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Localization and activity of multidrug resistance protein 1 in the secretory pathway of Leishmania parasites
Author(s) -
Dodge Matthew A.,
Waller Ross F.,
Chow Larry M. C.,
Zaman Muhammad M.,
Cotton Leanne M.,
McConville Malcolm J.,
Wirth Dyann F.
Publication year - 2004
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.2003.03927.x
Subject(s) - biology , calcein , golgi apparatus , microbiology and biotechnology , secretion , endoplasmic reticulum , rhodamine 123 , green fluorescent protein , secretory pathway , organelle , lysosome , leishmania , endosome , multidrug resistance associated protein 2 , leishmania mexicana , multiple drug resistance , biochemistry , atp binding cassette transporter , transporter , parasite hosting , enzyme , membrane , world wide web , gene , computer science , intracellular , antibiotics
Summary Upregulation of the multidrug resistance protein 1 (LeMDR1) in the protozoan parasite, Leishmania enriettii , confers resistance to hydrophobic drugs such as vinblastine, but increases the sensitivity of these parasites to the mitochondrial drug, rhodamine 123. In order to investigate the mechanism of action of LeMDR1, the subcellular localization of green fluorescent protein (GFP)‐tagged versions of LeMDR1 and the fate of the traceable‐fluorescent LeMDR1 substrate calcein AM were examined in both Leishmania mexicana and L. enriettii LeMDR1 –/– and overexpressing cell lines. The LeMDR1‐GFP chimera was localized by fluorescence microscopy to a number of secretory and endocytic compartments, including the Golgi apparatus, endoplasmic reticulum (ER) and a multivesicular tubule (MVT)‐lysosome. Pulse–chase labelling experiments with calcein AM suggested that the Golgi and ER pools, but not the MVT‐lysosome pool, of LeMDR1 were active in pumping calcein AM out of the cell. Cells labelled with calcein AM under conditions that slow vesicular transport (low temperature and stationary growth) inhibited export and resulted in the accumulation of fluorescent calcein in both the Golgi and the mitochondria. We propose that LeMDR1 substrates are pumped into secretory compartments and exported from the parasite by exocytosis. Accumulation of MDR substrates in the ER can result in alternative transport to the mitochondrion, explaining the reciprocal sensitivity of drug‐resistant Leishmania to vinblastine and rhodamine 123.