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Transcriptome analysis during the process of in vitro differentiation of Leishmania donovani using genomic microarrays

  • G. SRIVIDYA (a1), R. DUNCAN (a2), P. SHARMA (a1), B. V. S. RAJU (a1), H. L. NAKHASI (a2) and P. SALOTRA (a1)...
Abstract
SUMMARY

Leishmania donovani causes visceral disease (kala-azar), a major health problem throughout the tropics with 500 000 new cases every year. Leishmania differentiates from the promastigote to the amastigote form to establish infection in a mammalian host. To understand the process of differentiation, we assessed the global variation in gene expression in promastigotes, an intermediate stage of differentiation (PA24) and axenic amastigotes in culture using an L. donovani genomic microarray with 4224 clones printed in triplicate. During an intermediate stage of differentiation 24 h after shifting the promastigotes into amastigotes (PA24), there were 41 (∼1%) clones with expression ⩾2·0-fold higher than promastigotes, whereas in terminally differentiated amastigotes there were 130 (∼3%) such clones. Of particular interest were certain genes that exhibited a transient increase or decrease in expression at the PA24 stage. Kinases showed a transient increase, and surface molecules, PSA and amino acid permease, were prominent clones among those showing a brief decrease at the PA24 stage. The microarray results have been validated using Northern blots or RT-PCR. In summary, our results provide important clues about the genes involved in the differentiation process of L. donovani that may contribute to virulence.

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Corresponding author
*Corresponding author: Institute of Pathology (ICMR), Safdarjung Hospital Campus, New Delh-110029, India. Tel: +91 11 26166124. Fax: +91 11 26166124. E-mail: salotra@vsnl.com
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N. S. Akopyants , S. W. Clifton , J. Martin , D. Pape , T. Wylie , L. Li , J. C. Kissinger , D. S. Roos and S. M. Beverley (2001). A survey of the Leishmania major Friedlin strain V1 genome by shotgun sequencing: a resource for DNA microarrays and expression profiling. Molecular and Biochemical Parasitology 113, 337340.

N. S. Akopyants , R. S. Matlibs , E. N. Bukanova , M. R. Smeds , B. H. Brownstein , G. D. Stormo and S. M. Beverley (2004). Expression profiling using random genomic DNA microarray identifies differentially expressed genes associated with three major developmental stages of the protozoan parasite Leishmania major. Molecular and Biochemical Parasitology 136, 7186.

R. Almeida , B. J. Gilmartin , S. H. McCann , A. Norrish , A. C. Ivens , D. Lawson , M. P. Levick , D. F. Smith , S. D. Dyall , D. Vetrie , T. C. Freeman , R. M. Coulson , I. Sampaio , H. Schneider and J. M. Blackwell (2004). Expression profiling of the Leishmania life cycle: cDNA arrays identify developmentally regulated genes present but not annotated in the genome. Molecular and Biochemical Parasitology 136, 87100.

J. K. Beetham , J. E. Donelson and R. R. Dahlin (2003). Surface glycoprotein PSA (GP46) expression during short and long-term culture of Leishmania chagasi. Molecular and Biochemical Parasitology 131, 109117.

N. Boucher , Y. Wu , C. Dumas , M. Dube , D. Sereno , M. Breton and B. Papadopoulou (2002). A common mechanism of stage-regulated gene expression in Leishmania mediated by a conserved 3′-untranslated region element. Journal of Biological Chemistry 277, 1951119520.

S. Brandau , A. Dresel and J. Clos (2003). High constitutive levels of heat-shock proteins in human-pathogenic parasites of the genus Leishmania. The Biochemical Journal 310, 225232.

C. Cheadle , M. P. Vawter , W. J. Freed and K. G. Becker (2003). Analysis of microarray data using Z-score transformation. Journal of Molecular Diagnostics 5, 7381.

R. M. R. Coulson and D. F. Smith (1990). Isolation of genes showing increased or unique expression in the infective promastigotes of Leishmania major. Molecular and Biochemical Parasitology 40, 6376.

A. Debrabant , M. B. Joshi , P. F. Pimenta and D. M. Dwyer (2004). Generation of axenic amastigotes: their growth and cultural characteristics. International Journal for Parasitology 34, 205317.

P. Desjeux (2001). Worldwide increasing risk factors for leishmaniasis. Medical Microbiology and Immunology 190, 7779.

R. Duncan , P. Salotra , N. Goyal , N. Akopyants , S. M. Beverley and H. L. Nakhasi (2004). The application of gene expression microarray technology to kinetoplastid research. Current Molecular Medicine 4, 611621.

S. Dutoya , S. Gibert , G. Lemercier , X. Santarelli , D. Baltz , T. Baltz and N. Bakalara (2001). A novel C-terminal kinesin is essential for maintaining functional acidocalcisomes in Trypanosoma brucei. Journal of Biological Chemistry 276, 4911749124.

N. M. El-Sayed , P. Hegde , J. Quackenbush , S. E. Melville and J. E. Donelson (2000). The African trypanosome genome. International Journal for Parasitology 30, 329345.

M. Erdmann , A. Scholz , I. M. Melzer , C. Schmetz and M. Wiese (2006). Interacting protein kinases involved in regulation of flagellar length. Molecular Biology of the Cell 17, 20352045.

M. V. Geraldo , A. M. Silber , C. A. Pereira and S. R. Uliana (2005). Characterization of a developmentally regulated amino acid transporter gene from Leishmania amazonensis. FEMS Microbiology Letters 242, 275280.

N. Goyal , R. Duncan , A. Selvapandiyan , A. Debrabant , M. S. Baig and H. L. Nakhasi (2006). Cloning and characterization of angiotensin converting enzyme related dipeptidylcarboxypeptidase from Leishmania donovani. Molecular and Biochemical Parasitology 145, 147157.

R. T. Holzer , W. R. McMaster and J. D. Forney (2006). Expression profiling by whole genome interspecies microarray hybridization reveals differential gene expression in procyclic promastigotes, lesion derived amastigotes and axenic amastigotes in Leishmania mexicana. Molecular and Biochemical Parasitology 146, 198218.

K. Imai , T. Mimori , M. Kawai and H. Koga (2005). Microarray analysis of host gene-expression during intracellular nests formation of Trypanosoma cruzi amastigotes. Microbiology and Immunology 49, 623631.

A. C. Ivens , C. S. Peacock , E. A. Worthey , L. Murphy , G. Aggarwal , M. Berriman , E. Sisk , M. A. Rajandream , E. Adlem , R. Aert , A. Anupama , Z. Apostolou , P. Attipoe , N. Bason , C. Bauser , A. Beck , S. M. Beverley , G. Bianchettin , K. Borzym , G. Bothe , C. V. Bruschi , M. Collins , E. Cadag , L. Ciarloni , C. Clayton , R. M. Coulson , A. Cronin , A. K. Cruz , R. M. Davies , J. De Gaudenzi , D. E. Dobson , A. Duesterhoeft , G. Fazelina , N. Fosker , A. C. Frasch , A. Fraser , M. Fuchs , C. Gabel , A. Goble , A. Goffeau , D. Harris , C. Hertz-Fowler , H. Hilbert , D. Horn , Y. Huang , S. Klages , A. Knights , M. Kube , N. Larke , L. Litvin , A. Lord , T. Louie , M. Marra , D. Masuy , S. K. Matthews , S. Michaeli , J. C. Mottram , S. Muller-Auer , H. Munden , S. Nelson , H. Norbertczak , K. Oliver , S. O'neil , M. Pentony , T. M. Pohl , C. Price , B. Purnelle , M. A. Quail , E. Rabbinowitsch , R. Reinhardt , M. Rieger , J. Rinta , J. Robben , L. Robertson , J. C. Ruiz , S. Rutter , D. Saunders , M. Schafer , J. Schein , D. C. Schwartz , K. Seeger , A. Seyler , S. Sharp , H. Shin , D. Sivam , R. Squares , S. Squares , V. Tosato , C. Vogt , G. Volckaert , R. Wambutt , T. Warren , H. Wedler , J. Woodward , S. Zhou , W. Zimmermann , D. F. Smith , J. M. Blackwell , K. D. Stuart , B. Barrell and P. J. Myler (2005). The genome of the kinetoplastid parasite Leishmania major. Science 309, 436442.

M. Joshi , D. M. Dwyer and H. L. Nakhasi (1993). Cloning and characterization of differentially expressed genes from in vitro grown ‘amastigotes’ of Leishmania donovani. Molecular and Biochemical Parasitology 58, 345354.

P. J. Kolodziejski , A. Musial , J. S. Koo and N. T. Eissa (2002). Ubiquitination of inducible nitric oxide synthase is required for its degradation. Proceedings of the National Academy of Sciences, USA 99, 1231512320.

S. Krobitsch , S. Brandau , C. Hoyer , C. Schmetz , A. Hubel and J. Clos (1998). Leishmania donovani heat shock protein 100: Characterization and function in amastigote stage differentiation. Journal of Biological Chemistry 273, 64886494.

L. M. Lincoln , M. Ozaki , J. E. Donelson and J. K. Beetham (2004). Genetic complementation of Leishmania deficient in PSA (GP46) restores their resistance to lysis by complement. Molecular and Biochemical Parasitology 137, 185189.

T. Mitani , M. Terashima , H. Yoshimura , Y. Nariai and Y. Tanigawa (2005). TGF-beta 1 enhances degradation of IFN-gamma-induced iNOS protein via proteasomes in RAW 2647 cells. Nitric Oxide: Biology and Chemistry 13, 7887.

A. Piani , T. Ilg , A. G. Elefanty , J. Curtis and E. Handman (1999). Leishmania major proteophosphoglycan is expressed by amastigotes and has an immunomodulatory effect on macrophage function. Microbes and Infection 1, 589599.

C. O. Rodrigues , D. A. Scott and R. Docampo (1999). Presence of a vacuolar H+ pyrophosphatase in promastigotes of Leishmania donovani and its localization to a different compartment from the vacuolar H+ ATPase. The Biochemical Journal 340, 759766.

P. Salotra , R. C. Duncan , R. Singh , B. V. Subba Raju , G. Sreenivas and H. L. Nakhasi (2006). Upregulation of surface proteins in Leishmania donovani isolated from patients of post-kala-azar dermal leishmaniasis. Microbes and Infection 8, 637644.

P. Salotra , G. Sreenivas , G. P. Pogue , N. Lee , H. L. Nakhasi , V. Ramesh and N. S. Negi (2001). Development of a species-specific PCR assay for detection of Leishmania donovani in clinical samples from patients with kala-azar and post-kala-azar dermal leishmaniasis. Journal of Clinical Microbiology 39, 849854.

A. Saxena , E. A. Worthey , S. Yan , A. Leland , K. D. Stuart and P. J. Myler (2003). Evaluation of differential gene expression in Leishmania major Friedlin procyclics and metacyclics using DNA microarray analysis. Molecular and Biochemical Parasitology 129, 103114.

G. Sreenivas , R. Singh , A. Selvapandian , N. S. Negi , H. L. Nakhasi and P. Salotra (2004). Arbitrary-primed PCR for genomic fingerprinting and identification of differentially regulated genes in Indian isolates of Leishmania donovani. Experimental Parasitology 106, 110118.

J. Walker , J. J. Vasquez , M. A. Gomez , J. Drummelsmith , R. Burchmore , I. Girard and M. Ouellette (2006). Identification of developmentally regulated proteins in Leishmania panamensis by proteome profiling of promastigotes and axenic amastigotes. Molecular and Biochemical Parasitology 147, 6473.

M. Wiese (1998). A mitogen-activated protein (MAP) kinase homologue of Leishmania mexicana is essential for parasite survival in the infected host. The EMBO Journal 17, 26192628.

M. Wiese , D. D. Kuhn and C. G. Grünfelder (2003). Protein kinase involved in flagellar-length control. Eukaryotic Cell 2, 769777.

Y. Wu , Y. El-Fakhry , D. Sereno , S. Tamar and B. Papadopoulou (2000). A new developmentally regulated gene family in Leishmania amastigotes encoding a homolog of amastin surface proteins. Molecular and Biochemical Parasitology 110, 345357. www.geneDB.org

M. Yoshida and Y. Xia (2003). Heat shock protein 90 as an endogenous protein enhancer of inducible nitric-oxide synthase. Journal of Biological Chemistry 278, 3695336958.

J. A. Young , Q. L. Fivelman , P. L. Blair , P. de la Vega , K. G. Le Roch , Y. Zhou , D. J. Carucci , D. A. Baker and E. A. Winzeler (2005). The Plasmodium falciparum sexual development transcriptome: a microarray analysis using ontology-based pattern identification. Molecular and Biochemical Parasitology 143, 6779.

F. B. Zamora-Veyl , M. Kroemer , D. Zander and J. Clos (2005). Stage-specific expression of the mitochondrial co-chaperonin of Leishmania donovani CPN10. Kinetoplastid Biology and Disease DOI: 10.1186/1475-9292-4-3.

W. W. Zhang and G. Matlashewski (1997). Loss of virulence in Leishmania donovani deficient in an amastigote-specific protein A2. Proceedings of the National Academy of Sciences, USA 94, 88078811.

D. Zilberstein and M. Shapira (1994). The role of pH and temperature in the development of Leishmania parasites. Annual Review of Microbiology 48, 449470.

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Parasitology
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