Mutations in mexZ, encoding a negative regulator of the expression of the mexXY efflux pump genes, are frequently acquired by Pseudomonas aeruginosa at early stages of lung infection. Although traditionally related to resistance to the first-line drug tobramycin, mexZ mutations are associated with low-level aminoglycoside resistance when determined in the laboratory, suggesting that their selection during infection may not be necessarily, or only, related to tobramycin therapy. Here, we show that mexZ-mutated bacteria tend to accumulate inside the epithelial barrier of a human airway infection model, thus colonising the epithelium while being protected against diverse antibiotics. This phenotype is mediated by overexpression of lecA, a quorum sensing-controlled gene, encoding a lectin involved in P. aeruginosa tissue invasiveness. We find that lecA overexpression is caused by a disrupted equilibrium between the overproduced MexXY and another efflux pump, MexAB, which extrudes quorum sensing signals. Our results indicate that mexZ mutations affect the expression of quorum sensing-regulated pathways, thus promoting tissue invasiveness and protecting bacteria from the action of antibiotics within patients, something unnoticeable using standard laboratory tests.