Combinatorial interactions between viral proteins expand the potential functional landscape of the tomato yellow leaf curl virus proteome

Viruses manipulate the cells they infect in order to replicate and spread. Due to strict size restrictions, viral genomes have reduced genetic space; how the action of the limited number of viral proteins results in the cell reprogramming observed during the infection is a long-standing question. Here, we explore the hypothesis that combinatorial interactions may expand the functional landscape of the viral proteome. We show that the proteins encoded by a plant-infecting DNA virus, the geminivirus tomato yellow leaf curl virus (TYLCV), physically associate with one another in an intricate network, as detected by a number of protein-protein interaction techniques. Importantly, our results indicate that intra-viral protein-protein interactions can modify the subcellular localization of the proteins involved. Using one particular pairwise interaction, that between the virus-encoded C2 and CP proteins, as proof-of-concept, we demonstrate that the combination of viral proteins leads to novel transcriptional effects on the host cell. Taken together, our results underscore the importance of studying viral protein function in the context of the infection. We propose a model in which viral proteins might have evolved to extensively interact with other elements within the viral proteome, enlarging the potential functional landscape available to the pathogen. Author summary Viruses are obligate intracellular parasites that depend on the molecular machinery of their host cell to complete their life cycle. For this purpose, viruses co-opt host processes, modulating or redirecting them. Most viruses have small genomes, and hence limited coding capacity. During the viral invasion, virus-encoded proteins will be produced in large amounts and coexist in the infected cell, which enables physical or functional interactions among viral proteins, potentially expanding the virus-host functional interface by increasing the number of potential targets in the host cell and/or synergistically modulating the cellular environment. Examples of interactions between viral proteins have been recently documented for both animal and plant viruses; however, the hypothesis that viral proteins might have a combinatorial effect, which would lead to the acquisition of novel functions, lacks systematic experimental validation. Here, we use the geminivirus tomato yellow leaf curl virus (TYLCV), a plant-infecting virus with reduced proteome and causing devastating diseases in crops, to test the idea that combinatorial interactions between viral proteins exist and might underlie an expansion of the functional landscape of the viral proteome. Our results indicate that viral proteins prevalently interact with one another in the context of the infection, which can result in the acquisition of novel functions.