Activation of MAPK kinase pathway by Gal/GalNAc adherence lectin of E. histolytica: gateway to host response.

Amoebiasis caused by the protozoan parasite Entamoeba histolytica is one of the leading parasitic causes of morbidity and mortality in the developing countries. Among the variety of virulence factors, an adherence lectin (Gal/GalNAc, 260 kDa) has been known to mediate colonization and subsequent host responses. It is a major cell surface antigen which is universally recognized by the immune sera of patients with amoebic liver abscess (ALA). The role of this lectin in cytolysis and phagocytosis of human colonic mucin glycoproteins has also been established. The objective of the present study was to elucidate the signal transduction events induced in response to Entamoeba histolytica derived Gal/GalNAc lectin in the target epithelial cells. We have attempted to define a pathway in target cells that could link this immunodominant antigen to a known biological pathway for target cell activation and triggering of subsequent disease pathology/parasite survival. Lectin stimulated cells showed immediate rise in (Ca2+)i concentration corresponding to 1517.31+/-16.3 nM (approximately) at 0-2 min. The intracellular calcium also extruded from the cells as was measured by increase in calcium green-1 fluorescence. Expression of several protein kinases was checked by western blotting to delineate the signaling pathway. Results showed that the expression of PLA2, PI3K, Ras p21, Ras GAP, ERK-MAPK, p38MAPK and PKC was significantly increased. Expression of Raf-1 and MEK-1 was also found to be significant, as determined by intensity analysis. Overall, it indicated activation of MAPKinase pathway which is implicated in a variety of cellular functions. On the basis of our observations it can be stated that there is a calcium mediated activation of PKC in target cells, by lectin, which inturn activates cyclic nucleotides and other protein kinases. These protein kinases further phosphorylated downstream signals in a sequential manner, thus leading to the activation of MAPKinase cascade. Activation of MAPK cascade, in our studies, is implicated in a variety of physiological cellular functions including apoptosis, proliferation, cytoskeleton rearrangements and permeability changes. However, future screening of the genes responsible for the transcription and translation of new proteins and their biological functions in response to lectin stimulation will prove useful in understanding this host-parasite relationship.