Autorepressor PsrA is required for optimal Legionella pneumophila growth in Acanthamoeba castellanii protozoa

• Published in: Molecular microbiology 2021-08, Vol.116 (2), p.624-647
• Main Authors: Graham, Christopher I., Patel, Palak G., Tanner, Jennifer R., Hellinga, Jacqueline, MacMartin, Teassa L., Hausner, Georg, Brassinga, Ann Karen C.
• Format: Article
• Language: English
• Subjects: Acanthamoeba castellanii; Acanthamoeba castellanii - microbiology; Azotobacter; Bacterial Proteins - genetics; Binding sites; cyst differentiation; Cysts; Gene expression; Gene Expression Regulation, Bacterial - genetics; Genes; Humans; Hydroxybutyrates - metabolism; Inclusions; infection host model; Intracellular; intracellular growth; Legionella pneumophila; Legionella pneumophila - genetics; Legionella pneumophila - growth & development; Legionnaires' disease bacterium; Life cycle analysis; Macrophages; Macrophages - microbiology; Morphogenesis; Polyesters - metabolism; Promoter Regions, Genetic - genetics; Protozoa; Pseudomonas; Repressor Proteins - genetics; Sigma factor; Sigma Factor - genetics; Stationary phase; Stringent response; Transcription; Transcription Factors - genetics; transcription regulator; Transcription, Genetic - genetics; Virulence
• ISSN: 0950-382X; 1365-2958
• DOI: 10.1111/mmi.14760

Legionella pneumophila possesses a unique intracellular lifecycle featuring distinct morphological stages that include replicative forms and transmissive cyst forms. Expression of genes associated with virulence traits and cyst morphogenesis is concomitant, and governed by a complex stringent response based‐regulatory network and the stationary phase sigma factor RpoS. In Pseudomonas spp., rpoS expression is controlled by the autorepressor PsrA, and orthologs of PsrA and RpoS are required for cyst formation in Azotobacter. Here we report that the L. pneumophila psrA ortholog, expressed as a leaderless monocistronic transcript, is also an autorepressor, but is not a regulator of rpoS expression. Further, the binding site sequence recognized by L. pneumophila PsrA is different from that of Pseudomonas PsrA, suggesting a repertoire of target genes unique to L. pneumophila. While PsrA was dispensable for growth in human U937‐derived macrophages, lack of PsrA affected bacterial intracellular growth in Acanthamoeba castellanii protozoa, but also increased the quantity of poly‐3‐hydroxybutyrate (PHB) inclusions in matured transmissive cysts. Interestingly, overexpression of PsrA increased the size and bacterial load of the replicative vacuole in both host cell types. Taken together, we report that PsrA is a host‐specific requirement for optimal temporal progression of L. pneumophila intracellular lifecycle in A. castellanii. Legionella pneumophila is an intracellular parasite of Acanthamoeba castellanii protozoa, in which it undergoes a biphasic lifecycle. Intracellular bacterial replication is supported by host‐derived nutrients that once depleted, triggers a regulatory network to mediate the switch to development of transmissive resilient cyst‐like forms that are released upon lysis of the protozoan host cell. We show that precise levels of transcriptional regulator PsrA are required for temporal progression and completion of the lifecycle. Elevated PsrA levels impede the switch, whereas absence of PsrA fosters low bacterial load of cyst‐like forms featuring extra PHB granules.