Identification of a Gal/GalNAc Lectin in the Protozoan Hartmannella vermiformis as a Potential Receptor for Attachment and Invasion by the Legionnaires' Disease Bacterium

Abstract

The Legionnaire's disease bacterium, Legionella pneumophila, is a facultative intracellular pathogen which invades and replicates within two evolutionarily distant hosts, free-living protozoa and mammalian cells. Invasion and intracellular replication within protozoa are thought to be major factors in the transmission of Legionnaire's disease. Although attachment and invasion of human macrophages by L. pneumophila is mediated in part by the complement receptors CR1 and CR3, the protozoan receptor involved in bacterial attachment and invasion has not been identified. To define the molecular events involved in invasion of protozoa by L. pneumophila, we examined the role of protein tyrosine phosphorylation of the protozoan host Hartmannella vermiformis upon attachment and invasion by L. pneumophila. Bacterial attachment and invasion were associated with a time-dependent tyrosine dephosphorylation of multiple host cell proteins. This host cell response was highly specific for live L. pneumophila, required contact with viable bacteria, and was completely reversible following washing off the bacteria from the host cell surface. Tyrosine dephosphorylation of host proteins was blocked by a tyrosine phosphatase inhibitor but not by tyrosine kinase inhibitors. One of the tyrosine dephosphorylated proteins was identified as the 170-kD galactose/N-acetylgalactosamine–inhibitable lectin (Gal/GalNAc) using immunoprecipitation and immunoblotting by antibodies generated against the Gal/GalNAc lectin of the protozoan Entamoeba histolytica. This Gal/GalNAc–inhibitable lectin has been shown previously to mediate adherence of E. histolytica to mammalian epithelial cells. Uptake of L. pneumophila by H. vermiformis was specifically inhibited by two monovalent sugars, Gal and GalNAc, and by mABs generated against the 170-kD lectin of E. histolytica. Interestingly, inhibition of invasion by Gal and GalNAc was associated with inhibition of bacterial-induced tyrosine dephosphorylation of H. vermiformis proteins. High stringency DNA hybridization confirmed the presence of the 170-kD lectin gene in H. vermiformis. We conclude that attachment of L. pneumophila to the H. vermiformis 170-kD lectin is required for invasion and is associated with tyrosine dephosphorylation of the Gal lectin and other host proteins. This is the first demonstration of a potential receptor used by L. pneumophila to invade protozoa.

Legionella: from environmental habitats to disease pathology, detection and control

“…A type IV pilus, designated the competence and adherence-associated pilus (CAP), is involved in the initial adherence of Legionella to protozoan cells and may provide Legionella with a selective advantage in adhering to surfaces and bio®lms in the environment (Stone and Abu Kwaik, 1998). Hartmannella has been shown to have a lectin receptor that is involved in the attachment and invasion of Legionella (Venkataraman et al, 1997). Synthesis of Hartmannella vermiformis proteins but not of Acanthamoeba polyphaga proteins is required for the uptake of L. pneumophila (Harb et al, 1998).…”

“…Invasion and intracellular replication of Legionella pneumophila within protozoa in the environment plays a major role in the transmission of Legionnaires' disease (Harb et al, 1998). After intracellular replication within protozoa, L. pneumophila exhibits resistance to conditions of stress, including high temperature, acidity and biocides, which may contribute to its environmental persistence (Abu Kwaik et al, 1997). L. pneumophila-infected H. vermiformis organisms are more pathogenic than an equivalent number of free living L. pneumophila cells (Brieland et al, 1997).…”

Flagellum of Legionella pneumophila Positively Affects the Early Phase of Infection of Eukaryotic Host Cells

“…Uptake by Hartmannella is accomplished by a microfilamentindependent mechanism that is sensitive to methylamine, which is an inhibitor of receptor-mediated endocytosis (28). So far, one receptor of Hartmanella vermiformis, a Gal/GalNAc lectin, could be identified (46). Attachment of L. pneumophila to this lectin results in tyrosine dephosphorylation of multiple host cell proteins.…”

Lectins and Tetrahymena – A review

“…This can help the engulfment and use as nourishment of the prey, bacteria for the predator [63], Tetrahymena. It was demonstrated earlier that amoebae are using galactose-binding lectins for attaching Legionella pneumophila [64,65] as well as Acanthamoeba castellanii, which uses mannose-binding lectin for phagocytizing yeast [66]. Marine planktonic dinoflagellates, Oxyrrhis marina and others, use also a mannose-binding lectin to recognize and attach phytoplanktonic preys [66].…”