The targeting of lysosomal enzymes to their final destination is
directed by a series of protein and recognition signals. Although,
Giardia lamblia lacks a recognized Golgi apparatus or a typical
endosome/lysosome system, it might have a specific and conserved mechanism by means the soluble hydrolases are sorted. In this work, we describe the presence of a Vps10p -like receptor and its function in sorting of lysosomal enzymes. Bioinformatics and proteinase K assays allowed us to determine that the gVps10p receptor is a type I transmembrana protein with a WD40 motif in the luminal portion and a cytoplasmic tyrosine motif (YQII). Immunoblot assay showed a ?60 kDa but also a ?120 kDa band, possibly related to dimerization. IFA and confocal microscopy of trophozoites expressing gVps10p- HA fusion protein, showed that it localize around the nuclei colocalizing with the endoplasmic reticulum marker BIP (Immunoglobulin Binding Protein). Co-expression of gVPS10p- HA and gAcPh-V5 fusion proteins showed colocalization around nuclei and in the lysosomal peripheral vacuoles. On other hand, gVps10p-YQII-HA fusion protein, lacking the cytoplasmic tyrosine motif, showed a different subcelular distribution possibly related with changes in protein trafficking. This work will contribute to understand how the endosomal/lysosomal pathway works in the early divergent parasite G. lamblia.
endosome/lysosome system, it might have a specific and conserved
mechanism by means the soluble hydrolases are sorted. In this work,
we describe the presence of a Vps10p -like receptor and its function
in sorting of lysosomal enzymes. Bioinformatics and proteinase K
assays allowed us to determine that the gVps10p receptor is a type I
transmembrana protein with a WD40 motif in the luminal portion
and a cytoplasmic tyrosine motif (YQII). Immunoblot assay showed
a ?60 kDa but also a ?120 kDa band, possibly related to dimerization.
IFA and confocal microscopy of trophozoites expressing gVps10p-
HA fusion protein, showed that it localize around the nuclei
colocalizing with the endoplasmic reticulum marker BIP
(Immunoglobulin Binding Protein). Co-expression of gVPS10p-
HA and gAcPh-V5 fusion proteins showed colocalization around
nuclei and in the lysosomal peripheral vacuoles. On other hand,
gVps10p-YQII-HA fusion protein, lacking the cytoplasmic tyrosine
motif, showed a different subcelular distribution possibly related
with changes in protein trafficking. This work will contribute to
understand how the endosomal/lysosomal pathway works in the
early divergent parasite G. lamblia.
lacks a recognized Golgi apparatus or a typicalendosome/lysosome system, it might have a specific and conserved
mechanism by means the soluble hydrolases are sorted. In this work,
we describe the presence of a Vps10p -like receptor and its function
in sorting of lysosomal enzymes. Bioinformatics and proteinase K
assays allowed us to determine that the gVps10p receptor is a type I
transmembrana protein with a WD40 motif in the luminal portion
and a cytoplasmic tyrosine motif (YQII). Immunoblot assay showed
a ?60 kDa but also a ?120 kDa band, possibly related to dimerization.
IFA and confocal microscopy of trophozoites expressing gVps10p-
HA fusion protein, showed that it localize around the nuclei
colocalizing with the endoplasmic reticulum marker BIP
(Immunoglobulin Binding Protein). Co-expression of gVPS10p-
HA and gAcPh-V5 fusion proteins showed colocalization around
nuclei and in the lysosomal peripheral vacuoles. On other hand,
gVps10p-YQII-HA fusion protein, lacking the cytoplasmic tyrosine
motif, showed a different subcelular distribution possibly related
with changes in protein trafficking. This work will contribute to
understand how the endosomal/lysosomal pathway works in the
early divergent parasite G. lamblia.
G. lamblia.