Adenylosuccinate synthetase and adenylosuccinate lyase deficiencies trigger growth and infectivity deficits in Leishmania donovani

Jan M. Boitz, Rona Strasser, Phillip Yates, Armando Jardim, Buddy Ullman

Research output: Contribution to journalArticle

22 Citations (Scopus)

Abstract

Leishmania are auxotrophic for purines, and consequently purine acquisition from the host is a requisite nutritional function for the parasite. Both adenylosuccinate synthetase (ADSS) and adenylosuccinate lyase (ASL) have been identified as vital components of purine salvage in Leishmania donovani, and therefore Δadss and Δasl null mutants were constructed to test this hypothesis. Unlike wild type L. donovani, Δadss and Δasl parasites in culture exhibited a profoundly restricted growth phenotype in which the only permissive growth conditions were a 6-aminopurine source in the presence of α′-deoxycoformycin, an inhibitor of adenine aminohydrolase activity. Although both knock-outs showed a diminished capacity to infect murine peritoneal macrophages, only the Δasl null mutant was profoundly incapacitated in its ability to infect mice. The enormous discrepancy in parasite loads observed in livers and spleens from mice infected with either Δadss or Δasl parasites can be explained by selective accumulation of adenylosuccinate in the Δasl knock-out andconsequent starvation for guanylate nucleotides. Geneticcomplementation of a Δasl lesion in Escherichia coli implied that the L. donovaniASLcould also recognize 5-aminoimidazole-(N-succinylocarboxamide) ribotide as a substrate, and purified recombinant ASL displayed an apparent Km of ∼24 μM for adenylosuccinate. Unlike many components of the purine salvage pathway of L. donovani, both ASL and ADSS are cytosolic enzymes. Overall, these data underscore the paramount importance of ASL to purine salvage by both life cycle stages of L. donovani and authenticate ASL as a potential drug target in Leishmania.

Original languageEnglish (US)
Pages (from-to)8977-8990
Number of pages14
JournalJournal of Biological Chemistry
Volume288
Issue number13
DOIs
StatePublished - Mar 29 2013

Fingerprint

Adenylosuccinate Synthase
Adenylosuccinate Lyase
Leishmania donovani
Salvaging
Growth
Parasites
Leishmania
adenine deaminase
Pentostatin
Parasite Load
Purines
Macrophages
Peritoneal Macrophages
Starvation
Life Cycle Stages
Liver
Escherichia coli
Life cycle
Spleen
Nucleotides

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Adenylosuccinate synthetase and adenylosuccinate lyase deficiencies trigger growth and infectivity deficits in Leishmania donovani. / Boitz, Jan M.; Strasser, Rona; Yates, Phillip; Jardim, Armando; Ullman, Buddy.

In: Journal of Biological Chemistry, Vol. 288, No. 13, 29.03.2013, p. 8977-8990.

Research output: Contribution to journalArticle

@article{4ca36480a373491b86a3db973b2ac2ad,
title = "Adenylosuccinate synthetase and adenylosuccinate lyase deficiencies trigger growth and infectivity deficits in Leishmania donovani",
abstract = "Leishmania are auxotrophic for purines, and consequently purine acquisition from the host is a requisite nutritional function for the parasite. Both adenylosuccinate synthetase (ADSS) and adenylosuccinate lyase (ASL) have been identified as vital components of purine salvage in Leishmania donovani, and therefore Δadss and Δasl null mutants were constructed to test this hypothesis. Unlike wild type L. donovani, Δadss and Δasl parasites in culture exhibited a profoundly restricted growth phenotype in which the only permissive growth conditions were a 6-aminopurine source in the presence of α′-deoxycoformycin, an inhibitor of adenine aminohydrolase activity. Although both knock-outs showed a diminished capacity to infect murine peritoneal macrophages, only the Δasl null mutant was profoundly incapacitated in its ability to infect mice. The enormous discrepancy in parasite loads observed in livers and spleens from mice infected with either Δadss or Δasl parasites can be explained by selective accumulation of adenylosuccinate in the Δasl knock-out andconsequent starvation for guanylate nucleotides. Geneticcomplementation of a Δasl lesion in Escherichia coli implied that the L. donovaniASLcould also recognize 5-aminoimidazole-(N-succinylocarboxamide) ribotide as a substrate, and purified recombinant ASL displayed an apparent Km of ∼24 μM for adenylosuccinate. Unlike many components of the purine salvage pathway of L. donovani, both ASL and ADSS are cytosolic enzymes. Overall, these data underscore the paramount importance of ASL to purine salvage by both life cycle stages of L. donovani and authenticate ASL as a potential drug target in Leishmania.",
author = "Boitz, {Jan M.} and Rona Strasser and Phillip Yates and Armando Jardim and Buddy Ullman",
year = "2013",
month = "3",
day = "29",
doi = "10.1074/jbc.M112.431486",
language = "English (US)",
volume = "288",
pages = "8977--8990",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "13",

}

TY - JOUR

T1 - Adenylosuccinate synthetase and adenylosuccinate lyase deficiencies trigger growth and infectivity deficits in Leishmania donovani

AU - Boitz, Jan M.

AU - Strasser, Rona

AU - Yates, Phillip

AU - Jardim, Armando

AU - Ullman, Buddy

PY - 2013/3/29

Y1 - 2013/3/29

N2 - Leishmania are auxotrophic for purines, and consequently purine acquisition from the host is a requisite nutritional function for the parasite. Both adenylosuccinate synthetase (ADSS) and adenylosuccinate lyase (ASL) have been identified as vital components of purine salvage in Leishmania donovani, and therefore Δadss and Δasl null mutants were constructed to test this hypothesis. Unlike wild type L. donovani, Δadss and Δasl parasites in culture exhibited a profoundly restricted growth phenotype in which the only permissive growth conditions were a 6-aminopurine source in the presence of α′-deoxycoformycin, an inhibitor of adenine aminohydrolase activity. Although both knock-outs showed a diminished capacity to infect murine peritoneal macrophages, only the Δasl null mutant was profoundly incapacitated in its ability to infect mice. The enormous discrepancy in parasite loads observed in livers and spleens from mice infected with either Δadss or Δasl parasites can be explained by selective accumulation of adenylosuccinate in the Δasl knock-out andconsequent starvation for guanylate nucleotides. Geneticcomplementation of a Δasl lesion in Escherichia coli implied that the L. donovaniASLcould also recognize 5-aminoimidazole-(N-succinylocarboxamide) ribotide as a substrate, and purified recombinant ASL displayed an apparent Km of ∼24 μM for adenylosuccinate. Unlike many components of the purine salvage pathway of L. donovani, both ASL and ADSS are cytosolic enzymes. Overall, these data underscore the paramount importance of ASL to purine salvage by both life cycle stages of L. donovani and authenticate ASL as a potential drug target in Leishmania.

AB - Leishmania are auxotrophic for purines, and consequently purine acquisition from the host is a requisite nutritional function for the parasite. Both adenylosuccinate synthetase (ADSS) and adenylosuccinate lyase (ASL) have been identified as vital components of purine salvage in Leishmania donovani, and therefore Δadss and Δasl null mutants were constructed to test this hypothesis. Unlike wild type L. donovani, Δadss and Δasl parasites in culture exhibited a profoundly restricted growth phenotype in which the only permissive growth conditions were a 6-aminopurine source in the presence of α′-deoxycoformycin, an inhibitor of adenine aminohydrolase activity. Although both knock-outs showed a diminished capacity to infect murine peritoneal macrophages, only the Δasl null mutant was profoundly incapacitated in its ability to infect mice. The enormous discrepancy in parasite loads observed in livers and spleens from mice infected with either Δadss or Δasl parasites can be explained by selective accumulation of adenylosuccinate in the Δasl knock-out andconsequent starvation for guanylate nucleotides. Geneticcomplementation of a Δasl lesion in Escherichia coli implied that the L. donovaniASLcould also recognize 5-aminoimidazole-(N-succinylocarboxamide) ribotide as a substrate, and purified recombinant ASL displayed an apparent Km of ∼24 μM for adenylosuccinate. Unlike many components of the purine salvage pathway of L. donovani, both ASL and ADSS are cytosolic enzymes. Overall, these data underscore the paramount importance of ASL to purine salvage by both life cycle stages of L. donovani and authenticate ASL as a potential drug target in Leishmania.

UR - http://www.scopus.com/inward/record.url?scp=84875973362&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84875973362&partnerID=8YFLogxK

U2 - 10.1074/jbc.M112.431486

DO - 10.1074/jbc.M112.431486

M3 - Article

C2 - 23404497

AN - SCOPUS:84875973362

VL - 288

SP - 8977

EP - 8990

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 13

ER -