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Quantitative assessment of sense and antisense transcripts from genes involved in antigenic variation (vsp genes) and encystation (cwp 1 gene) of Giardia lamblia clone GS/M-83-H7

Published online by Cambridge University Press:  02 December 2004

N. von ALLMEN ,
M. BIENZ ,
A. HEMPHILL  and
N. MÜLLER
N. von ALLMEN
Affiliation:
Institute of Parasitology, University of Berne, Länggass-Strasse 122, CH-3012 Berne, Switzerland
M. BIENZ
Affiliation:
Hematology, Inselspital (University Hospital), University of Berne, Switzerland
A. HEMPHILL
Affiliation:
Institute of Parasitology, University of Berne, Länggass-Strasse 122, CH-3012 Berne, Switzerland
N. MÜLLER
Affiliation:
Institute of Parasitology, University of Berne, Länggass-Strasse 122, CH-3012 Berne, Switzerland
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Abstract

Antigenic variation of the intestinal protozoan parasite Giardia lamblia is caused by an exchange of the parasite's variant surface protein (VSP) coat. Many investigations on antigenic variation were performed with G. lamblia clone GS/M-83-H7 which produces surface antigen VSP H7. To generate novel information on giardial vsp gene transcription, vsp RNA levels were assessed by quantitative reverse transcription-(RT)-PCR in both axenic VSP H7-type trophozoites and subvariants obtained after negative selection of GS/M-83-H7 trophozoites by treatment with a cytotoxic, VSP H7-specific monoclonal antibody. Our investigation was not restricted to the assessment of the sense vsp transcript levels but also included an approach aimed at the detection of complementary antisense vsp transcripts within the two trophozoite populations. We found that sense vsp H7 RNA predominated in VSP H7-type trophozoites while sense RNA from only one (vsp IVg) of 8 subvariant vsp genes totally analysed predominated in subvariant-type trophozoites. Interestingly, the two trophozoite populations exhibited a similar relative distribution regarding the vsp H7 and vsp IVg antisense RNA molecules. An analogous sense versus antisense RNA pattern was also observed when the transcripts of gene cwp 1 (encoding cyst wall protein 1) were investigated. Here, both types of RNA molecules only appeared after cwp 1 had been induced through in vitro encystation of the parasite. These findings for the first time demonstrated that giardial antisense RNA production did not occur in a constitutive manner but was directly linked to complementary sense RNA production after activation of the respective gene systems.

Keywords

Giardia lambliaantigenic variationencystationtranscriptionantisense RNA
Type
Research Article
Information
Parasitology , Volume 130 , Issue 4 , April 2005 , pp. 389 - 396
Copyright
2004 Cambridge University Press

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References

REFERENCES

AGGARWAL, A., MERRITT, J. W. & NASH, T. E. ( 1989). Cysteine-rich variant surface proteins of Giardia lamblia. Molecular and Biochemical Parasitology 32, 39–48.CrossRefGoogle Scholar
BIENZ, M., SILES-LUCAS, M., WITTWER, P. & MÜLLER, N. ( 2001). Vsp gene expression by Giardia lamblia clone GS/M-83-H7 during antigenic variation in vivo and in vitro. Infection and Immunity 69, 5278–5285.CrossRefGoogle Scholar
CHENG, N., UPCROFT, J. A. & UPCROFT, P. ( 1996). A new cysteine-rich protein-encoding gene family in Giardia duodenalis. Gene 169, 33–38.Google Scholar
DAVIS-HAYMAN, S. R., HAYMAN, J. R. & NASH, T. E. ( 2003). Encystation-specific regulation of the cyst wall protein 2 gene in Giardia lamblia by multiple cis-acting elements. International Journal for Parasitology 33, 1005–1012.CrossRefGoogle Scholar
ELMENDORF, H. G., SINGER, S. M. & NASH, T. E. ( 2001). The abundance of sterile transcripts in Giardia lamblia. Nucleic Acids Research 29, 4674–4683.CrossRefGoogle Scholar
GOTTSTEIN, B., HARRIMAN, G. R., CONRAD, J. T. & NASH, T. E. ( 1990). Antigenic variation in Giardia lamblia: Cellular and humoral immune response in a mouse model. Parasite Immunology 12, 659–673.CrossRefGoogle Scholar
HEHL, A. B., MARTI, M. & KÖHLER, P. ( 2000). Stage-specific expression and targeting of cyst wall protein-green fluorescent protein chimeras in Giardia. Molecular Biology of the Cell 11, 1789–1800.CrossRefGoogle Scholar
KANE, A. V., WARD, H. D., KEUSCH, G. T. & PEREIRA, M. E. ( 1991). In vitro encystation of Giardia lamblia: large-scale production of in vitro cysts and strain and clone differences in encystation efficiency. International Journal for Parasitology 77, 974–981.CrossRefGoogle Scholar
KEISTER, D. B. ( 1983). Axenic culture of Giardia lamblia in TYI-S-33 medium supplemented with bile. Transactions of the Royal Society of Tropical Medicine and Hygiene 77, 487–488.CrossRefGoogle Scholar
LUJAN, H. D., MOWATT, M. R., CONRAD, J. T., BOWERS, B. & NASH, T. E. ( 1995). Identification of a novel Giardia lamblia cyst wall protein with leucine-rich repeats. Implications for secretory granule formation and protein assembly into the cyst wall. Journal of Biological Chemistry 270, 29307–29313.Google Scholar
LUJAN, H. D. & TOUZ, M. C. ( 2003). Protein trafficking in Giardia lamblia. Cellular Microbiology 5, 427–434.CrossRefGoogle Scholar
MARTI, M., REGOS, A., LI, Y., SCHRANER, E. M., WILD, P., MÜLLER, N., KNOPF, L. G. & HEHL, A. B. ( 2003). An ancestral secretory apparatus in the protozoan parasite Giardia intestinalis. Journal of Biological Chemistry 278, 24837–24848.CrossRefGoogle Scholar
MOWATT, M. R., LUJAN, H. D., COTTEN, D. B., BOWERS, B., YEE, J., NASH, T. E. & STIBBS, H. H. ( 1995). Developmentally regulated expression of a Giardia lamblia cyst wall protein gene. Molecular Microbiology 15, 955–963.CrossRefGoogle Scholar
MÜLLER, N. & GOTTSTEIN, B. ( 1998). Antigenic variation and the murine immune response to Giardia lamblia. International Journal for Parasitology 28, 1829–1839.CrossRefGoogle Scholar
NASH, T. E. ( 2002). Surface antigenic variation in Giardia lamblia. Molecular Microbiology 45, 585–590.CrossRefGoogle Scholar
NASH, T. E., CONRAD, J. T. & MOWATT, M. R. ( 1995). Giardia lamblia: identification and characterization of a variant-specific surface protein gene family. Journal of Eukaryotic Microbiology 42, 604–619.CrossRefGoogle Scholar
NASH, T. E., LUJAN, H. T., MOWATT, M. R. & CONRAD, J. T. ( 2001). Variant-specific surface protein switching in Giardia lamblia. Infection and Immunity 69, 1922–1923.CrossRefGoogle Scholar
NASH, T. E. & MOWATT, M. R. ( 1992). Characterization of a Giardia lamblia variant-specific surface protein (VSP) gene from isolate GS/M and estimation of the VSP gene repertoire size. Molecular and Biochemical Parasitology 51, 219–228.CrossRefGoogle Scholar
RIRIE, K. M., RASMUSSEN, R. P. & WITTWER, C. T. ( 1997). Product differentiation by analysis of DNA melting curve during the polymerase chain reaction. Analytical Biochemistry 245, 154–160.CrossRefGoogle Scholar
SAMBROOK, J. & RUSSEL, D. W. ( 2001). Molecular Cloning: A Laboratory Manual, 3rd Edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.
SUN, C. H., PALM, D., McARTHUR, A. G., SVÄRD, S. G. & GILLIN, F. D. ( 2002). A novel Myb-related protein involved in transcriptional activation of encystation genes in Giardia lamblia. Molecular Microbiology 46, 971–984.CrossRefGoogle Scholar
TIJSTERMAN, M. & PLASTERK, R. A. H. ( 2004). Dicer at RISC: The mechanism of RNAi. Cell 117, 1–4.Google Scholar
TOUZ, M. C., GOTIG, N., NASH, T. E. & LUJAN, H. D. ( 2002). Identification and characterization of a novel secretory granule calcium-binding protein from the early branching eukaryote Giardia lamblia. Journal of Biological Chemistry 52, 50557–50563.CrossRefGoogle Scholar
ULLU, E., TSCHUDI, C. & CHKRABORTX, T. ( 2004). RNA interference in protozoan parasites. Cellular Microbiology 6, 509–519.CrossRefGoogle Scholar
UPCROFT, P., CHENG, N. & UPCROFT, J. A. ( 1997). Telomeric organization of a variable and inducible toxin gene family in the ancient eukaryote Giardia duodenalis. Genome Research 7, 37–46.CrossRefGoogle Scholar
von ALLMEN, N., BIENZ, M., HEMPHILL, A. & MÜLLER, N. ( 2004). Experimental infections of neonatal mice with cysts of Giardia lamblia clone GS/M-83-H7 are associated with an antigenic reset of the parasite. Infection and Immunity 72, 4763–4771.CrossRefGoogle Scholar
WITTWER, C. T., RIRIE, K. M., ANDREW, R. V., DAVID, D. A., GUNDRY, R. A. & BALIS, U. J. ( 1997). The LightCycler™: a microvolume multisample fluorimeter with rapid temperature control. Biotechniques 22, 176–178.Google Scholar