary to the expectation of random phenotype assortment in bulk reactions, I am investigating how such coupling emerges in the absence of cellular compartmentalisation.
The experimental system utilised involves two phage mutant genomes, differing by a single tail fiber mutation that confers infectivity to distinct host bacteria (therefore conferring orthogonal infectivity system). An expected outcome would be random packaging of genomes and phenotypes produced by co-assembly in bulk, thus precluding g/P coupling and selection. Yet, Levrier et al. observed, genotype and phenotype tend to link, allowing for direct selection of desired phenotypes from a mixture. This challenges the current paradigm of cellular compartmentalisation as the basis of phages’ genotype-phenotype linkage, suggesting fundamental questions about inherent capacity of self-assembly, physical crowding, and evolutionary constraints in non-compartmentalised environments.
In parallel to the experimental efforts, we are developing a mechanistic model to describe the non-equilibrium dynamics of phage assembly, incorporating spatial diffusion, local synthesis, and the kinetics of capsid and tail fiber integration. My goal is to understand the physicochemical and evolutionary parameters that drive coupling of the corresponding genomes and proteins in bulk, and the minimal and optimal conditions for selection to operate.
