Identifying human targets for Legionella pneumophila effector kinases by combining human protein arrays with a non-radioactive ATP labeling technique

FARE Award Winner

Authors

• P LEE
• MP Machner

Abstract

Protein phosphorylation is the most abundant post-translational modification within living cells. Hundreds of kinases and thousands of kinase substrates have been predicted in human cells, highlighting the importance and complexity of phosphorylation networks. During infection, many bacterial pathogens interfere with cellular signaling pathways in order to avoid clearance by the host. Legionella pneumophila causes a form of bacterial pneumonia called Legionnaires’ disease. Upon phagocytosis by alveolar macrophages, L. pneumophila uses a type IV secretion system (T4SS) to translocate more than 300 effector proteins into the host. How these effectors alter cellular functions in order to create conditions favorable for L. pneumophila replication has remained largely unclear. Using a bioinformatics approach, we discovered five novel L. pneumophila effectors with putative protein kinase domains, three of which we already validated using a non-radioactive ATP analog labeling technique. In order to identify host targets phosphorylated by these effector kinases, we combined the ATP analog labeling technique with a protein array containing more than 9,300 human proteins. Using this approach, we discovered that the L. pneumophila effector kinase 6, LegK6, specifically phosphorylates several human proteins. Interestingly, pathway analysis revealed that host proteins involved in cytoskeleton rearrangement are the main targets of LegK6. Since L. pneumophila alters host endocytosis/phagocytosis pathway to create a unique legionella-containing vesicle for its intracellular replication, our future studies are aimed at investigating, in a cellular context, how the downstream signaling events that are manipulated by L. pneumophila effector kinases promote bacterial infectivity.

Scientific Focus Area: Microbiology and Infectious Diseases

This page was last updated on Friday, March 26, 2021