Hostname: page-component-586b7cd67f-rdxmf Total loading time: 0 Render date: 2024-11-26T04:07:59.401Z Has data issue: false hasContentIssue false
  • English
  • Français

The knockdown of each component of the cysteine proteinase-adhesin complex of Entamoeba histolytica (EhCPADH) affects the expression of the other complex element as well as the in vitro and in vivo virulence

Published online by Cambridge University Press:  02 November 2015

RAMÓN OCÁDIZ-RUIZ ,
WENDY FONSECA ,
ALICIA S. LINFORD ,
TIMOTHY P. YOSHINO ,
ESTHER OROZCO  and
MARIO A. RODRÍGUEZ
RAMÓN OCÁDIZ-RUIZ
Affiliation:
Departamento de Infectómica y Patogénesis Molecular, CINVESTAV-IPN, México, D.F. México
WENDY FONSECA
Affiliation:
Departamento de Infectómica y Patogénesis Molecular, CINVESTAV-IPN, México, D.F. México
ALICIA S. LINFORD
Affiliation:
Department of Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, Virginia 22908, USA
TIMOTHY P. YOSHINO
Affiliation:
Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin, Madison, Wisconsin 53706, USA
ESTHER OROZCO
Affiliation:
Departamento de Infectómica y Patogénesis Molecular, CINVESTAV-IPN, México, D.F. México
MARIO A. RODRÍGUEZ*
Affiliation:
Departamento de Infectómica y Patogénesis Molecular, CINVESTAV-IPN, México, D.F. México
*
*Corresponding author: Departamento de Infectómica y Patogénesis Molecular, CINVESTAV-IPN. A.P. 14-740, México, D.F. 07000, México. E-mail: [email protected]
Get access Rights & Permissions [Opens in a new window]

Summary

Entamoeba histolytica is the protozoan parasite causative of human amoebiasis, disease responsible for 40 000–100 000 deaths annually. The cysteine proteinase-adhesin complex of this parasite (EhCPADH) is a heterodimeric protein formed by a cysteine protease (EhCP112) and an adhesin (EhADH) that plays an important role in the cytopathic mechanism of this parasite. The coding genes for EhCP112 and EhADH are adjacent in the E. histolytica genome, suggesting that their expression may be co-regulated, but this hypothesis has not yet been confirmed. Here, we performed the knockdown of EhCP112 and EhADH using gene-specific short-hairpin RNAs (shRNA), and the effect of these knockdowns on the expression of both complex components as well as on the in vitro and in vivo virulence was analysed. Results showed that the knockdown of one of the EhCPADH components produced a simultaneous downregulation of the other protein. Accordingly, a concomitant reduction in the overall expression of the complex was observed. The downregulation of each component also produced a significant decrease in the in vitro and in vivo virulence of trophozoites. These results demonstrated that the expression of EhCP112 and EhADH is co-regulated and confirmed that the EhCPADH complex plays an important role in E. histolytica virulence.

Keywords

Entamoeba histolyticaEhCPADH complexvirulencedownregulationRNA interferenceshRNAexpression co-regulation
Type
Research Article
Information
Parasitology , Volume 143 , Issue 1 , January 2016 , pp. 50 - 59
Check for updates
Copyright
Copyright © Cambridge University Press 2015 

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

REFERENCES

Ali, I. K., Clark, C. G. and Petri, W. A. Jr. (2008). Molecular epidemiology of amebiasis. Infection, Genetics and Evolution: Journal of Molecular Epidemiology and Evolutionary Genetics in Infectious Diseases 8, 698–707.Google Scholar
Arroyo, R. and Orozco, E. (1987). Localization and identification of an Entamoeba histolytica adhesin. Molecular and Biochemical Parasitology 23, 151–158.Google Scholar
Azuara-Liceaga, E., Flores-Soto, E., Lopez-Camarillo, C. and Orozco, E. (2005). Entamoeba histolytica: structural and functional analysis of the Ehadh112 gene promoter. Experimental Parasitology 110, 280–285.Google Scholar
Banuelos, C., Garcia-Rivera, G., Lopez-Reyes, I. and Orozco, E. (2005). Functional characterization of EhADH112: an Entamoeba histolytica Bro1 domain-containing protein. Experimental Parasitology 110, 292–297.CrossRefGoogle ScholarPubMed
Betanzos, A., Javier-Reyna, R., Garcia-Rivera, G., Banuelos, C., Gonzalez-Mariscal, L., Schnoor, M. and Orozco, E. (2013). The EhCPADH112 complex of Entamoeba histolytica interacts with tight junction proteins occludin and claudin-1 to produce epithelial damage. PLoS ONE 8, e65100.CrossRefGoogle ScholarPubMed
Boycheva, S., Daviet, L., Wolfender, J. L. and Fitzpatrick, T. B. (2014). The rise of operon-like gene clusters in plants. Trends in Plant Science 19, 447–459.Google Scholar
Collemare, J., Pianfetti, M., Houlle, A. E., Morin, D., Camborde, L., Gagey, M. J., Barbisan, C., Fudal, I., Lebrun, M. H. and Bohnert, H. U. (2008). Magnaporthe grisea avirulence gene ACE1 belongs to an infection-specific gene cluster involved in secondary metabolism. New Phytologist 179, 196–208.Google Scholar
Dandekar, T., Snel, B., Huynen, M. and Bork, P. (1998). Conservation of gene order: a fingerprint of proteins that physically interact. Trends in Biochemical Sciences 23, 324–328.Google Scholar
Deng, Y., Wang, C. C., Choy, K. W., Du, Q., Chen, J., Wang, Q., Li, L., Chung, T. K. and Tang, T. (2014). Therapeutic potentials of gene silencing by RNA interference: principles, challenges, and new strategies. Gene 538, 217–227.Google Scholar
Diamond, L. S., Harlow, D. R. and Cunnick, C. C. (1978). A new medium for the axenic cultivation of Entamoeba histolytica and other Entamoeba . Transactions of the Royal Society of Tropical Medicine and Hygiene 72, 431–432.Google Scholar
Faust, D. M. and Guillen, N. (2012). Virulence and virulence factors in Entamoeba histolytica, the agent of human amoebiasis. Microbes and Infection/Institut Pasteur 14, 1428–1441.Google Scholar
Flores-Soto, E., Azuara-Liceaga, E., Lopez-Camarillo, C. and Orozco, E. (2005). The Entamoeba histolytica Ehcp112 gene has a distal and weak promoter. Experimental Parasitology 110, 286–291.Google Scholar
Foda, B. M. and Singh, U. (2015). Dimethylated H3K27 is a repressive epigenetic histone mark in the protist Entamoeba histolytica and is significantly enriched in genes silenced via the RNAi pathway. Journal of Biological Chemistry 290, 21114–21130.Google Scholar
Garcia-Rivera, G., Rodriguez, M. A., Ocadiz, R., Martinez-Lopez, M. C., Arroyo, R., Gonzalez-Robles, A. and Orozco, E. (1999). Entamoeba histolytica : a novel cysteine protease and an adhesin form the 112 kDa surface protein. Molecular Microbiology 33, 556–568.Google Scholar
Garcia-Rivera, G., Sanchez, T., Orozco, E. and Guarneros, G. (1982). Isolation of clones of E. histolytica deficient in adhesion to human erythrocytes. Archivos de Investigacion Medica 13(Suppl. 3), 129–136.Google Scholar
Gaunt, S. J. (2015). The significance of Hox gene collinearity. International Journal of Developmental Biology 59, 159–170.Google Scholar
Hittinger, G. T., Rokas, E. and Carroll, S. B. (2004). Parallel inactivation of multiple GAL pathway genes and ecological diversification in yeasts. Proceedings of the National Academy of Sciences USA 101, 14144–14149.Google Scholar
Horton, R., Wilming, L., Rand, V., Lovering, R. C., Bruford, E. A., Khodiyar, V. K., Lush, M. J., Povey, S., Talbot, C. C. Jr., Wright, M. W., Wain, H. M., Trowsdale, J., Ziegler, A. and Beck, S. (2004). Gene map of the extended human MHC. Nature Reviews Genetics 5, 889–899.Google Scholar
Lewis, E. B. (1978). A gene complex controlling segmentation in Drosophila. Nature 276, 565–570.Google Scholar
Linford, A. S., Moreno, H., Good, K. R., Zhang, H., Singh, U. and Petri, W. A. Jr. (2009). Short hairpin RNA-mediated knockdown of protein expression in Entamoeba histolytica . BMC Microbiology 9, 38.Google Scholar
Livak, K. J. and Schmittgen, T. D. (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods 25, 402–408.Google Scholar
Lushbaugh, W. B., Kairalia, A. B., Hofbauer, A. F., Cantey, J. R. and Pittman, F. E. (1978). Cytotoxic activity of a cell-free extract of Entamoeba histolytica . Transactions of the Royal Society of Tropical Medicine and Hygiene 72, 105–106.Google Scholar
Madriz, X., Martinez, M. B., Rodriguez, M. A., Sierra, G., Martinez-Lopez, C., Riveron, A. M., Flores, L. and Orozco, E. (2004). Expression in fibroblasts and in live animals of Entamoeba histolytica polypeptides EhCP112 and EhADH112. Microbiology 150, 1251–1260.Google Scholar
Martinez-Lopez, C., Orozco, E., Sanchez, T., Garcia-Perez, R. M., Hernandez-Hernandez, F. and Rodriguez, M. A. (2004). The EhADH112 recombinant polypeptide inhibits cell destruction and liver abscess formation by Entamoeba histolytica trophozoites. Cellular Microbiology 6, 367–376.CrossRefGoogle ScholarPubMed
Martinez, M. B., Rodriguez, M. A., Garcia-Rivera, G., Sanchez, T., Hernandez-Pando, R., Aguilar, D. and Orozco, E. (2009). A pcDNA-Ehcpadh vaccine against Entamoeba histolytica elicits a protective Th1-like response in hamster liver. Vaccine 27, 4176–4186.Google Scholar
Ocadiz, R., Orozco, E., Carrillo, E., Quintas, L. I., Ortega-Lopez, J., Garcia-Perez, R. M., Sanchez, T., Castillo-Juarez, B. A., Garcia-Rivera, G. and Rodriguez, M. A. (2005). EhCP112 is an Entamoeba histolytica secreted cysteine protease that may be involved in the parasite-virulence. Cellular Microbiology 7, 221–232.CrossRefGoogle ScholarPubMed
Ocadiz-Ruiz, R., Fonseca, W., Martinez, M. B., Ocadiz-Quintanar, R., Orozco, E. and Rodriguez, M. A. (2013). Effect of the silencing of the Ehcp112 gene on the in vitro virulence of Entamoeba histolytica . Parasites and Vectors 6, 248.CrossRefGoogle ScholarPubMed
Orozco, E., Martinez-Palomo, A. and Lopez-Revilla, R. (1978). An in vitro model for the quantitative study of E. histolytica virulence. Archivos de Investigacion Medica 9 (Suppl. 1), 257–260.Google Scholar
Orozco, E., Guarneros, G., Martinez-Palomo, A. and Sanchez, T. (1983). Entamoeba histolytica. Phagocytosis as a virulence factor. Journal of Experimental Medicine 158, 1511–1521.Google Scholar
Ravdin, J. I., Croft, B. Y. and Guerrant, R. L. (1980). Cytopathogenic mechanisms of Entamoeba histolytica . Journal of Experimental Medicine 152, 377–390.Google Scholar
Rodriguez, M. A. and Orozco, E. (1986). Isolation and characterization of phagocytosis - and virulence-deficient mutants of Entamoeba histolytica . Journal of Infectious Diseases 154, 27–32.Google Scholar
Sahoo, N., Bhattacharya, S. and Bhattacharya, A. (2003). Blocking the expression of a calcium binding protein of the protozoan parasite Entamoeba histolytica by tetracycline regulatable antisense-RNA. Molecular and Biochemical Parasitology 126, 281–284.Google Scholar
Solis, C. F. and Guillen, N. (2008). Silencing genes by RNA interference in the protozoan parasite Entamoeba histolytica . Methods in Molecular Biology 442, 113–128.Google Scholar
Voigt, H., Olivo, J. C., Sansonetti, P. and Guillen, N. (1999). Myosin IB from Entamoeba histolytica is involved in phagocytosis of human erythrocytes. Journal of Cell Science 112(Pt 8), 1191–1201.Google Scholar
Ximenez, C., Moran, P., Rojas, L., Valadez, A. and Gomez, A. (2009). Reassessment of the epidemiology of amebiasis: state of the art. Infection, Genetics and evolution: Journal of Molecular Epidemiology and Evolutionary Genetics in Infectious Diseases 9, 1023–1032.Google Scholar
Supplementary material: PDF

Ocadiz-Ruiz supplementary material

Tables S1 and S2

Download Ocadiz-Ruiz supplementary material(PDF)
PDF 13.5 KB