The Legionella pneumophila effector Ceg4 is a phosphotyrosine phosphatase that attenuates activation of eukaryotic MAPK pathways

Abstract

Host colonization by Gram-negative pathogens often involves delivery of bacterial proteins called "effectors" into the host cell. The pneumonia-causing pathogen Legionella pneumophila delivers more than 330 effectors into the host cell via its type IVB Dot/Icm secretion system. The collective functions of these proteins is the establishment of a replicative niche from which Legionella can recruit cellular materials to grow while evading lysosomal fusion inhibiting its growth. Using a combination of structural, biochemical, and in vivo approaches, we show that one of these translocated effector proteins, Ceg4, is a phosphotyrosine phosphatase harboring a haloacid dehalogenase-hydrolase domain. Ceg4 could dephosphorylate a broad range of phosphotyrosine-containing peptides in vitro and attenuated activation of MAPKcontrolled pathways in both yeast and human cells. Our findings indicate that L. pneumophila's infectious program includes manipulation of phosphorylation cascades in key host pathways. The structural and functional features of the Ceg4 effector unraveled here, provide first insight into its function as a phosphotyrosine phosphatase, paving the way to further studies into L. pneumophila pathogenicity.Host colonization by Gram-negative pathogens often involves delivery of specific sets of bacterial proteins called "effectors" into the host cell by specialized secretion systems. Acting in concert, effectors secure the pathogen's survival and replication, via manipulation of host processes and the establishment of subcellular environments that favor the pathogen. Legionella phosphotyrosine phosphatase activity towards MAPKs 2 different pathogens. For example, plague and intestinal disease-causing Yersinia species encode less than ten effectors delivered by the type III secretion system (4) while the similar secretion system in Shigella spp. is involved in translocation of close to 30 effectors (5). Notably, sequence-related effectors are found in pathogens with very diverse invasion strategies suggesting that these effector families are involved in common host manipulation tactics.A causative agent of severe pneumonia in humans, the Legionella pneumophila genome encodes over 330 effector proteins, that are translocated via the Dot/Icm (defect in organelle trafficking/ intracellular multiplication), type IVB secretion system (e.g. (6-11), or see (12) for a recent review). Legionella's effectors account for more than 10% of its proteome (13) and represents the largest effector set known to the bacterial world. In their natural habitat of fresh water reservoirs Legionella spp. invade diverse amoebae by preventing formation of the phagolysosome (14) followed by modification of the newly co-opted compartment into an organelle ideal for intracellular replication of the bacteria, the Legionella containing vacuole (LCV) (15). The ability to apply the same invasion strategy to invade human alveolar macrophages raises the intriguing possibility that Legionella's effectors target host processes that are conser...

Legionella pneumophila effector Lem4 is a membrane-associated protein tyrosine phosphatase

“…Recently, the structure of another member of the HAD superfamily present in L. pneumophila, Ceg4, was reported (44). Like Lem4 (reported here), Ceg4 was also identified as a tyrosine phosphatase based on the structural information and was confirmed experimentally.…”

Legionnaires’ Disease: State of the Art Knowledge of Pathogenesis Mechanisms of Legionella

“…Indeed, L. pneumophila activates this pathway in a T4SS-dependent manner (117,118) by secreting five effectors that inhibit host translation and lead to MAPK activation, thus shaping the transcriptional response of the host cell (119). Another secreted effector, Ceg4, can modulate the phosphorylation state of eukaryotic MAPKs through its haloacid dehalogenase-like phosphatase domain (120).…”

Budowa IV systemu sekrecji Legionella pneumophilai jego znaczenie w patogenezie

“…Domena Qc może działać jako sensor, który umożliwia kompleksowi IcmR-IcmQ interakcję z krytycznym składnikiem systemu sekrecji, podczas gdy wewnątrzkomórkowy poziom NAD + jest wystarczający do zajścia reakcji wiązania. Niski wewnątrzkomórkowy poziom NAD + prowadzi w tym przypadku do zwiększonego tempa zużycia białka IcmQ, co może nastąpić w czasie wzrostu bakterii, w którym poziom innych białek związanych z wirulencją jest również obniżony [44]. Kompleks IcmR-IcmQ wiąże się z błonami zawierającymi kwaśne fosfolipidy [8].…”

“…Pięć efektorów L. pneumophila wydzielanych za pomocą IV systemu sekrecji hamuje translację białek gospodarza przez aktywację szlaku kinazy białkowej aktywowanej mitogenami (MAPK), modyfikując w ten sposób odpowiedź transkrypcyjną komórki gospodarza [76]. Fosfataza fosfotyrozynowa Ceg4 zawierająca domenę o aktywności dehalogenazy-hydrolazy halo-kwasowej defosforyluje wiele peptydów zawierających fosfotyrozynę i w ten sposób może osłabić aktywację szlaków kontrolowanych przez MAPK zarówno w komórkach drożdży, jak i ludzkich [44].…”