Hostname: page-component-8448b6f56d-dnltx Total loading time: 0 Render date: 2024-04-25T01:46:32.814Z Has data issue: false hasContentIssue false
  • English
  • Français

In vitro selection of Phytomonas serpens cells resistant to the calpain inhibitor MDL28170: alterations in fitness and expression of the major peptidases and efflux pumps

Published online by Cambridge University Press:  17 October 2017

SIMONE S. C. OLIVEIRA ,
INÊS C. GONÇALVES ,
VITOR ENNES-VIDAL ,
ANGELA H. C. S. LOPES ,
RUBEM F. S. MENNA-BARRETO ,
CLAUDIA M. D’ÁVILA-LEVY ,
ANDRÉ L. S. SANTOS  and
MARTA H. BRANQUINHA
SIMONE S. C. OLIVEIRA
Affiliation:
Laboratório de Investigação de Peptidases, Departamento de Microbiologia Geral, Instituto de Microbiologia Paulo de Góes (IMPG), Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil
INÊS C. GONÇALVES
Affiliation:
Laboratório de Bioquímica de Microrganismos, Departamento de Microbiologia Geral, IMPG, UFRJ, Rio de Janeiro, Brazil
VITOR ENNES-VIDAL
Affiliation:
Laboratório de Estudos Integrados em Protozoologia, Coleção de Protozoários, IOC, FIOCRUZ, Rio de Janeiro, Brazil
ANGELA H. C. S. LOPES
Affiliation:
Laboratório de Bioquímica de Microrganismos, Departamento de Microbiologia Geral, IMPG, UFRJ, Rio de Janeiro, Brazil
RUBEM F. S. MENNA-BARRETO
Affiliation:
Laboratório de Biologia Celular, Instituto Oswaldo Cruz (IOC), Fundação Oswaldo Cruz (FIOCRUZ), Rio de Janeiro, Brazil
CLAUDIA M. D’ÁVILA-LEVY
Affiliation:
Laboratório de Estudos Integrados em Protozoologia, Coleção de Protozoários, IOC, FIOCRUZ, Rio de Janeiro, Brazil
ANDRÉ L. S. SANTOS
Affiliation:
Laboratório de Investigação de Peptidases, Departamento de Microbiologia Geral, Instituto de Microbiologia Paulo de Góes (IMPG), Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil Programa de Pós-Graduação em Bioquímica, Instituto de Química, UFRJ, Rio de Janeiro, Brazil
MARTA H. BRANQUINHA*
Affiliation:
Laboratório de Investigação de Peptidases, Departamento de Microbiologia Geral, Instituto de Microbiologia Paulo de Góes (IMPG), Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil
*
*Corresponding author: Departamento de Microbiologia Geral, Instituto de Microbiologia Paulo de Góes, Universidade Federal do Rio de Janeiro, Av. Carlos Chagas Filho 373, Centro de Ciências da Saúde, 21941-902, Rio de Janeiro, Brazil. E-mail: mbranquinha@micro.ufrj.br
Get access Rights & Permissions [Opens in a new window]

Summary

The species Phytomonas serpens is known to express some molecules displaying similarity to those described in trypanosomatids pathogenic to humans, such as peptidases from Trypanosoma cruzi (cruzipain) and Leishmania spp. (gp63). In this work, a population of P. serpens resistant to the calpain inhibitor MDL28170 at 70 µm (MDLR population) was selected by culturing promastigotes in increasing concentrations of the drug. The only relevant ultrastructural difference between wild-type (WT) and MDLR promastigotes was the presence of microvesicles within the flagellar pocket of the latter. MDLR population also showed an increased reactivity to anti-cruzipain antibody as well as a higher papain-like proteolytic activity, while the expression of calpain-like molecules cross-reactive to anti-Dm-calpain (from Drosophila melanogaster) antibody and calcium-dependent cysteine peptidase activity were decreased. Gp63-like molecules also presented a diminished expression in MDLR population, which is probably correlated to the reduction in the parasite adhesion to the salivary glands of the insect vector Oncopeltus fasciatus. A lower accumulation of Rhodamine 123 was detected in MDLR cells when compared with the WT population, a phenotype that was reversed when MDLR cells were treated with cyclosporin A and verapamil. Collectively, our results may help in the understanding of the roles of calpain inhibitors in trypanosomatids.

Keywords

Phytomonascalpain-like proteinscalpain inhibitorsresistancepeptidases
Type
Research Article
Information
Parasitology , Volume 145 , Issue 3 , March 2018 , pp. 355 - 370
Check for updates
Copyright
Copyright © Cambridge University Press 2017 

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

Al-Mohammed, H. I., Chance, M. L. and Bates, P. A. (2005). Production and characterization of stable amphotericin-resistant amastigotes and promastigotes of Leishmania mexicana . Antimicrobial Agents and Chemotherapy 49, 3274–3280.CrossRefGoogle ScholarPubMed
Andrade, H. M., Murta, S. M., Chapeaurouge, A., Perales, J., Nirdé, P. and Romanha, A. J. (2008). Proteomic analysis of Trypanosoma cruzi resistance to benznidazole. Journal of Proteome Research 7, 2357–2367.CrossRefGoogle ScholarPubMed
Atsma, D. E., Bastiaanse, E. M., Jerzewski, A., Van der Valk, L. J. and Van der Laarse, A. (1995). Role of calcium-activated neutral protease (calpain) in cell death in cultured neonatal rat cardiomyocytes during metabolic inhibition. Circulation Research 76, 1071–1078.CrossRefGoogle ScholarPubMed
Beyette, J. R., Emori, Y. and Mykles, D. L. (1997). Immunological analysis of two calpain-like Ca2+- dependent proteinases from lobster striated muscles: relationship to mammalian and Drosophila calpains. Archives of Biochemistry and Biophysics 15, 337–341.Google Scholar
Branquinha, M. H., Marinho, F. A., Sangenito, L. S., Oliveira, S. S., Gonçalves, K. C., Ennes-Vidal, V., D'Avila-Levy, C. M. and Santos, A. L. (2013). Calpains: potential targets for alternative chemotherapeutic intervention against human pathogenic trypanosomatids. Current Medicinal Chemistry 20, 3174–3185.CrossRefGoogle ScholarPubMed
Brotherton, M. C., Bourassa, S., Leprohon, P., Légaré, D., Poirier, G. G., Droit, A. and Ouellette, M. (2013). Proteomic and genomic analyses of antimony-resistant Leishmania infantum mutant. PLoS ONE 8, e81899.CrossRefGoogle ScholarPubMed
Caljon, G., De Muylder, G., Durnez, L., Jennes, W., Vanaerschot, M. and Dujardin, J. C. (2016). Alice in microbe's land: adaptations and counter-adaptations of vector-borne parasitic protozoa and their hosts. FEMS Microbiology Review 40, 664–685.CrossRefGoogle ScholarPubMed
Campos, M. C., Castro-Pinto, D. B., Ribeiro, G. A., Berredo-Pinho, M. M., Gomes, L. H., da Silva Bellieny, M. S., Goulart, C. M., Echevarria, A. and Leon, L. L. (2013). P-glycoprotein efflux pump plays an important role in Trypanosoma cruzi drug resistance. Parasitology Research 112, 2341–2351.CrossRefGoogle ScholarPubMed
Cazzulo, J. J., Cazzulo Franke, M. C., Martínez, J. and Franke De Cazzulo, B. M. (1990). Some kinetic properties of a cysteine proteinase (cruzipain) from Trypanosoma cruzi . Biochimica et Biophysica Acta 1037, 186–191.CrossRefGoogle ScholarPubMed
Cunha, V., Burkhardt-Medicke, K., Wellner, P., Santos, M. M., Moradas-Ferreira, P., Luckenbach, T. and Ferreira, M. (2017). Effects of pharmaceuticals and personal care products (PPCPs) on multixenobiotic resistance (MXR) related efflux transporter activity in zebrafish (Danio rerio). Ecotoxicology and Environmental Safety 136, 14–23.CrossRefGoogle ScholarPubMed
d'Avila-Levy, C. M., Altoé, E. C., Uehara, L. A., Branquinha, M. H. and Santos, A. L. (2014). GP63 function in the interaction of trypanosomatids with the invertebrate host: facts and prospects. Subcellular Biochemistry 74, 253–270.CrossRefGoogle ScholarPubMed
Donkor, I. O. (2015). An updated patent review of calpain inhibitors (2012–2014). Expert Opinion on Therapeutic Patents 25, 17–31.CrossRefGoogle Scholar
Emori, Y. and Saigo, K. (1994). Calpain localization changes in coordination with actin-related cytoskeletal changes during early embryonic development of Drosophila . Journal of Biological Chemistry 269, 25137–25142.CrossRefGoogle ScholarPubMed
Engel, J. C., Torres, C., Hsieh, I., Doyle, P. S. and McKerrow, J. H. (2000). Upregulation of the secretory pathway in cysteine protease inhibitor-resistant Trypanosoma cruzi . Journal of Cell Science 113, 1345–1354.CrossRefGoogle ScholarPubMed
Ennes-Vidal, V., Menna-Barreto, R. F., Branquinha, M. H., Santos, A. L. S. and D'Avila-Levy, C. M. (2017). Why calpain inhibitors are interesting leading compounds to search for new therapeutic options to treat leishmaniasis? Parasitology 144, 117–123.CrossRefGoogle ScholarPubMed
Ersfeld, K., Barraclough, H. and Gull, K. (2005). Evolutionary relationships and protein domain architecture in an expanded calpain superfamily in kinetoplastid parasites. Journal of Molecular Evolution 61, 742–757.CrossRefGoogle Scholar
Forster, S., Thumser, A. E., Hood, S. R. and Plant, N. (2012). Characterization of rhodamine-123 as a tracer dye for use in in vitro drug transport assays. PLoS ONE 7, e33253.CrossRefGoogle ScholarPubMed
Gueiros-Filho, F. J., Viola, J. P., Gomes, F. C., Farina, M., Lins, U., Bertho, A. L., Wirth, D. F. and Lopes, U. G. (1995). Leishmania amazonensis: multidrug resistance in vinblastine-resistant promastigotes is associated with rhodamine 123 efflux, DNA amplification, and RNA overexpression of a Leishmania mdr1 gene. Experimental Parasitology 81, 480–490.CrossRefGoogle ScholarPubMed
Hayes, P., Varga, V., Olego-Fernandez, S., Sunter, J., Ginger, M. L. and Gull, K. (2014). Modulation of a cytoskeletal calpain-like protein induces major transitions in trypanosome morphology. Journal of Cellular Biology 206, 377–384.CrossRefGoogle ScholarPubMed
Hertz-Fowler, C., Ersfeld, K. and Gull, K. (2001). CAP5·5, a life-cycle-regulated, cytoskeleton-associated protein is a member of a novel family of calpain-related proteins in Trypanosoma brucei . Molecular and Biochemical Parasitology 116, 25–34.CrossRefGoogle ScholarPubMed
Heussen, C. and Dowdle, E. B. (1980). Electrophoretic analysis of plasminogen activators in polyacrylamide gels containing sodium dodecyl sulphate and copolymerized substrates. Analytical Biochemistry 102, 196–202.CrossRefGoogle ScholarPubMed
Jaskowska, E., Butler, C., Preston, G. and Kelly, S. (2015). Phytomonas: trypanosomatids adapted to plant environments. PLoS Pathogens 11, e1004484.CrossRefGoogle ScholarPubMed
Lowry, O. H., Rosebrough, N. J., Farr, A. L. and Randall, R. J. (1951). Protein measurement with the Folin phenol reagent. Journal of Biological Chemistry 193, 265–275.CrossRefGoogle ScholarPubMed
Marinho, F. A., Gonçalves, K. C., Oliveira, S. S., Gonçalves, D. S., Matteoli, F. P., Seabra, S. H., Oliveira, A. C., Bellio, M., Oliveira, S. S., Souto-Padrón, T., d'Avila-Levy, C. M., Santos, A. L. and Branquinha, M. H. (2014). The calpain inhibitor MDL28170 induces the expression of apoptotic markers in Leishmania amazonensis promastigotes. PLoS ONE 9, e87659.CrossRefGoogle ScholarPubMed
Morgan, G. W., Hall, B. S., Denny, P. W., Carrington, M. and Field, M. C. (2002). The kinetoplastida endocytic apparatus. Part I: a dynamic system for nutrition and evasion of host defences. Trends in Parasitology 18, 491–496.CrossRefGoogle Scholar
Olego-Fernandez, S., Vaughan, S., Shaw, M. K., Gull, K. and Ginger, M. L. (2009). Cell morphogenesis of Trypanosoma brucei requires the paralogous, differentially expressed calpain-related proteins CAP5·5 and CAP5·5V . Protist 60, 576–590.CrossRefGoogle Scholar
Oliveira, S. S. C., Goncalves, D. S., Garcia-Gomes, A. S., Goncalves, I. C., Seabra, S. H., Menna-Barreto, R. F. S., Lopes, A. H. C. S., d'Avila-Levy, C. M., Santos, A. L. S. and Branquinha, M. H. (2017). Susceptibility of Phytomonas serpens to calpain inhibitors in vitro: interference on the proliferation, ultrastructure, cysteine peptidase expression and interaction with the invertebrate host. Memórias do Instituto Oswaldo Cruz 112, 31–43.CrossRefGoogle ScholarPubMed
Ono, Y. and Sorimachi, H. (2012). Calpains: an elaborate proteolytic system. Biochimica et Biophysica Acta 1824, 224–236.CrossRefGoogle ScholarPubMed
Porcel, B. M., Denoeud, F., Opperdoes, F., Noel, B., Madoui, M. A., Hammarton, T. C., Field, M. C., da Silva, C., Couloux, A., Poulain, J., Katinka, M., Jabbari, K., Aury, J. M., Campbell, D. A., Cintron, R., Dickens, N. J., Docampo, R., Sturm, N. R., Koumandou, V. L., Fabre, S., Flegontov, P., Lukeš, J., Michaeli, S., Mottram, J. C., Szöőr, B., Zilberstein, D., Bringaud, F., Wincker, P. and Dollet, M. (2014). The streamlined genome of Phytomonas spp. relative to human pathogenic kinetoplastids reveals a parasite tailored for plants. PLoS Genetics 10, e1004007.CrossRefGoogle ScholarPubMed
Rai, S., Bhaskar, E., Goel, S. K., Nath Dwivedi, U., Sundar, S. and Goyal, N. (2013). Role of efflux pumps and intracellular thiols in natural antimony resistant isolates of Leishmania donovani . PloS ONE 8, e74862.CrossRefGoogle ScholarPubMed
Rami, A., Ferger, D. and Krieglstein, J. (1997). Blockade of calpain proteolytic activity rescues neurons from glutamate excitotoxicity. Neuroscience Research 27, 93–97.CrossRefGoogle ScholarPubMed
Romeiro, A., Solé-Cava, A., Sousa, M. A., De Souza, W. and Attias, M. (2000). Ultrastructural and biochemical characterization of promastigote and cystic forms of Leptomonas wallacei n. sp. isolated from the intestine of its natural host Oncopeltus fasciatus (Hemiptera: Lygaeidae). Journal of Eukaryotic Microbiology 47, 208–220.CrossRefGoogle Scholar
Santos, A. L. S., D´Avila-Levy, C. M., Elias, C. G. R., Vermelho, A. B. and Branquinha, M. H. (2007). Phytomonas serpens: immunological similarities with the human trypanosomatid pathogens. Microbes and Infection 9, 915–921.CrossRefGoogle ScholarPubMed
Vanaerschot, M., Huijben, S., Van den Broeck, F. and Dujardin, J. C. (2014). Drug resistance in vectorborne parasites: multiple actors and scenarios for an evolutionary arms race. FEMS Microbiology Review 38, 41–55.CrossRefGoogle ScholarPubMed
Vergnes, B., Gourbal, B., Girard, I., Sundar, S., Drummelsmith, J. and Ouellette, M. (2007). A proteomics screen implicates HSP83 and a small kinetoplastid calpain-related protein in drug resistance in Leishmania donovani clinical field isolates by modulating drug-induced programmed cell death. Molecular and Cellular Proteomics 6, 88–101.CrossRefGoogle Scholar
Wang, K. K., Nath, R., Posner, A., Raser, K. J., Buroker-Kilgore, M., Hajimohammadreza, I., Probert, A. W., Marcoux, F. W., Ye, Q., Takano, E., Hatanaka, M., Maki, M., Caner, H., Collins, J. L., Fergus, A., Lee, K. S., Lunney, E. A., Hays, S. J. and Yuen, P. (1996). An alpha-mercaptoacrylic acid derivative is a selective nonpeptide cel-permeable calpain inhibitor and is neuroprotective. Proceedings of the National Academy of Sciences USA 93, 6687–6692.CrossRefGoogle ScholarPubMed
Xiong, J., Mao, D. A. and Liu, L. Q. (2015). Research progress on the role of ABC transporters in the drug resistance mechanism of intractable epilepsy. BioMed Research International 2015, 194541.CrossRefGoogle ScholarPubMed
Yasinzai, M., Khan, M., Nadhman, A. and Shahnaz, G. (2013). Drug resistance in leishmaniasis: current drug-delivery systems and future perspectives. Future Medicinal Chemistry 5, 1877–1888.CrossRefGoogle ScholarPubMed
Yong, V., Schmitz, V., Vannier-Santos, M. A., De Lima, A. P., Lalmanach, G., Juliano, L., Gauthier, F. and Scharfstein, J. (2000). Altered expression of cruzipain and a cathepsin B-like target in a Trypanosoma cruzi cell line displaying resistance to synthetic inhibitors of cysteine-proteinases. Molecular and Biochemical Parasitology 109, 47–59.CrossRefGoogle Scholar
Supplementary material: File

Oliveira et al supplementary material

Figure S1

Download Oliveira et al supplementary material(File)
File 374.3 KB
Supplementary material: File

Oliveira et al supplementary material

Table S1

Download Oliveira et al supplementary material(File)
File 13.2 KB
Supplementary material: File

Oliveira et al supplementary material

Table S2

Download Oliveira et al supplementary material(File)
File 13.1 KB