INIMEC - CONICET   05467
INSTITUTO DE INVESTIGACION MEDICA MERCEDES Y MARTIN FERREYRA
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
The traffic of soluble hydrolases is mediated by VPS10P-like receptor in Giardia lamblia.
Autor/es:
MIRAS S; RIVERO MR; QUIROGA R; ZAMPONI N; FELIZIANI C; ROPOLO AS; TOUZ MC
Reunión:
Congreso; SAIB; 2011
Resumen:
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.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.G. lamblia.