Cellubrevin is present in the basolateral endocytic compartment of hepatocytes and follows the transcytotic pathway after IgA internalization.
The endocytic compartment of polarized cells is organized in basolateral and apical endosomes plus those endocytic structures specialized in recycling and transcytosis, which are still poorly characterized. The complexity of the various populations of endosomes has been demonstrated by the exquisite repertoire of endogenous proteins. In this study we examined the distribution of cellubrevin in the endocytic compartment of hepatocytes, since its intracellular location and function in polarized cells are largely unknown. Highly purified rat liver endosomes were isolated from estradiol-treated rats, and the early/sorting endosomal fraction was further subfractionated in a multistep sucrose density gradient, and studied. Analysis of dissected endosomal fractions showed that cellubrevin was located in early/sorting endosomes, with Rab4, annexins II and VI, and transferrin receptor, but in a specific subpopulation of these early endosomes with the same density range as pIgA and Raf-1. Interestingly, only in those isolated endosomal fractions, endosomes enriched in transcytotic structures (of livers loaded with IgA), the polymeric immunoglobulin receptor specifically co-immunoprecipitated with cellubrevin. In addition, confocal and immuno-electron microscopy identification of cellubrevin in tubular structures underneath the sinusoidal plasma membrane together with the re-organization of cellubrevin, in the endocytic compartment, after the IgA loading, strongly suggest the involvement of cellubrevin in the transcytosis of pIgA.
Calvo
M
M
Departament de Biologia Cel.lular, Institut de Investigacions Biomèdiques August Pi i Sunyer, Facultat de Medicina, Universitat de Barcelona, 08036 Barcelona, Spain.
Pol
A
A
Lu
A
A
Ortega
D
D
Pons
M
M
Blasi
J
J
Enrich
C
C
D000818:Animals; D017302:Annexins / Q000302:isolation & purification; D015415:Biomarkers; D002451:Cell Compartmentation; D002458:Cell Fractionation; D004705:Endocytosis; D011992:Endosomes / Q000737:chemistry* / Q000648:ultrastructure; D004958:Estradiol / Q000494:pharmacology; D005455:Fluorescent Antibody Technique; D007070:Immunoglobulin A / Q000378:metabolism*; D008099:Liver / Q000378:metabolism* / Q000648:ultrastructure; D008297:Male; D008565:Membrane Proteins / Q000302:isolation & purification* / Q000378:metabolism; D016253:Microscopy, Immunoelectron; D011233:Precipitin Tests; D051381:Rats; D017207:Rats, Sprague-Dawley; D019056:Receptors, Polymeric Immunoglobulin / Q000302:isolation & purification; D011990:Receptors, Transferrin / Q000302:isolation & purification; D051040:Vesicle-Associated Membrane Protein 3; D020695:rab4 GTP-Binding Proteins / Q000302:isolation & purification
D017302:Annexins; D015415:Biomarkers; D007070:Immunoglobulin A; D008565:Membrane Proteins; D019056:Receptors, Polymeric Immunoglobulin; D011990:Receptors, Transferrin; D051040:Vesicle-Associated Membrane Protein 3; D004958:Estradiol; D020695:rab4 GTP-Binding Proteins