Comparative Proteomic Profiling of Leishmania tropica: Investiga¬tion of a Case Infected with Simultaneous Cutaneous and Viscero¬tropic Leishmaniasis by 2-Dimentional Electrophoresis and Mass Spectrometry
AbstractBackground: Viscerotropic leishmaniasis caused by Leishmania tropica poses a significant problem in the diagnosis and treatment management. Since differential gene expression is more important in outcome of the infection, we employed proteomic approach to identify potential proteins involved in visceralization of L. tropica.Methods: The proteomes profiling of L. tropica isolated from cutaneous and visceral tissues of one host were compared by 2-DE/MS proteomics study. Moreover, the transcript level of some identified proteins was confirmed using real-time RT-PCR.Results: Of the 700 protein spots that were detected reproducibly on each gel, 135 were found to be differentially expressed (P≤ 0.05). Most of responsive proteins in visceral isolate changed in less abundant compared to cutaneous isolate. Among differentially expressed proteins, 56 proteins were confidently identified and classified according to the biological process. The largest groups consist of proteins involved in carbohydrate metabolism and protein synthesis. Most of the identified proteins, which implicated in energy metabolism, cell signaling and virulence were down-regulated, whereas some proteins that have a role in protein folding, antioxidant defense and proteolysis were up-regulated in visceral form. Moreover, the transcript level of some identified proteins such as co-chaperon was confirmed using real-time RT-PCR.Conclusion: L. tropica probably uses different mechanisms for survival and multiplication in viscera to establish viscerotropic leishmaniasis. The current study provides some clues into the mechanisms underlying the dissemination of L. tropica.
Alvar J, Vélez ID, Bern C, Herrero M, Desjeux P, Cano J, et al. Leishmaniasis worldwide and global estimates of its incidence. PloS One.2012; 7, e35671.
Reithinger R, Dujardin JC, Louzir H, Pirmez C, Alexander B, Brooker S, et al. Cutaneous leishmaniasis. Lancet Infect Dis. 2007; 7: 581-96.
Foucher AL, Papadopoulou B, Ouellette M. Prefractionation by Digitonin Extraction Increases Representation of the Cytosolic and Intracellular Proteome of Leishmania infantum. J Proteome Res. 2006; 5: 1741-50.
Hajjaran H, Mohebali M, Mamishi S, Vasighe F, Oshaghi MA, Naddaf SR, et al. Molecular Identification and Polymorphism Determina-tion of Cutaneous and Visceral Leishmaniasis Agents Isolated from Human and Animal Hosts in Iran. BioMed Research International. 2013; ID 789326,7pages.
Control of the leishmaniases: report of a meeting of the WHO Expert Committee on the Control of Leishmaniases, Geneva, 22-26 March 2010. WHO Technical Report No. 949.
Badirzadeh A, Mohebali M, Ghasemian M, Amini H, Zarei Z, Akhoundi B, et al. Cutaneous and post kala-azar dermal leishmaniasis caused by Leishmania infantum in endemic areas of visceral leishmaniasis, northwestern Iran 2002‐2011: a case series. Pathog Glob Health. 2013; 107: 194-7.
Magill A J, Grogl M, Gasser Jr R A, Sun W, Oster CN. Visceral infection caused by Leishmania tropica in veterans of Operation Desert Storm. N Engl J Med. 1993; 328: 1383-87.
Zijlstra EE, Musa AM, Khalil EA, el- Hassan IM, el-Hassan AM. Post-kala-azar dermal leishmaniasis. Lancet Infect Dis.2003; 3: 87-98.
Dillon DC, Day CH, Whittle JA, Magill AJ, Reed SG. Characterization of a Leishmania tropica antigen that detects immune responses in Desert Storm viscerotropic leishmaniasis patients. Proc Natl Acad Sci USA.1995; 7981-85.
Fasel N, Acestor N, Fadili-Kundig A, Gonzalez IS. The Leishmania proteom. In: Leishmania After the Genome. Caiser Academic Press: 2008, p 55-65.
Kazemi-Rad E, Mohebali M, Khadem Erfan MB, Saffari M, Raoofian R, Hajjaran H, et al. Identification of antimony resistance markers in Leishmania tropica field isolates through a cDNA-AFLP approach. Exp Parasitol. 2013; 135: 344-49.
Paape D, Aebischer T. Contribution of proteomics of Leishmania spp. to the understanding of differentiation, drug resistance mechanisms, vaccine and drug development. J Proteomics.2011; 74: 1614-24.
Mohebali M, Malmasi A, Hajjaran H, Jamshidi S, Akhoundi B, Rezaei M, et al. Disseminated leishmaniasis caused by Leishmania tropica in a puppy from Karaj, Central Iran. Iran J Parasi-tol. 2011; 6: 69-73.
Hajjaran H, Mohebali M, Teimouri A, Oshaghi M A, Mirjalali H et al. Identification and phy-logenetic relationship of Iranian strains of var-ious Leishmania species isolated from cuta-neous and visceral cases of leishmaniasis based on N-acetyl glucosamine-1-phosphate trans-ferase gene. Infect Genet Evol. 2014; 26: 203-12.
Bazargani MM, Sarhadi E, Bushehri AA, Matros A, Mock HP, Naghavi MR, et al. A proteomics view on the role of drought-induced senescence and oxidative stress defense in enhanced stem reserves remobiliza-tion in wheat. J Proteomics. 2011;74: 1959-73.
Blum H, Beier H, Gross H J. Improved silver staining of plant proteins, RNA and DNA in polyacrylamide gels. Electrophoresis. 1987; 8: 93-9.
Neuhoff V, Arold N, Taube D, Ehrhardt W. Improved staining of proteins in polyacrylamide gels including isoelectric focusing gels with clear background at nanogram sensitivity using Coomassie Brilliant Blue G-250 and R-250. Electrophoresis. 1988; 9: 255-62
Daneshvar H, Wyllie S, Phillips S, Hagan P, Burchmore R. Comparative proteomics profiling of a gentamicin-attenuated Leishmania infantum cell line identifies key changes in parasite thiol-redox metabolism. J Proteomics. 2012; 75: 1463-71.
Livak K J, Schmittgen TD. Analysis of relative gene expression data using real- time quantitative PCR and the 2 (Delta Delta C(T)) method. Methods. 2001; 25: 402-8.
Opperdoes FR, Coombs GH. Metabolism of Leishmania: proven and predicted. Trends Parasitol. 2007; 23 : 149-58.
Kushawaha PK, Gupta R, Tripathi CDP, Khare P, Jaiswal AK, Sundar S, et al. Leishmania donovani Triose Phosphate Isomerase: A Potential Vaccine Target against Visceral Leishmaniasis. PLoS One. 2012; 7 : e45766.
Kramer S. Developmental regulation of gene expression in the absence of transcriptional control: the case of kinetoplastids. Mol Biochem Parasitol. 2012; 181: 61-72.
Condeelis J. Elongation factor 1 alpha, translation and the cytoskeleton. Trends Biochem Sci. 1995; 20 : 169-70.
Nandan D, Yi T, Lopez M, Lai C, Reiner NE. Leishmania EF-1alpha activates the Src homology 2 domain containing tyrosine phosphatase SHP-1 leading to macrophage deactivation. J Biol Chem.2002; 277 : 50190-7.
Kamoun-Essghaier S, Guizani I, Strub J. M., Van Dorsselaer A, Mabrouk K, Ouelhazi L, Dellagi K. Proteomic approach for characterization of immunodominant membrane-associated 30-to 36-kilodalton fraction antigens of Leishmania infantum promastigotes, reacting with sera from mediterranean visceral leishmaniasis patients. Clin Diagn Lab Immunol.2005; 12, 310-320.
Chin D, Means A R. Calmodulin: a prototypical calcium sensor. Trends Cell Biol.2000; 10: 322-8.
Lu HG, Zhong L, Chang KP, Docampo R. Intracellular Ca2+ pool content and signaling and expression of a calcium pump are linked to virulence in Leishmania mexicana amazonesis amastigotes. J Biol Chem.1997; 272: 9464-73.
Seaman MN, Marcusson EG, Cereghino JL, Emr S D. Endosome to Golgi retrieval of the vacuolar protein sorting receptor, Vps10p,
requires the function of the VPS29, VPS30, and VPS35 gene products. J Cell Biol.1997; 137, 79-92.
Besteiro S, Williams RA, Morrison LS, Coombs GH, Mottram JC. Endosome sorting and autophagy are essential for differentiation and virulence of Leishmania major. J Biol Chem. 2006; 281, 11384-96.
Shang F, Taylor A. Ubiquitin–proteasome pathway and cellular responses to oxidative stress. Free Radic Biol Med. 2011; 51, 5-16.
Cheng L, Watt R, Piper P. Polyubiquitin gene expression contributes to oxidative stress resistance in respiratory yeast (Saccharomyces cerevisiae). Mol Gen Genet.1994; 243, 358-62.
Kazemi-Rad E, Mohebali M, Khadem-Erfan MB, Hajjaran H, Hadighi R, Khamesipour A, et al. Overexpression of Ubiquitin and Amino Acid Permease Genes in Association with Antimony Resistance in Leishmania tropica Field Isolates. Korean J Parasitol. 2013; 51, 413-9.
Nogoceke E, Gommel DU, Kieß M, Kalisz HM, Flohé L. A unique cascade of oxidoreductases catalyses trypanothione-mediated peroxide metabolism in Crithidia fasciculata. Biol Chem. 1997; 378, 827-36.
Castro H, Sousa C, Novais M, Santos M, Budde H, Cordeiro-da-Silva A, et al. Two linked genes of Leishmania infantum encode tryparedoxins localised to cytosol and mitochondrion. Mol Biochem Parasitol. 2004; 136, 137-47.
Rhee SG, Woo HA. Multiple functions of peroxiredoxins: peroxidases, sensors and regulators of the intracellular messenger H2O2, and protein chaperones. Antioxid. Redox Signal. 2011; 15, 781-94.
Lin YC, Hsu JY, Chiang SC, Lee ST. Distinct overexpression of cytosolic and mitochondrial tryparedoxin peroxidases results in preferential detoxification of different oxidants in arsenite-resistant Leishmania amazonensis with and without DNA amplification. Mol Biochem Parasitol. 2005;142, 66-75.
Caplan AJ. What is a co-chaperone? Cell Stress Chaperones. 2003; 8, 105-7.
Mayer MP, Bukau B. Hsp70 chaperones: cellular functions and molecular mechanism. Cell Mol Life Sci. 2005; 62, 670-84.
Bellmann K, Jäättelä M, Wissing D, Burkart V. Kolb, H. Heat shock protein hsp70 overexpression confers resistance against nitric oxide. FEBS Lett. 1996; 391, 185-88.
Chong KY, Lai CC, Lille S, Chang C, Su C Y. Stable overexpression of the constitutive form of heat shock protein 70 confers oxidative protection. J Mol Cell Cardiol. 1998; 30, 599-608.
Ono S. Mechanism of Depolymerization and Severing of Actin Filaments and Its Significance in Cytoskeletal Dynamics. Int Rev Cytol. 2007; 258, 1-82.
Tammana TVS, Sahasrabuddhe A A, Mitra K, Bajpai VK, Gupta CM. Actin-depolymerizing factor, ADF/cofilin, is essentially required in assembly of Leishmania flagellum. Mol Microbiol. 2008; 70, 837-52.
Hiam A, Sebastien D, George B, Arlette F, Kalil J, Le Pape P. Microtubule target for new antileishmanial drugs based on ethyl 3-haloacetamidobenzoates. J Enzyme Inhib Med Chem. 2006; 21, 305- 12.
Theinert SM, Basu R, Forgber M, Roy S, Sundar S, Walden P. Identification of New Antigens in Visceral Leishmaniasis by Expression Cloning and Immunoblotting with Sera of Kala-Azar Patients from Bihar, India. Infect Immun. 2005;73, 7018-21.
Yan H, Tsai M. Nucleoside monophosphate kinases: structure, mechanism, and substrate specificity. Adv Enzymol Relat Areas Mol Biol. 1999; 73, 103-34.
Villa H, Pérez-Pertejo Y, García-Estrada C, Reguera RM, Requena JM, Tekwani BL, et al. Molecular and functional characterization of adenylate kinase 2 gene from Leishmania donovani. Eur J Biochem.2003; 270, 4339-47.
Feliciano PR, Cordeiro AT, Costa-Filho AJ, Nonato MC. Cloning, expression, purification, and characterization of Leishmania major dihydroorotate dehydrogenase. Protein Expr Purif. 2006; 48, 98-103.
Cuervo P, de Jesus JB, Junqueira M, Mendonca-Lima L, Gonzalez LJ, Betancourt L, et al. Proteome analysis of Leishmania (Viannia) braziliensis by two-dimensional gel ele-ctrophoresis and mass spectrometry. Mol Biochem Parasitol.2007; 154, 6-21.
Kilmartin JV. Sfi1p has conserved centrin-binding sites and an essential function in budding yeast spindle pole body duplication. J Cell Biol. 2003; 162, 1211-21.
Selvapandiyan A, Debrabant A, Duncan R, Muller J, Salotra P, Sreenivas G, et al. Centrin gene disruption impairs stage-specific basal body duplication and cell cycle progression in Leishmania. J Biol Chem. 2004; 279, 25703-10.
Leipe DD, Wolf YI, Koonin EV, Aravind L. Classification and evolution of P-loop GTPases and related ATPases. J Mol Biol. 2002; 317, 41-72.
Saito TT, Lui DY, Kim HM, Meyer K, Colaiácovo MP. Interplay between structure-specific endonucleases for crossover control during Caenorhabditis elegans meiosis. PLoS Genet. 2013; 9, e1003586.