Hostname: page-component-8448b6f56d-xtgtn Total loading time: 0 Render date: 2024-04-19T15:39:56.547Z Has data issue: false hasContentIssue false
  • English
  • Français

Trypanosoma brucei vacuolar protein sorting 41 (VPS41) is required for intracellular iron utilization and maintenance of normal cellular morphology

Published online by Cambridge University Press:  19 June 2007

S. LU ,
T. SUZUKI ,
N. IIZUKA ,
S. OHSHIMA ,
Y. YABU ,
M. SUZUKI ,
L. WEN  and
N. OHTA
S. LU
Affiliation:
Department of Molecular Parasitology, Nagoya City University, Graduate School of Medical Sciences, Nagoya 467-8601, Japan Institute of Parasitic Diseases, Zhejiang Academy of Medical Sciences, HangZhou 310013, China
T. SUZUKI*
Affiliation:
Department of Molecular Parasitology, Nagoya City University, Graduate School of Medical Sciences, Nagoya 467-8601, Japan
N. IIZUKA
Affiliation:
Department of Virology, Nagoya City University, Graduate School of Medical Sciences, Nagoya 467-8601, Japan
S. OHSHIMA
Affiliation:
Department of Molecular Parasitology, Nagoya City University, Graduate School of Medical Sciences, Nagoya 467-8601, Japan
Y. YABU
Affiliation:
Department of Molecular Parasitology, Nagoya City University, Graduate School of Medical Sciences, Nagoya 467-8601, Japan
M. SUZUKI
Affiliation:
Department of Molecular Parasitology, Nagoya City University, Graduate School of Medical Sciences, Nagoya 467-8601, Japan
L. WEN
Affiliation:
Institute of Parasitic Diseases, Zhejiang Academy of Medical Sciences, HangZhou 310013, China
N. OHTA
Affiliation:
Department of Molecular Parasitology, Nagoya City University, Graduate School of Medical Sciences, Nagoya 467-8601, Japan
*
*Corresponding author: Department of Molecular Parasitology, Nagoya City University, Graduate School of Medical Sciences, Nagoya 467-8601, Japan. Tel: +81 52 853 8186. Fax: +81 52 842 0149. E-mail: hakuna@med.nagoya-cu.ac.jp (T. Suzuki).
Get access Rights & Permissions [Opens in a new window]

Summary

Procyclic forms of Trypanosoma brucei brucei remain and propagate in the midgut of tsetse fly where iron is rich. Additional iron is also required for their growth in in vitro culture. However, little is known about the genes involved in iron metabolism and the mechanism of iron utilization in procyclic-form cells. Therefore, we surveyed the genes involved in iron metabolism in the T. b. brucei genome sequence database. We found a potential homologue of vacuole protein sorting 41 (VPS41), a gene that is required for high-affinity iron transport in Saccharomyces cerevisiae and cloned the full-length gene (TbVPS41). Complementation analysis of TbVPS41 in ΔScvps41 yeast cells showed that TbVPS41 could partially suppress the inability of ΔScvps41 yeast cells to grow on low-iron medium, but it could not suppress the fragmented vacuole phenotype. Further RNA interference (RNAi)-mediated gene knock-down in procyclic-form cells resulted in a significant reduction of growth in low-iron medium; however, no change in growth was observed in normal culture medium. Transmission electron microscopy showed that RNAi caused T. b. brucei cells to have larger numbers of small intracellular vesicles, similar to the fragmented vacuoles observed in ΔScvps41 yeast cells. The present study demonstrates that TbVPS41 plays an important role in the intracellular iron utilization system as well as in the maintenance of normal cellular morphology.

Keywords

Trypanosoma brucei bruceiVPS41 geneRNAiiron-utilizationcellular morphology
Type
Research Article
Information
Parasitology , Volume 134 , Issue 11 , October 2007 , pp. 1639 - 1647
Copyright
Copyright © Cambridge University Press 2007

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

Breidbach, T., Scory, S., Krauth-Siegel, R. L. and Steverding, D. (2002). Growth inhibition of bloodstream forms of Trypanosoma brucei by the iron chelator deferoxamine. International Journal for Parasitology 32, 473–479.Google Scholar
Brun, R. and Schönenberger, M. (1979). Cultivation and in vitro cloning of procyclic culture forms of Trypanosoma brucei in a semi-defined medium. Acta Tropica 36, 289–292.Google Scholar
Clarkson, A. B., Bienen, E. J., Pollakis, G. and Grady, R. W. (1989). Respiration of bloodstream forms of the parasite Trypanosoma brucei brucei is dependent on a plant-like alternative oxidase. Journal of Biological Chemistry 264, 17770–17776.Google Scholar
Darsow, T., Katzmann, D. J., Cowles, C. R. and Emr, S. D. (2001). Vps41p function in the alkaline phosphatase pathway requires homo-oligomerization and interaction with AP-3 through two distinct domains. Molecular and Cellular Biology 12, 37–51.Google Scholar
Dormeyer, M., Schoneck, R., Dittmar, G. A. and Krauth-Siegel, R. L. (1997). Cloning, sequencing and expression of ribonucleotide reductase R2 from Trypanosoma brucei. FEBS Letters 414, 449–453.Google Scholar
Eide, D. and Guarente, L. (1992). Increased dosage of a transcriptional acativator gene enhances iron-limited growth of Saccharomyces cerevisiae. Journal of General Microbiology 138, 347–354.Google Scholar
Field, M. C. and Carrington, M. (2004). Intracellular membrane transport systems in Trypanosoma brucei. Traffic 5, 905–913.Google Scholar
Garcia-Salcedo, J. A., Perez-Morga, D., Gijon, P., Dilbeck, V., Pays, E. and Nolan, D. P. (2004). A differential role for actin during the life cycle of Trypanosoma brucei. EMBO Journal 23, 780–789.Google Scholar
Grab, D. J., Wells, C. W., Shaw, M. K., Webster, P. and Russo, D. C. (1992). Endocytosed transferrin in African trypanosomes is delivered to lysosomes and may not be recycled. European Journal of Cell Biology 59, 398–404.Google Scholar
Grab, D. J., Shaw, M. K., Wells, C. W., Verjee, Y., Russo, D. C., Webster, P., Naessens, J. and Fish, W. R. (1993). The transferrin receptor in African trypanosomes: identification, partial characterization and subcellular localization. European Journal of Cell Biology 62, 114–126.Google Scholar
Hall, B. S., Smith, E., Langer, W., Jacobs, L. A., Goulding, D. and Field, M. C. (2005). Developmental variation in Rab11-dependent trafficking in Trypanosoma brucei. Eukaryotic Cell 4, 971–980.Google Scholar
Le Trant, N., Meshnick, S. R., Kitchener, K., Eaton, J. W. and Cerami, A. (1983). Iron-containing superoxide dismutase from Crithidia fasciculata. Purification, characterization, and similarity to Leishmanial and trypanosomal enzymes. Journal of Biological Chemistry 258, 125–130.Google Scholar
Li, L., Chen, O. S., McVey Ward, D. and Kaplan, J. (2001). CCC1 is a transporter that mediates vacuolar iron storage in yeast. Journal of Biological Chemistry 276, 29515–29519.CrossRefGoogle ScholarPubMed
Lipman, D. J. and Pearson, D. J. (1985). Rapid and sensitive protein similarity searches. Science 227, 1435–1441.Google Scholar
Liu, J., Qiao, X., Du, D. and Lee, M. G. (2000). Receptor-mediated endocytosis in the procyclic form of Trypanosoma brucei. Journal of Biological Chemistry 275, 12032–12040.Google Scholar
Radisky, D. C., Snyder, W. B., Emr, S. D. and Kaplan, J. (1997). Characterization of VPS41, a gene required for vacuolar tracfficking and high-affinity iron transport in yeast. Proceedings of the National Academy of Sciences, USA 94, 5662–5666.Google Scholar
Richardson, S. C., Winistorfer, S. C., Poupon, V., Luzio, J. P. and Piper, R. C. (2004). Mammalian late vacuole protein sorting orthologues participate in early endosomal fusion and interact with the cytoskeleton. Molecular Biology of the Cell 15, 1197–1210.CrossRefGoogle ScholarPubMed
Saas, J., Ziegelbauer, K., von Haeseler, A., Fast, B. and Boshart, M. (2000). A developmentally regulated aconitase related to iron-regulatory protein-1 is localized in the cytoplasm and in the mitochondrion of Trypanosoma brucei. Journal of Biological Chemistry 275, 2745–2755.Google Scholar
Sherman, F., Fink, G. R. and Lawrence, L. W. (1986). Laboratory Course Manual for Methods in Yeast Genetics. Cold Spring Harbor Lab, NY, USA.Google Scholar
Sikorski, R. S. and Hieter, P. (1989). A system of shutter vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. Genetics 122, 19–27.Google Scholar
Stearman, R., Yuan, D. S., Yamaguchi-Iwai, Y., Klausner, R. D. and Dancis, A. (1996). A permease-oxidase complex involved in high-affinity iron uptake in yeast. Science 271, 1552–1557.Google Scholar
Steverding, D., Stierhof, Y. D., Chaudhri, M., Ligtenberg, M., Schell, D., Beck-Sickinger, A. G. and Overath, P. (1994). ESAG 6 and 7 products of Trypanosoma brucei form a transferrin binding protein complex. European Journal of Cell Biology 64, 78–87.Google Scholar
Steverding, D., Stierhof, Y. D., Fuchs, H., Tauber, R. and Overath, P. (1995). Transferrin-binding protein complex is the receptor for transferrin uptake in Trypanosoma brucei. Journal of Cell Biology 131, 1173–1182.CrossRefGoogle ScholarPubMed
Wada, Y. O., Ohsumi, Y. and Anraku, Y. (1992). Genes for directing vacuolar morphogenesis in Saccharomyces cerevisiae. Journal of Biological Chemistry 267, 18665–18670.Google Scholar
Wang, Z. F., Morris, J. C., Drew, M. E. and Englund, P. T. (2000). Inhibition of Trypanosoma brucei gene expression by RNA interference using an integratable vector with opposing T7 promoters. Journal of Biological Chemistry 275, 40174–40179.Google Scholar
Warner, T. S., Sinclair, D. A., Fitzpatrick, K. A., Singh, M., Devlin, R. H. and Honda, B. M. (1998). The light gene of Drosophila melanogaster encodes a homologue of VPS41, a yeast gene involved in cellular-protein trafficking. Genome 41, 236–243.Google Scholar
Yuan, D. S., Stearman, R., Dancis, A., Dunn, T., Beeler, T. and Klausner, R. D. (1995). The Menkes/Wilson disease gene homologue in yeast provides copper to a ceruloplasmin-like oxidase required for iron uptake. Proceedings of the National Academy of Sciences, USA 92, 2632–2636.Google Scholar
Zheng, B., Wu, J. N., Schober, W., Lewis, D. E. and Vida, T. (1998). Isolation of yeast mutants defective for localization of vacuolar vital dyes. Proceedings of the National Academy of Sciences, USA 95, 11721–11726.CrossRefGoogle ScholarPubMed