title: Functional characterization of three aquaglyceroporins from Trypanosoma brucei in osmoregulation and glycerol transport.
abstract: Previous studies using bloodstream form Trypanosoma brucei have shown that glycerol transport in this parasite occurs via specific membrane proteins, namely a glycerol transporter and glycerol channels [1]. Later, we cloned, expressed and characterized the transport properties of all three aquaglyceroporins (AQP1-3) [2], which were found permeable for water, glycerol and other small uncharged solutes like dihydroxyacetone [3]. Here, we report on the cellular localization of TbAQP1 and TbAQP3 in bloodstream form trypanosomes. Indirect immunofluorescence analysis showed that TbAQP1 is exclusively localized in the flagellar membrane, whereas TbAQP3 was found in the plasma membrane.In addition, we analyzed the functions of all 3 AQPs, using an inducible inheritable double-stranded RNA interference methodology. All AQP knockdown cell lines were still able to survive hypo-osmotic stress conditions, except AQP2 knockdown parasites. Depleted TbAQP2 negatively impacted cell growth and the regulatory volume recovery, whereas AQP1 und 3 knockdown trypanosomes displayed phenotypes consistent with their localization in external membranes. A simultaneous knockdown of all 3 AQPs showed that the cells were able to substitute the missing glycerol uptake capability through a putative glycerol transporter.
authors.affiliation: Interfaculty Institute for Biochemistry, University of Tübingen, Tübingen, Germany.
authors.forename: Björn
authors.initials: B
authors.lastname: Bassarak
authors.forename: Nestor L
authors.initials: NL
authors.lastname: Uzcátegui
authors.forename: Caroline
authors.initials: C
authors.lastname: Schönfeld
authors.forename: Michael
authors.initials: M
authors.lastname: Duszenko
mesh_terms: D051397:Aquaglyceroporins / Q000032:analysis / Q000235:genetics / Q000502:physiology*; D051398:Aquaporin 1 / Q000032:analysis / Q000235:genetics / Q000502:physiology; D051399:Aquaporin 2 / Q000032:analysis / Q000235:genetics / Q000502:physiology; D051400:Aquaporin 3 / Q000032:analysis / Q000235:genetics / Q000502:physiology; D001692:Biological Transport; D002460:Cell Line; D019084:Fluorescent Antibody Technique, Indirect; D055785:Gene Knockdown Techniques; D005990:Glycerol / Q000378:metabolism* / Q000494:pharmacology; D015800:Protozoan Proteins / Q000032:analysis / Q000235:genetics / Q000502:physiology*; D011773:Pyruvates / Q000378:metabolism; D014346:Trypanosoma brucei brucei / Q000378:metabolism*; D014882:Water-Electrolyte Balance
chemical_list: D051397:Aquaglyceroporins; D051399:Aquaporin 2; D015800:Protozoan Proteins; D011773:Pyruvates; D051398:Aquaporin 1; D051400:Aquaporin 3; D005990:Glycerol