Abstract
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. (Mol Cell Biochem 268: 93–101, 2005)
Similar content being viewed by others
References
Ravdin JI: Amebiasis. Clinical Infec Dis 20: 1453–1466, 1995
Ravdin JI, Murphy CF, Guerrant RL, Long-Krug SA: Effect of calcium and phospholipase A antagonists on the cytopathogenicity of Entamoeba histolytica. J Infect Dis 152: 542–549, 1985
Vazquez J, Franco E, Reyes G, Meza I: Characterization of adhesion plates induced by the interaction of Entamoeba histolytica trophozoites with fibronectin. Cell Motil Cytoskeleton 32: 37–45, 1995
Perez E, Munoz ML, Ortega A: E. histolytica: Involvement of pp125FAK in collagen induced signal transduction. Exp Parasitol 82: 164–170, 1996
Ravdin JI, Moreau R, Sullivan JA, Petri WA Jr, Mandell GL: Relationship of free intracellular calcium to the cytolytic activity of E. histolytica. Infec Immun 56: 1505–1512, 1988
Diamond LS, Harlow DR, Cunnick, CC: A new medium for the axenic cultivation of E. histolytica and other Entamoeba. Trans Royal Soc Trop Med Hygiene 72: 431–432, 1978
Petri WA Jr, Smith RD, Schlesinger PH, Murphy CF, Ravdin JI: Isolation of the galactose binding lectin which mediates the in vitro adherence of E. histolytica. J Clinical Investigation 80: 1238–1244, 1987
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ: Protein measurement with the Folin phenol reagent. J Biol Chem 193: 265–275, 1951
Laemmli UK: Cleavage of structural proteins during the assembly of bacteriophage T4. Nature 227: 680–685, 1970
Petri WA, Schnaar RL: Identification of adhesins and receptors. Methods Enzymol 253: 98–104, 1995
Towbin H, Stachelin T, Gordan J: Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: Procedure and some applications. Proc National Acad Sci USA 76: 4350–4355, 1979
Kao JPY, Harootunian AT, Tsien RY: Photochemically generated cytosolic calcium pulses and their detection by Fluo-3. J Biol Chem 264: 8179–8184, 1989
Eberhard M, Erne P: Calcium binding to fluorescent calcium indicators: Calcium green, calcium orange and calcium crimson. Biochem Biophys Res Commun 180: 209–215, 1991
Sugimoti Y, Whiteman M, Cantley LC, Erikson RL: Evidence that RSV transforming gene product phosphorylates phosphatidylinositol and diacylglycerol. Proc National Acad Sci USA 81: 2117–2121, 1984
Leroy A, Bruyne GD, Mareel M, Nokkaew C, Bailey G, Nelis H: Contact-dependent transfer of the galactose-specific lectin of Entamoeba histolytica to the lateral surface of enterocytes in culture. Infect Immun 63: 4253–4260, 1995
Ravdin JI: Entamoeba histolytica: From adherence to enteropathy. J Infect Dis 159: 420–429, 1989
Bailey GB, Nudelman ED, Day DB, Harper CF, Gilmour JR: Specificity of glycosphingolipid recognition by E. histolytica trophozoites. Infec Immun 58: 43–47, 1990
Guillen N: Role of signaling and cytoskeletal rearrangements in the pathogenesis of Entamoeba histolytica. Trends Microbiol 4: 191–197, 1996
Olivier M: Modulation of host cell intracellular Ca2+. Parasitology Today 12: 145–150, 1996
Olivier M, Baimbridge KG, Reiner NE: Stimulus response coupling in monocytes infected with Leishmania. Attenuation of calcium transients is related to defective agonist-induced accumulation of inositol phosphates. J Immunol 148: 1188–1196, 1992
Turco SJ, Descoteaux A: The lipophosphoglycan of Leishmania parasites. Annu Rev Microbiol 46: 65–94, 1992
McGowan TA, Madesh M, Zhu Y, Wang L, Russo M, Deelman L, Henning R, Joseph S, Hajnoczky G, Sharma K: TGF-beta-induced Ca(2+) influx involves the type III IP (3) receptor and regulates actin cytoskeleton. Am J Ren Physiol 282: F910–F920, 2002
Banan A, Fields JZ, Farhadi A, Talmage DA, Zhang I, Keshavarzian A: The beta 1 isoform of protein kinase C mediates the protective effects of epidermal growth factor on the dynamic assembly of F–actin cytoskeleton and normalization of calcium homeostasis in human colonic cells. J Pharmacol Exp Ther 301: 852–866, 2002
Li C, Fultz ME, Parkash J, Rhoten WB, Wrigh GL: Ca2+ dependent actin remodeling in the A7r5 cells. J Muscle Res Cell Motil 22: 521–534, 2001
Exton JH, Taylor SJ, Blank JS, Bocckino SB: Regulation of phosphoinositide and phosphatidyl choline phospholipases by G proteins. In: Interactions among Cell Signaling Systems, Wiley, Chichester (Ciba Found. Symp. 164), pp. 36–49, 1992
Rasmussen H, Isales C, Ganesan S, Calle R, Zawalich W: Ca2+ cyclic AMP interactions in sustained cellular responses. In: Interactions among Cell Signaling Systems. Wiley, Chichester (Ciba Found. Symp. 164), 1992, pp. 98–112
Kramer RM: Structure, function and regulation of mammalian phospholipase A2. Adv Second Messenger Phosphoprotein Res 28: 81–89, 1993
Sanchez-Ramirez B, Escalante B, Rosales-Encina JL, Talamas-Rhana P: Role of prostaglandin E2 an amoebic liver abscess formation in hamsters. Prostaglandins C53(6): 411–421, 1997
Stenson WF, Zhang Z, Riehl T, Stanley Jr. SL: Amoebic infection in the human colon induces Cox-2. Infect Immun 69: 3382–3388, 2001
Beubler E, Bukare K, Rask-Madsen J: Significance of calcium for the prostaglandin E2 mediated secretory response to 5-hydroxytryptamine in small intestine of rats in vivo. Gastroenterology 90: 1972–1977, 1986
Wang W, Chadee K: E. histolytica alters arachidonic acid metabolism in macrophages in vitro and in vivo. Immunolog{y} 76: 242–250, 1992
Takai Y, Sasaki T, Matozaki T: Small GTP binding proteins. Physiol Rev 81: 154–188, 2001
Kyriakis JM, Avruch J: Mammalian mitogen-activated protein kinase signal transduction pathways activated by stress and inflammation. Physiol Rev 81: 808–859, 2001
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Rawal, S., Majumdar, S. & Vohra, H. Activation of MAPK Kinase pathway by Gal/GalNAc adherence lectin of E. histolytica: Gateway to host response. Mol Cell Biochem 268, 93–101 (2005). https://doi.org/10.1007/s11010-005-3698-4
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/s11010-005-3698-4