Skip to main content
NCBI home page
Biochemical Journal logoLink to Biochemical Journal
. 1992 Oct 15;287(Pt 2):383–389. doi: 10.1042/bj2870383

Expression of CRABP-I and -II in human epidermal cells. Alteration of relative protein amounts is linked to the state of differentiation.

G Siegenthaler 1, I Tomatis 1, D Chatellard-Gruaz 1, S Jaconi 1, U Eriksson 1, J H Saurat 1
PMCID: PMC1133176  PMID: 1332671

Abstract

The physiological role of cellular retinoic acid-binding proteins (CRABPs) may be to influence the intracellular level of free retinoic acid in the cell. In the present study two isoforms of CRABP, CRABP-I and CRABP-II were partially characterized in various human Malpighian epithelia and in human cultured keratinocytes expressing various patterns of differentiation. We have developed a new sensitive radiobinding assay using a PAGE/autoradioblotting technique which effectively separates CRABP-I and CRABP-II. This method allows the simultaneous quantification of these proteins. We show that CRABP-I and -II have similar M(r) values (15,000), but differ in their dissociation constant towards retinoic acid (Kd of 16.6 nM and 50 nM respectively), in pI (4.86 and 5.13) and in their relative mobilities (RF) on PAGE under nondenaturating conditions (RF values 0.65 and 0.44). In addition, we show that CRABP-II is the major isoform expressed in human keratinocytes, in vivo as in vitro. Furthermore, we demonstrate that CRABP-II is actually the CRABP previously studied in epidermal cells by a PAGE assay (Siegenthaler & Saurat (1987) Eur. J Biochem. 166, 209-214) and whose levels are dramatically increased by retinoic acid and its analogues in human epidermis. Keratinocytes, in the absence of full terminal differentiation, as well as hyperplasia, such as cultured human differentiating keratinocytes, psoriatic plaques, and non-keratinized oral mucosa, contained high levels of CRABP-II. CRABP-I was not detected in cultured keratinocytes, whereas normal skin (at full terminal differentiation) expressed CRABP-I and CRABP-II at a ratio of approx. 1:1.4. This value was approx. 1:17 in lesional psoriatic skin and 1:8 in oral mucosa. These observations suggest that CRABP-I and -II are regulated differently in human keratinocytes. The sharp increases in CRABP-II levels are associated with an alteration in the differentiation programme, as well as with cell response to retinoic acid overload, whereas CRABP-I might be a marker for terminal differentiation.

Full text

PDF
383

Images in this article

386Fig. 3.
on p.386

Selected References

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

  1. Aström A., Pettersson U., Krust A., Chambon P., Voorhees J. J. Retinoic acid and synthetic analogs differentially activate retinoic acid receptor dependent transcription. Biochem Biophys Res Commun. 1990 Nov 30;173(1):339–345. doi: 10.1016/s0006-291x(05)81062-9. [DOI] [PubMed] [Google Scholar]
  2. Aström A., Tavakkol A., Pettersson U., Cromie M., Elder J. T., Voorhees J. J. Molecular cloning of two human cellular retinoic acid-binding proteins (CRABP). Retinoic acid-induced expression of CRABP-II but not CRABP-I in adult human skin in vivo and in skin fibroblasts in vitro. J Biol Chem. 1991 Sep 15;266(26):17662–17666. [PubMed] [Google Scholar]
  3. Bailey J. S., Siu C. H. Purification and partial characterization of a novel binding protein for retinoic acid from neonatal rat. J Biol Chem. 1988 Jul 5;263(19):9326–9332. [PubMed] [Google Scholar]
  4. Bailey J. S., Siu C. H. Unique tissue distribution of two distinct cellular retinoic acid binding proteins in neonatal and adult rat. Biochim Biophys Acta. 1990 Mar 26;1033(3):267–272. doi: 10.1016/0304-4165(90)90131-f. [DOI] [PubMed] [Google Scholar]
  5. Brand N., Petkovich M., Krust A., Chambon P., de Thé H., Marchio A., Tiollais P., Dejean A. Identification of a second human retinoic acid receptor. Nature. 1988 Apr 28;332(6167):850–853. doi: 10.1038/332850a0. [DOI] [PubMed] [Google Scholar]
  6. Busch C., Sakena P., Funa K., Nordlinder H., Eriksson U. Tissue distribution of cellular retinol-binding protein and cellular retinoic acid-binding protein: use of monospecific antibodies for immunohistochemistry and cRNA for in situ localization of mRNA. Methods Enzymol. 1990;189:315–324. doi: 10.1016/0076-6879(90)89303-y. [DOI] [PubMed] [Google Scholar]
  7. Chytil F., Ong D. E. Intracellular vitamin A--binding proteins. Annu Rev Nutr. 1987;7:321–335. doi: 10.1146/annurev.nu.07.070187.001541. [DOI] [PubMed] [Google Scholar]
  8. Cogan U., Kopelman M., Mokady S., Shinitzky M. Binding affinities of retinol and related compounds to retinol binding proteins. Eur J Biochem. 1976 May 17;65(1):71–78. doi: 10.1111/j.1432-1033.1976.tb10390.x. [DOI] [PubMed] [Google Scholar]
  9. Crettaz M., Baron A., Siegenthaler G., Hunziker W. Ligand specificities of recombinant retinoic acid receptors RAR alpha and RAR beta. Biochem J. 1990 Dec 1;272(2):391–397. doi: 10.1042/bj2720391. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Didierjean L., Durand B., Saurat J. H. Cellular retinoic acid-binding protein type 2 mRNA is overexpressed in human psoriatic skin as shown by in situ hybridization. Biochem Biophys Res Commun. 1991 Oct 15;180(1):204–208. doi: 10.1016/s0006-291x(05)81277-x. [DOI] [PubMed] [Google Scholar]
  11. Elder J. T., Fisher G. J., Zhang Q. Y., Eisen D., Krust A., Kastner P., Chambon P., Voorhees J. J. Retinoic acid receptor gene expression in human skin. J Invest Dermatol. 1991 Apr;96(4):425–433. doi: 10.1111/1523-1747.ep12469889. [DOI] [PubMed] [Google Scholar]
  12. Eriksson U., Hansson E., Nordlinder H., Busch C., Sundelin J., Peterson P. A. Quantitation and tissue localization of the cellular retinoic acid-binding protein. J Cell Physiol. 1987 Dec;133(3):482–490. doi: 10.1002/jcp.1041330308. [DOI] [PubMed] [Google Scholar]
  13. Eriksson U., Sundelin J., Rask L., Peterson P. A. The NH2-terminal amino acid sequence of cellular retinoic-acid binding protein from rat testis. FEBS Lett. 1981 Nov 30;135(1):70–72. doi: 10.1016/0014-5793(81)80945-3. [DOI] [PubMed] [Google Scholar]
  14. FELL H. B., MELLANBY E. Metaplasia produced in cultures of chick ectoderm by high vitamin A. J Physiol. 1953 Mar;119(4):470–488. doi: 10.1113/jphysiol.1953.sp004860. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Fiorella P. D., Napoli J. L. Expression of cellular retinoic acid binding protein (CRABP) in Escherichia coli. Characterization and evidence that holo-CRABP is a substrate in retinoic acid metabolism. J Biol Chem. 1991 Sep 5;266(25):16572–16579. [PubMed] [Google Scholar]
  16. Fuchs E. Epidermal differentiation: the bare essentials. J Cell Biol. 1990 Dec;111(6 Pt 2):2807–2814. doi: 10.1083/jcb.111.6.2807. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Fuchs E., Green H. Regulation of terminal differentiation of cultured human keratinocytes by vitamin A. Cell. 1981 Sep;25(3):617–625. doi: 10.1016/0092-8674(81)90169-0. [DOI] [PubMed] [Google Scholar]
  18. Giguère V., Lyn S., Yip P., Siu C. H., Amin S. Molecular cloning of cDNA encoding a second cellular retinoic acid-binding protein. Proc Natl Acad Sci U S A. 1990 Aug;87(16):6233–6237. doi: 10.1073/pnas.87.16.6233. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hirschel-Scholz S., Siegenthaler G., Saurat J. H. Isotretinoin differs from other synthetic retinoids in its modulation of human cellular retinoic acid binding protein (CRABP). Br J Dermatol. 1989 May;120(5):639–644. doi: 10.1111/j.1365-2133.1989.tb01349.x. [DOI] [PubMed] [Google Scholar]
  20. Kaikaus R. M., Bass N. M., Ockner R. K. Functions of fatty acid binding proteins. Experientia. 1990 Jun 15;46(6):617–630. doi: 10.1007/BF01939701. [DOI] [PubMed] [Google Scholar]
  21. Kopan R., Fuchs E. The use of retinoic acid to probe the relation between hyperproliferation-associated keratins and cell proliferation in normal and malignant epidermal cells. J Cell Biol. 1989 Jul;109(1):295–307. doi: 10.1083/jcb.109.1.295. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Krust A., Kastner P., Petkovich M., Zelent A., Chambon P. A third human retinoic acid receptor, hRAR-gamma. Proc Natl Acad Sci U S A. 1989 Jul;86(14):5310–5314. doi: 10.1073/pnas.86.14.5310. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  24. Lotan R. Effects of vitamin A and its analogs (retinoids) on normal and neoplastic cells. Biochim Biophys Acta. 1980 Mar 12;605(1):33–91. doi: 10.1016/0304-419x(80)90021-9. [DOI] [PubMed] [Google Scholar]
  25. Maden M., Ong D. E., Summerbell D., Chytil F., Hirst E. A. Cellular retinoic acid-binding protein and the role of retinoic acid in the development of the chick embryo. Dev Biol. 1989 Sep;135(1):124–132. doi: 10.1016/0012-1606(89)90163-2. [DOI] [PubMed] [Google Scholar]
  26. Maden M., Ong D. E., Summerbell D., Chytil F. Spatial distribution of cellular protein binding to retinoic acid in the chick limb bud. Nature. 1988 Oct 20;335(6192):733–735. doi: 10.1038/335733a0. [DOI] [PubMed] [Google Scholar]
  27. Ong D. E., Chytil F. Cellular retinoic acid-binding protein from rat testis. Purification and characterization. J Biol Chem. 1978 Jul 10;253(13):4551–4554. [PubMed] [Google Scholar]
  28. Rheinwald J. G., Green H. Serial cultivation of strains of human epidermal keratinocytes: the formation of keratinizing colonies from single cells. Cell. 1975 Nov;6(3):331–343. doi: 10.1016/s0092-8674(75)80001-8. [DOI] [PubMed] [Google Scholar]
  29. Siegenthaler G. Retinoic acid formation from retinol and retinal metabolism in epidermal cells. Methods Enzymol. 1990;189:530–536. doi: 10.1016/0076-6879(90)89332-c. [DOI] [PubMed] [Google Scholar]
  30. Siegenthaler G., Samson J., Bernard J. P., Fiore-Donno G., Saurat J. H. Retinoid-binding proteins in human oral mucosa. J Oral Pathol. 1988 Mar;17(3):106–112. doi: 10.1111/j.1600-0714.1988.tb01895.x. [DOI] [PubMed] [Google Scholar]
  31. Siegenthaler G., Saurat J. H. A slab gel electrophoresis technique for measurement of plasma retinol-binding protein, cellular retinol-binding and retinoic-acid-binding proteins in human skin. Eur J Biochem. 1987 Jul 1;166(1):209–214. doi: 10.1111/j.1432-1033.1987.tb13503.x. [DOI] [PubMed] [Google Scholar]
  32. Siegenthaler G., Saurat J. H., Hotz R., Camenzind M., Merot Y. Cellular retinoic acid- but not cellular retinol-binding protein is elevated in psoriatic plaques. J Invest Dermatol. 1986 Jan;86(1):42–45. doi: 10.1111/1523-1747.ep12283788. [DOI] [PubMed] [Google Scholar]
  33. Siegenthaler G., Saurat J. H., Ponec M. Retinol and retinal metabolism. Relationship to the state of differentiation of cultured human keratinocytes. Biochem J. 1990 Jun 1;268(2):371–378. doi: 10.1042/bj2680371. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Siegenthaler G., Saurat J. H., Ponec M. Terminal differentiation in cultured human keratinocytes is associated with increased levels of cellular retinoic acid-binding protein. Exp Cell Res. 1988 Sep;178(1):114–126. doi: 10.1016/0014-4827(88)90383-7. [DOI] [PubMed] [Google Scholar]
  35. Siegenthaler G., Saurat J. H. Retinoid binding proteins and human skin. Pharmacol Ther. 1989;40(1):45–54. doi: 10.1016/0163-7258(89)90073-9. [DOI] [PubMed] [Google Scholar]
  36. Siegenthaler G., Saurat J. H. Retinol-binding protein in human serum: conformational changes induced by retinoic acid binding. Biochem Biophys Res Commun. 1987 Mar 13;143(2):418–423. doi: 10.1016/0006-291x(87)91370-2. [DOI] [PubMed] [Google Scholar]
  37. Siegenthaler G., Saurat J. H. Therapy with a synthetic retinoid--(Ro 10-1670) etretin--increases the cellular retinoic acid-binding protein in nonlesional psoriatic skin. J Invest Dermatol. 1986 Jul;87(1):122–124. doi: 10.1111/1523-1747.ep12523628. [DOI] [PubMed] [Google Scholar]
  38. Stoner C. M., Gudas L. J. Mouse cellular retinoic acid binding protein: cloning, complementary DNA sequence, and messenger RNA expression during the retinoic acid-induced differentiation of F9 wild type and RA-3-10 mutant teratocarcinoma cells. Cancer Res. 1989 Mar 15;49(6):1497–1504. [PubMed] [Google Scholar]
  39. Strickland S., Sawey M. J. Studies on the effect of retinoids on the differentiation of teratocarcinoma stem cells in vitro and in vivo. Dev Biol. 1980 Jul;78(1):76–85. doi: 10.1016/0012-1606(80)90319-x. [DOI] [PubMed] [Google Scholar]
  40. Wolf G. Multiple functions of vitamin A. Physiol Rev. 1984 Jul;64(3):873–937. doi: 10.1152/physrev.1984.64.3.873. [DOI] [PubMed] [Google Scholar]

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

RESOURCES