Skip to main content
NCBI home page
Biochemical Journal logoLink to Biochemical Journal
. 1993 Apr 1;291(Pt 1):187–191. doi: 10.1042/bj2910187

Receptor-mediated endocytosis of retinol-binding protein by liver parenchymal cells: interference by radioactive iodination.

L Malaba 1, G M Kindberg 1, K R Norum 1, T Berg 1, R Blomhoff 1
PMCID: PMC1132500  PMID: 8471038

Abstract

Retinol-binding protein (RBP) was iodinated directly by radio-iodine substitution on the tyrosyl residues by the sodium hypochlorite (NaOCl) or the Enzymobead (EB) methods, or indirectly by linkage of 125I-tyramine-cellobiose (TC) or 125I-N-succinimidyl-3-(4- hydroxyphenyl)propionic acid ester (SHPP) adduct on to free amino residues of RBP. Binding, uptake and degradation of iodinated RBP were studied in isolated rat and rabbit liver parenchymal cells. The amount of ligand bound to cells at 4 degrees C was dependent on the type of labelling, in that the 125I-TC ligand was bound to a lesser extent than NaClO-labelled 125I-RBP, EB-labelled 125I-RBP and 125I-SHPP-RBP. At 37 degrees C, the 125I-SHPP-RBP and the EB-labelled 125I-RBP became cell-associated more rapidly than the other two ligands. The higher cell association at 37 degrees C than at 4 degrees C suggests that internalization of the ligand occurred at the higher temperature. The degradation of the ligands was also different. The EB-labelled 125I-RBP, the 125I-TC-RBP and the 125I-SHPP-RBP showed an apparent lag phase before a steady increase in acid-soluble radioactivity was observed. Much less of EB-labelled 125I-RBP and 125I-TC-RBP were degraded (about 6%) than of the other two ligands (about 16%) after 120 min. About 50% of the acid-soluble radioactivity in these experiments could be accounted for by degradation in the medium, suggesting that about half of the degradation observed was intracellular. The present study therefore shows that the different labelling techniques yield varying estimates of the cellular handling of RBP. In addition, a rapid release of RBP was observed in experiments where cells were pulsed with radioactive RBP at 4 degrees C, washed and incubated further at 37 degrees C. Between 50% and 70% was released after 5 min of incubation. By increasing the temperature during the pulse to 37 degrees C, or by lowering the temperature during the chase to 4 degrees C, much less RBP was released from the cells. These data suggest that the release process represents recycling of internalized ligand from an early endosome.

Full text

PDF
187

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Blomhoff R., Berg T. Isolation and cultivation of rat liver stellate cells. Methods Enzymol. 1990;190:58–71. doi: 10.1016/0076-6879(90)90009-p. [DOI] [PubMed] [Google Scholar]
  2. Blomhoff R., Green M. H., Berg T., Norum K. R. Transport and storage of vitamin A. Science. 1990 Oct 19;250(4979):399–404. doi: 10.1126/science.2218545. [DOI] [PubMed] [Google Scholar]
  3. Blomhoff R., Green M. H., Green J. B., Berg T., Norum K. R. Vitamin A metabolism: new perspectives on absorption, transport, and storage. Physiol Rev. 1991 Oct;71(4):951–990. doi: 10.1152/physrev.1991.71.4.951. [DOI] [PubMed] [Google Scholar]
  4. Blomhoff R., Norum K. R., Berg T. Hepatic uptake of [3H]retinol bound to the serum retinol binding protein involves both parenchymal and perisinusoidal stellate cells. J Biol Chem. 1985 Nov 5;260(25):13571–13575. [PubMed] [Google Scholar]
  5. Bolton A. E., Hunter W. M. The labelling of proteins to high specific radioactivities by conjugation to a 125I-containing acylating agent. Biochem J. 1973 Jul;133(3):529–539. doi: 10.1042/bj1330529. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Creek K. E., Silverman-Jones C. S., De Luca L. M. Comparison of the uptake and metabolism of retinol delivered to primary mouse keratinocytes either free or bound to rat serum retinol-binding protein. J Invest Dermatol. 1989 Feb;92(2):283–289. doi: 10.1111/1523-1747.ep12276867. [DOI] [PubMed] [Google Scholar]
  7. Gjøen T., Bjerkelund T., Blomhoff H. K., Norum K. R., Berg T., Blomhoff R. Liver takes up retinol-binding protein from plasma. J Biol Chem. 1987 Aug 15;262(23):10926–10930. [PubMed] [Google Scholar]
  8. Goldstein J. L., Brown M. S. The low-density lipoprotein pathway and its relation to atherosclerosis. Annu Rev Biochem. 1977;46:897–930. doi: 10.1146/annurev.bi.46.070177.004341. [DOI] [PubMed] [Google Scholar]
  9. Green M. H., Uhl L., Green J. B. A multicompartmental model of vitamin A kinetics in rats with marginal liver vitamin A stores. J Lipid Res. 1985 Jul;26(7):806–818. [PubMed] [Google Scholar]
  10. HUNTER W. M., GREENWOOD F. C. Preparation of iodine-131 labelled human growth hormone of high specific activity. Nature. 1962 May 5;194:495–496. doi: 10.1038/194495a0. [DOI] [PubMed] [Google Scholar]
  11. Heller J. Characterization of bovine plasma retinol-binding protein and evidence for lack of binding between it and other bovine plasma proteins. J Biol Chem. 1975 Aug 25;250(16):6549–6554. [PubMed] [Google Scholar]
  12. Heller M., Bok D. A specific receptor for retinol binding protein as detected by the binding of human and bovine retinol binding protein to pigment epithelial cells. Am J Ophthalmol. 1976 Jan;81(1):93–97. doi: 10.1016/0002-9394(76)90198-7. [DOI] [PubMed] [Google Scholar]
  13. Jones P. G., Heller J. Specific binding of fluorescein labelled serum retinol-binding protein to its cell surface receptor in isolated, purified bovine pigment epithelial cells. Exp Eye Res. 1980 May;30(5):489–499. doi: 10.1016/0014-4835(80)90033-0. [DOI] [PubMed] [Google Scholar]
  14. Kanai M., Raz A., Goodman D. S. Retinol-binding protein: the transport protein for vitamin A in human plasma. J Clin Invest. 1968 Sep;47(9):2025–2044. doi: 10.1172/JCI105889. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. McGuire B. W., Orgebin-Crist M. C., Chytil F. Autoradiographic localization of serum retinol-binding protein in rat testis. Endocrinology. 1981 Feb;108(2):658–667. doi: 10.1210/endo-108-2-658. [DOI] [PubMed] [Google Scholar]
  16. Ottonello S., Petrucco S., Maraini G. Vitamin A uptake from retinol-binding protein in a cell-free system from pigment epithelial cells of bovine retina. Retinol transfer from plasma retinol-binding protein to cytoplasmic retinol-binding protein with retinyl-ester formation as the intermediate step. J Biol Chem. 1987 Mar 25;262(9):3975–3981. [PubMed] [Google Scholar]
  17. Pittman R. C., Carew T. E., Glass C. K., Green S. R., Taylor C. A., Jr, Attie A. D. A radioiodinated, intracellularly trapped ligand for determining the sites of plasma protein degradation in vivo. Biochem J. 1983 Jun 15;212(3):791–800. doi: 10.1042/bj2120791. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Rask L., Peterson P. A. In vitro uptake of vitamin A from the retinol-binding plasma protein to mucosal epithelial cells from the monkey's small intestine. J Biol Chem. 1976 Oct 25;251(20):6360–6366. [PubMed] [Google Scholar]
  19. Redshaw M. R., Lynch S. S. An improved method for the preparation of iodinated antigens for radioimmunoassay. J Endocrinol. 1974 Mar;60(3):527–528. doi: 10.1677/joe.0.0600527. [DOI] [PubMed] [Google Scholar]
  20. Seglen P. O. Preparation of isolated rat liver cells. Methods Cell Biol. 1976;13:29–83. doi: 10.1016/s0091-679x(08)61797-5. [DOI] [PubMed] [Google Scholar]
  21. Senoo H., Stang E., Nilsson A., Kindberg G. M., Berg T., Roos N., Norum K. R., Blomhoff R. Internalization of retinol-binding protein in parenchymal and stellate cells of rat liver. J Lipid Res. 1990 Jul;31(7):1229–1239. [PubMed] [Google Scholar]
  22. Sivaprasadarao A., Findlay J. B. The interaction of retinol-binding protein with its plasma-membrane receptor. Biochem J. 1988 Oct 15;255(2):561–569. [PMC free article] [PubMed] [Google Scholar]
  23. Sivaprasadarao A., Findlay J. B. The mechanism of uptake of retinol by plasma-membrane vesicles. Biochem J. 1988 Oct 15;255(2):571–579. [PMC free article] [PubMed] [Google Scholar]
  24. Tolleshaug H., Berg T., Nilsson M., Norum K. R. Uptake and degradation of 125I-labelled asialo-fetuin by isolated rat hepatocytes. Biochim Biophys Acta. 1977 Aug 25;499(1):73–84. doi: 10.1016/0304-4165(77)90230-6. [DOI] [PubMed] [Google Scholar]
  25. Vahlquist A., Nilsson S. F., Peterson P. A. Isolation of the human retinol binding protein by affinity chromatography. Eur J Biochem. 1971 May 28;20(2):160–168. doi: 10.1111/j.1432-1033.1971.tb01374.x. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES