Surface-associated microbial systems are hotspots for the spread of plasmid-encoded antibiotic resistance, but how surface association affects plasmid transfer and proliferation remains unclear. Surface association enables prolonged spatial proximities between different populations, which promotes plasmid transfer between them. However, surface association also fosters strong metabolic interactions between different populations, which can direct their spatial self-organization with consequences for plasmid transfer and proliferation. Here, we hypothesize that metabolic interactions direct the spatial self-organization of different populations and, in turn, regulate the spread of plasmid-encoded antibiotic resistance. We show that resource competition causes populations to spatially segregate, which represses plasmid transfer. In contrast, resource cross-feeding causes populations to spatially intermix, which promotes plasmid transfer. We further show that the spatial positionings that emerge from metabolic interactions determine the proliferation of plasmid recipients. Our results demonstrate that metabolic interactions are important regulators of both the transfer and proliferation plasmid-encoded antibiotic resistance.