Characterization of farnesylated protein tyrosine phosphatase TcPRL-1 from Trypanosoma cruzi

Ileana C. Cuevas, Peter Rohloff, Daniel O. Sánchez, Roberto Docampo

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

Protein tyrosine kinases and phosphatases play important roles in the regulation of cell growth, development, and differentiation. We report here the identification in Trypanosoma cruzi of a gene (TcPRL-1) encoding a protein tyrosine phosphatase. The predicted protein (TcPRL-1) shares ca. 35% identity with the mammalian protein tyrosine phosphatase known as phosphatase of regenerating liver 1 (PRL-1). Four copies of this protein tyrosine phosphatase are present in the T. cruzi genome, and Northern blot assays showed a transcript of ∼750 bases. TcPRL-1 was detected by Western blot analysis only in amastigote extracts as a 21-kDa protein. TcPRL-1 was expressed in Escherichia coli, and its phosphatase activity was determined by using p- nitrophenylphosphate and a phosphorylated protein as substrates. In contrast to other PRLs, TcPRL-1 activity was not affected by pentamidine, and it was inhibited by very low concentrations of o-vanadate. TcPRL-1 has a C-terminal CAAX motif (CAVM) and is farnesylated in vitro by T. cruzi epimastigote extracts and in vivo according to the transfection results. After transfection of T. cruzi with a vector that expresses TcPRL-1 as a C-terminal fusion to green fluorescent protein, GFP-TcPRL-1 was detected in the endocytic pathway of epimastigotes, amastigotes, and trypomastigotes by colocalization with cruzipain and concanavalin A. Interestingly, a mutant form without the CAAX motif localized to the cytoplasm, in contrast to its mammalian counterparts that localize to the nucleus. The results of these studies on TcPRL-1 reveal that, even though the animal and parasite PRLs share similar kinetic properties, their susceptibilities to inhibitors, as well as their localization, are distinct, implying that they may be involved in different cellular processes.

Original languageEnglish (US)
Pages (from-to)1550-1561
Number of pages12
JournalEukaryotic Cell
Volume4
Issue number9
DOIs
StatePublished - Sep 2005

Fingerprint

protein-tyrosine-phosphatase
Protein Tyrosine Phosphatases
Trypanosoma cruzi
epimastigotes
amastigotes
transfection
Phosphoric Monoester Hydrolases
Transfection
cruzipain
Pentamidine
trypomastigotes
Proteins
protein-tyrosine kinases
Vanadates
proteins
extracts
concanavalin A
Concanavalin A
Green Fluorescent Proteins
Growth and Development

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Microbiology

Cite this

Characterization of farnesylated protein tyrosine phosphatase TcPRL-1 from Trypanosoma cruzi. / Cuevas, Ileana C.; Rohloff, Peter; Sánchez, Daniel O.; Docampo, Roberto.

In: Eukaryotic Cell, Vol. 4, No. 9, 09.2005, p. 1550-1561.

Research output: Contribution to journalArticle

Cuevas, Ileana C. ; Rohloff, Peter ; Sánchez, Daniel O. ; Docampo, Roberto. / Characterization of farnesylated protein tyrosine phosphatase TcPRL-1 from Trypanosoma cruzi. In: Eukaryotic Cell. 2005 ; Vol. 4, No. 9. pp. 1550-1561.
@article{ed8fa08b448d45f581688cc758eac9e8,
title = "Characterization of farnesylated protein tyrosine phosphatase TcPRL-1 from Trypanosoma cruzi",
abstract = "Protein tyrosine kinases and phosphatases play important roles in the regulation of cell growth, development, and differentiation. We report here the identification in Trypanosoma cruzi of a gene (TcPRL-1) encoding a protein tyrosine phosphatase. The predicted protein (TcPRL-1) shares ca. 35{\%} identity with the mammalian protein tyrosine phosphatase known as phosphatase of regenerating liver 1 (PRL-1). Four copies of this protein tyrosine phosphatase are present in the T. cruzi genome, and Northern blot assays showed a transcript of ∼750 bases. TcPRL-1 was detected by Western blot analysis only in amastigote extracts as a 21-kDa protein. TcPRL-1 was expressed in Escherichia coli, and its phosphatase activity was determined by using p- nitrophenylphosphate and a phosphorylated protein as substrates. In contrast to other PRLs, TcPRL-1 activity was not affected by pentamidine, and it was inhibited by very low concentrations of o-vanadate. TcPRL-1 has a C-terminal CAAX motif (CAVM) and is farnesylated in vitro by T. cruzi epimastigote extracts and in vivo according to the transfection results. After transfection of T. cruzi with a vector that expresses TcPRL-1 as a C-terminal fusion to green fluorescent protein, GFP-TcPRL-1 was detected in the endocytic pathway of epimastigotes, amastigotes, and trypomastigotes by colocalization with cruzipain and concanavalin A. Interestingly, a mutant form without the CAAX motif localized to the cytoplasm, in contrast to its mammalian counterparts that localize to the nucleus. The results of these studies on TcPRL-1 reveal that, even though the animal and parasite PRLs share similar kinetic properties, their susceptibilities to inhibitors, as well as their localization, are distinct, implying that they may be involved in different cellular processes.",
author = "Cuevas, {Ileana C.} and Peter Rohloff and S{\'a}nchez, {Daniel O.} and Roberto Docampo",
year = "2005",
month = "9",
doi = "10.1128/EC.4.9.1550-1561.2005",
language = "English (US)",
volume = "4",
pages = "1550--1561",
journal = "Eukaryotic Cell",
issn = "1535-9778",
publisher = "American Society for Microbiology",
number = "9",

}

TY - JOUR

T1 - Characterization of farnesylated protein tyrosine phosphatase TcPRL-1 from Trypanosoma cruzi

AU - Cuevas, Ileana C.

AU - Rohloff, Peter

AU - Sánchez, Daniel O.

AU - Docampo, Roberto

PY - 2005/9

Y1 - 2005/9

N2 - Protein tyrosine kinases and phosphatases play important roles in the regulation of cell growth, development, and differentiation. We report here the identification in Trypanosoma cruzi of a gene (TcPRL-1) encoding a protein tyrosine phosphatase. The predicted protein (TcPRL-1) shares ca. 35% identity with the mammalian protein tyrosine phosphatase known as phosphatase of regenerating liver 1 (PRL-1). Four copies of this protein tyrosine phosphatase are present in the T. cruzi genome, and Northern blot assays showed a transcript of ∼750 bases. TcPRL-1 was detected by Western blot analysis only in amastigote extracts as a 21-kDa protein. TcPRL-1 was expressed in Escherichia coli, and its phosphatase activity was determined by using p- nitrophenylphosphate and a phosphorylated protein as substrates. In contrast to other PRLs, TcPRL-1 activity was not affected by pentamidine, and it was inhibited by very low concentrations of o-vanadate. TcPRL-1 has a C-terminal CAAX motif (CAVM) and is farnesylated in vitro by T. cruzi epimastigote extracts and in vivo according to the transfection results. After transfection of T. cruzi with a vector that expresses TcPRL-1 as a C-terminal fusion to green fluorescent protein, GFP-TcPRL-1 was detected in the endocytic pathway of epimastigotes, amastigotes, and trypomastigotes by colocalization with cruzipain and concanavalin A. Interestingly, a mutant form without the CAAX motif localized to the cytoplasm, in contrast to its mammalian counterparts that localize to the nucleus. The results of these studies on TcPRL-1 reveal that, even though the animal and parasite PRLs share similar kinetic properties, their susceptibilities to inhibitors, as well as their localization, are distinct, implying that they may be involved in different cellular processes.

AB - Protein tyrosine kinases and phosphatases play important roles in the regulation of cell growth, development, and differentiation. We report here the identification in Trypanosoma cruzi of a gene (TcPRL-1) encoding a protein tyrosine phosphatase. The predicted protein (TcPRL-1) shares ca. 35% identity with the mammalian protein tyrosine phosphatase known as phosphatase of regenerating liver 1 (PRL-1). Four copies of this protein tyrosine phosphatase are present in the T. cruzi genome, and Northern blot assays showed a transcript of ∼750 bases. TcPRL-1 was detected by Western blot analysis only in amastigote extracts as a 21-kDa protein. TcPRL-1 was expressed in Escherichia coli, and its phosphatase activity was determined by using p- nitrophenylphosphate and a phosphorylated protein as substrates. In contrast to other PRLs, TcPRL-1 activity was not affected by pentamidine, and it was inhibited by very low concentrations of o-vanadate. TcPRL-1 has a C-terminal CAAX motif (CAVM) and is farnesylated in vitro by T. cruzi epimastigote extracts and in vivo according to the transfection results. After transfection of T. cruzi with a vector that expresses TcPRL-1 as a C-terminal fusion to green fluorescent protein, GFP-TcPRL-1 was detected in the endocytic pathway of epimastigotes, amastigotes, and trypomastigotes by colocalization with cruzipain and concanavalin A. Interestingly, a mutant form without the CAAX motif localized to the cytoplasm, in contrast to its mammalian counterparts that localize to the nucleus. The results of these studies on TcPRL-1 reveal that, even though the animal and parasite PRLs share similar kinetic properties, their susceptibilities to inhibitors, as well as their localization, are distinct, implying that they may be involved in different cellular processes.

UR - http://www.scopus.com/inward/record.url?scp=25144479943&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=25144479943&partnerID=8YFLogxK

U2 - 10.1128/EC.4.9.1550-1561.2005

DO - 10.1128/EC.4.9.1550-1561.2005

M3 - Article

VL - 4

SP - 1550

EP - 1561

JO - Eukaryotic Cell

JF - Eukaryotic Cell

SN - 1535-9778

IS - 9

ER -