Stable amplification of a linear extrachromosomal DNA in mycophenolic acid-resistant Leishmania donovani

Keith Wilson, Stephen M. Beverley, Buddy Ullman

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

Pulsed field gel electrophoretic analysis of chromosomes of MPA 100 cells, a strain of Leishmania donovani that possesses an approx. 15-fold amplified IMP dehydrogenase (IMPDH) gene copy number, revealed a new 280-kb extrachromosomal DNA, IMPDH-280, that was not present in wild type parental cells. Southern blots of these pulsed field gels revealed that the vast majority of the amplified impdh genes were localized on IMPDH-280. In addition to the 700-kb wild type chromosome, the impdh probe also recognized a 740-kb chromosome in the MPA100 genome. The pulse time-dependent relative mobility of IMPDH-280 in pulsed field gels, the failure of limited γ-irradiation to generate a new discrete DNA fragment, and the susceptibility of IMPDH-280 to λ-exonuclease digestion, demonstrated that IMPDH-280 was a linear molecule. IMPDH-280 was also recognized by a telomere probe but not by fragments derived from amplified DNAs found in other drug-resistant Leishmania. IMPDH-280 and the drug resistance phenotype remained stable when MPA100 cells were propagated in the absence of drug for 2 years. The appearance of IMPDH-280 in MPA 100 cells represents one of the first examples of an amplification of a linear extrachromosomal DNA element mediating drug resistance in Leishmania and the first instance of a linear DNA amplification that is stable in the absence of selective pressure.

Original languageEnglish (US)
Pages (from-to)197-206
Number of pages10
JournalMolecular and Biochemical Parasitology
Volume55
Issue number1-2
DOIs
StatePublished - 1992

Fingerprint

IMP Dehydrogenase
Mycophenolic Acid
Leishmania donovani
DNA
Chromosomes
Leishmania
Gels
Drug Resistance
Exonucleases
Gene Dosage
Telomere
Southern Blotting
Pharmaceutical Preparations
Digestion
Genome

Keywords

  • Drug resistance
  • Extrachromosomal element
  • Gene amplification
  • IMP dehydrogenase
  • Leishmania donovani
  • Pulsed field gel electrophoresis

ASJC Scopus subject areas

  • Molecular Biology
  • Parasitology

Cite this

Stable amplification of a linear extrachromosomal DNA in mycophenolic acid-resistant Leishmania donovani. / Wilson, Keith; Beverley, Stephen M.; Ullman, Buddy.

In: Molecular and Biochemical Parasitology, Vol. 55, No. 1-2, 1992, p. 197-206.

Research output: Contribution to journalArticle

Wilson, Keith ; Beverley, Stephen M. ; Ullman, Buddy. / Stable amplification of a linear extrachromosomal DNA in mycophenolic acid-resistant Leishmania donovani. In: Molecular and Biochemical Parasitology. 1992 ; Vol. 55, No. 1-2. pp. 197-206.
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