Localization and activity of multidrug resistance protein 1 in the secretory pathway of Leishmania parasites

• Published in: Molecular microbiology Vol. 51; no. 6; pp. 1563 - 1575
• Main Authors: Dodge, Matthew A., Waller, Ross F., Chow, Larry M. C., Zaman, Muhammad M., Cotton, Leanne M., McConville, Malcolm J., Wirth, Dyann F.
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Science Ltd 01.03.2004; Blackwell Science; Blackwell Publishing Ltd
• Subjects: Animals; Antiprotozoal Agents - pharmacology; ATP Binding Cassette Transporter, Subfamily B, Member 1 - analysis; ATP Binding Cassette Transporter, Subfamily B, Member 1 - genetics; ATP Binding Cassette Transporter, Subfamily B, Member 1 - metabolism; ATP-Binding Cassette Transporters - metabolism; Biological and medical sciences; Biological Transport; Cell Line; Drug Resistance, Multiple - genetics; Endoplasmic Reticulum - metabolism; Fluoresceins - pharmacology; Fundamental and applied biological sciences. Psychology; Golgi Apparatus - metabolism; Green Fluorescent Proteins; Leishmania enriettii; Leishmania enriettii - drug effects; Leishmania enriettii - metabolism; Leishmania mexicana; Leishmania mexicana - drug effects; Leishmania mexicana - metabolism; Leishmaniasis - drug therapy; Luminescent Proteins - metabolism; Microbiology; Protozoan Proteins - analysis; Protozoan Proteins - metabolism; Recombinant Fusion Proteins - metabolism; Rhodamine 123 - pharmacology; Vinblastine - pharmacology; Infection; Kinetoplastida; Protozoa; Microbiology; Leishmania; Multiple resistance; Secretory protein; Parasitosis; Localization
• ISSN: 0950-382X; 1365-2958
• DOI: 10.1111/j.1365-2958.2003.03927.x

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.