Pseudomonas aeruginosa is known to cause chronic infections in Cystic Fibrosis (CF) patients by establishing itself in their lungs for extended periods. The RNA degradosome is an important regulatory machinery in bacteria that controls environmental adaptation by acting on RNA. The RNA degradosome main component, the endoribonuclease RNase E, acts as a scaffold for other RNA degradosome components while possessing its own catalytic activity. The composition and importance of the P. aeruginosa RNA degradosome during lung colonization in CF patients are not yet established. Here, we address these questions by using different RNase E mutants and an artificial sputum medium (ASM) that mimics the lung mucus of CF patients. First, we performed phenotypic analyses of the mutants in ASM and in the traditional laboratory medium NYB to compare the two media. Second, we searched for new components of the RNA degradosome in ASM, as the machinery composition changes according to the bacterial environment. Third, we set up a transposon-sequencing protocol that will allow to study the contribution of RNA degradosome assembly to bacterial fitness. Overall, we showed that P. aeruginosa RNA degradosome action and composition can be different according to the growing medium, which may give importance to studying bacteria in a medium that mimics the infectious environment.