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

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.

A Heterocomplex Formed by the Calcium-binding Proteins MRP8 (S100A8) and MRP14 (S100A9) Binds Unsaturated Fatty Acids with High Affinity

“…Such a range in Ca 2ϩ concentrations is currently thought to be physiological. Taking in account that keratinocytes do not express detectable amounts of other FABPs than the epidermal type (29,30), which has no or low affinity for arachidonic and ␥-linolenic acid (6), our data suggest that these fatty acids mightbetransportedindifferentiatingkeratinocytesinacalciumdependent way by the highly expressed FA-p34. Whether more specific, yet unknown carriers for unsaturated fatty acids might co-exist in keratinocytes remains to be determined.…”

Increased retinoic acid levels through ablation of Cyp26b1 determine the processes of embryonic skin barrier formation and peridermal development

“…The retinoic-acid-binding proteins cellular retinoic acid binding protein II (CrabpII) and Rarb were upregulated 13.6-and 7.54-fold, respectively. Interestingly, when CRABPII was increased by RA treatment of human keratinocytes it was associated with absence of terminal differentiation (Siegenthaler et al, 1992). Lipid metabolism is known to be an integral part of barrier formation (Elias, 2005).…”

An Analysis of Retinoic Acid-induced Gene Expression and Metabolism in AB1 Embryonic Stem Cells

“…Higher (73,74). The amount of functional CRABPI and CRABPII protein expressed in a particular cell line can be visualized by virtue of its ability to bind to [ 3 H]RA.…”