Leishmania RAB7: characterisation of terminal endocytic stages in an intracellular parasite.
Leishmania species are intracellular parasites that inhabit a parasitophorous vacuole (PV) within host macrophages and engage with the host endo-membrane network to avoid clearance from the cell. Intracellular Leishmania amastigotes exhibit a high degree of proteolytic/lysosomal activity that may assist degradation of MHC class II molecules and subsequent interruption of antigen presentation. As an aid to further analysis of the endosomal/lysosomal events that could facilitate this process, we have characterised a Leishmania homologue of the late endosomal marker, Rab7, thought to be involved in the terminal steps of endocytosis and lysosomal delivery. The Leishmania major Rab7 (LmRAB7) protein is expressed throughout the life-cycle, shows 73 and 64% identity to Trypanosoma cruzi and Trypanosoma brucei Rab7s (TcRAB7 and TbRAB7), respectively, and includes a kinetoplastid-specific insertion. The recombinant protein binds GTP and polyclonal antibodies raised against this antigen recognise structures in the region of the cell between the nucleus and kinetoplast. By immunoelectron microscopy of axenic amastigotes, Leishmania mexicana Rab7 (LmexRAB7) is found juxtaposed to and overlapping membrane structures labelled for the megasomal marker, cysteine proteinase B, confirming a late-endosomal/lysosomal localisation.
Denny
Paul W
PW
Wellcome Trust Laboratory for Molecular Parasitology and Centre for Molecular Microbiology and Infection, Department of Biological Sciences, Imperial College of Science, Technology and Medicine, London SW7 2AZ, UK.
Lewis
Sharon
S
Tempero
Jane E
JE
Goulding
David
D
Ivens
Alasdair C
AC
Field
Mark C
MC
Smith
Deborah F
DF
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D003001:Cloning, Molecular; D004705:Endocytosis; D011992:Endosomes / Q000469:parasitology; D017125:Genes, Protozoan; D006790:Host-Parasite Interactions; D018320:Leishmania major / Q000378:metabolism / Q000502:physiology*; D007894:Leishmania mexicana / Q000378:metabolism / Q000502:physiology*; D008018:Life Cycle Stages; D008247:Lysosomes / Q000469:parasitology; D008264:Macrophages / Q000469:parasitology; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000378:metabolism; D016415:Sequence Alignment; D020691:rab GTP-Binding Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D000090802:rab7 GTP-Binding Proteins
D011994:Recombinant Proteins; D000090802:rab7 GTP-Binding Proteins; C000716465:rab7 GTP-binding proteins, mouse; D020691:rab GTP-Binding Proteins