Establishment of polarized endocytosis in differentiable intestinal HT29-18 subclones.
Subclones of the HT29-18 clone, derived from a human adenocarcinoma, are able to acquire an enterocyte-like phenotype depending on the culture conditions. To investigate fluid-phase and receptor-mediated endocytosis in the polarized subclone HT29-18-C1, we established culture conditions that allowed cell growth on permeable supports. HT29-18-C1 monolayers had an electrical resistance of 43 ohms.cm2 and developed a transepithelial potential of about 2 mV. Transferrin receptors were uniformly distributed on the entire cell surface of undifferentiated HT29-18 cells but were located on the basolateral membrane of differentiated cells. Transferrin had a high affinity (Kd = 2.5 x 10(-9) M) for its receptor independent of the state of differentiation. The number of transferrin receptors and the mRNA amounts encoding them were comparable in the undifferentiated and differentiated HT29-18 cells. Transferrin was quickly internalized and recycled back to the cell surface of undifferentiated HT29-18 cells. The same phenomenon also occurred in differentiated HT29-18 cells, but the receptors were limited to the basolateral membrane. In the presence of ammonium chloride, the process was slower but remained polarized. Fluid-phase uptake was also investigated with horseradish peroxidase (HRP) in differentiated HT29-18 C1 cells. HRP that was internalized in 1 hour from a given membrane domain preferentially recycled back to the same membrane domain. No significant accumulation of the enzyme in the late endosomes and lysosomes of the differentiated HT29-18-C1 cells was observed.
Godefroy
O
O
Département de Biologie Moléculaire, URA CNRS 1149, Institut Pasteur, Paris, France.
Huet
C
C
Ibarra
C
C
Dautry-Varsat
A
A
Louvard
D
D
D000230:Adenocarcinoma / Q000473:pathology*; D002454:Cell Differentiation; D002999:Clone Cells / Q000187:drug effects / Q000473:pathology; D003110:Colonic Neoplasms / Q000473:pathology*; D004553:Electric Conductivity; D004705:Endocytosis; D005947:Glucose / Q000494:pharmacology; D006735:Horseradish Peroxidase / Q000378:metabolism; D006801:Humans; D009363:Neoplasm Proteins / Q000378:metabolism; D011990:Receptors, Transferrin / Q000378:metabolism; D014168:Transferrin / Q000378:metabolism; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000473:pathology
D009363:Neoplasm Proteins; D011990:Receptors, Transferrin; D014168:Transferrin; D006735:Horseradish Peroxidase; D005947:Glucose