Cellular Retinoic Acid Binding Protein II Research Articles

Molecules from Nature: Modulating the Expression of Estrogen Receptor Genes in Breast Cancer Cells

Estrogen receptors (ERα and ERβ) belong to the nuclear receptor superfamily. They mediate the effects of estrogens by binding (as homodimers or heterodimers) either directly to DNA at their estrogen-response elements (EREs) or by protein-protein interactions with other transcription factors (i.e., AP-1, NF-kβ) bound to their cognate DNA sequences, thus regulating the transcription of estrogen-responsive genes [1]. The ER mediates estrogenic activity in a variety of organs, including those in the reproductive, cardiovascular, immune and central nervous systems. Estrogen provides neuroprotection against neurodegenerative diseases, including Parkinsons disease. Its effects may stem from interactions with neurons, astrocytes and microglia. Human breast cancer cell lines expressing the (ERα), all-trans-retinoic acid (ATRA) receptor α (RARα) and cellular retinoic acid binding protein II (CRABPII) genes are sensitive to ATRA-mediated growth inhibition, as well. Several naturally found polyphenols like, flavonoids, phytoestrogens, etc., interact with the ERα and ERβ, exhibit estrogenic/antiestrogenic activities, and may play protective roles in cancer, inflammation, heart disease and osteoporosis, etc., and other disease conditions. These molecules are considered

Ileal Bile Acid-binding Protein, Functionally Associated with the Farnesoid X Receptor or the Ileal Bile Acid Transporter, Regulates Bile Acid Activity in the Small Intestine

Bile acids secreted in the small intestine are reabsorbed in the ileum where they activate the nuclear farnesoid X receptor (FXR), which in turn stimulates expression of the ileal bile acid-binding protein (I-BABP). We first hypothesized that I-BABP may negatively regulate the FXR activity by competing for the ligands, bile acids. Reporter assays using stable HEK293 cell lines expressing I-BABP revealed that I-BABP enhances rather than attenuates FXR activity. In these cells I-BABP localizes predominantly in the cytosol and partially in the nucleus, a distribution that does not shift in response to FXR expression. In vitro binding assays reveal that recombinant I-BABP is able to bind 35S-labeled FXR and that chenodeoxycholic acid (CDCA) stimulates this interaction modestly. When FLAG-tagged FXR was expressed in stable cells, the FXR.I-BABP complex in the nuclear extracts was more efficiently immunoprecipitable with anti-FLAG antibodies in the presence of CDCA. These results indicate that I-BABP stimulates FXR activity through a mutual interaction augmented by bile acids. When stable cells were transfected with an expression plasmid of the ileal bile acid transporter 14(IBAT) essential for the reabsorption of conjugated bile acids, the C-labeled conjugated bile acid, glycocholic acid, was more efficiently imported via IBAT in the presence than absence of I-BABP, whereas no change was observed in 14C-labeled CDCA uptake, which is independent of IBAT. Immunofluorescent staining analysis revealed that these two proteins co-localize in the vicinity of the plasma membrane in stable cells. Taken together, the current data provide the first evidence that I-BABP is functionally associated with FXR and IBAT in the nucleus and on the membrane, respectively, stimulating FXR transcriptional activity and the conjugated bile acid uptake mediated by IBAT in the ileum.

A novel function of differentiation revealed by cDNA microarray profiling of p75 NTR-regulated gene expression

The expression of the p75 neurotrophin receptor (p75 NTR) is diminished in epithelial cells during progression of prostate cancer in vivo and in vitro. Previous studies have demonstrated a role for p75 NTR as a tumor suppressor in prostate growth. To better understand the molecular mechanism of p75 NTR on tumor suppression, we utilized a complementary deoxyribonucleic acid microarray composed of ∼6,000 human cancer-related genes to determine the gene expression pattern altered by re-introduction of p75 NTR into PC-3 prostate tumor cells. Comparison of the transcripts in the neo and p75 NTR-transfected cells revealed 52 differentially expressed genes, of which 21 were up-regulated and 31 were down-regulated in the presence of p75 NTR. Based on the known biological functions of the p75 NTR-regulated genes, we observed that p75 NTR modulated the expression of genes that are critically involved in the regulation of differentiation as well as cell adhesion, signal transduction, apoptosis, tumor cell invasion, and metastasis. Several differentially expressed genes identified by microarray were selected for confirmation using quantitative real-time polymerase chain reaction. Immunoblot analysis further confirmed increased cellular retinoic acid-binding protein I (CRABPI) and IGFBP5 protein levels and decreased level of PLAUR protein with increasing p75 NTR protein expression. As CRABPI was elevated far more than any other genes, we observed that the retinoids, all- trans retinoic acid and 9- cis retinoic acid, that bind CRABPI, promoted nitroblue tetrazolium-associated functional cell differentiation in p75 NTR PC-3 cells, but not in neo control PC-3 cells. Subsequent examination of the retinoic acid receptors (RARs) expression levels demonstrated an absence of RAR-β in the neo control cells and re-expression in the p75 NTR expressing cells, consistent with previous findings where RAR-β is believed to play a critical role as a tumor suppressor gene that is lost during de-differentiation of prostate epithelial cells. Whereas the RAR-α and -γ protein levels remained unchanged, retinoid X receptor (RXR)-α and -β also exhibited increasing protein levels with re-expression of the p75 NTR protein. Moreover, the ability of p75 NTR siRNA to knockdown levels of RAR-β, RXR-α, and RXR-β supports the specificity of the functional involvement of p75 NTR in differentiation. Hence, re-expression of the p75 NTR appears to partially reverse de-differentiation of prostate cancer cells by up-regulating the expression of CRABPI for localized sequestration of retinoids that are available to newly up-regulated RAR-β, RXR-α, and RXR-β.

Signal Sends Carrier to the Nucleus

The intracellular lipid-binding proteins (iLBPs) are thought to act as carriers that move small lipophilic compounds in the cell. Cellular retinoic acid binding protein-II (CRABP-II) is one such protein and helps augment the biological effects of retinoic acid to activate the type II nuclear receptor known as the retinoic acid receptor (RAR). CRABP-II is thought to interact with RARs also and, in essence, to hand off the retinoic acid ligand to its receptor. RARs are localized in the nucleus, and CRABP-II is found in the cytoplasm but moves to the nucleus when it binds retinoic acid. However CRABP-II contains no classical nuclear localization signal, so it has been unclear how this translocation occurs. Sessier and Noy therefore examined the crystal structures of ligand-bound and -unbound CRABP-II for differences that might account for the translocation. Comparison of combined electrostatic surface potentials across the molecular surfaces of the two proteins revealed a region that was neutrally charged in the unbound protein but became basic or positively charged when retinoic acid was bound. Furthermore, it appeared that reorganization of three key basic amino acid residues brought them into an alignment that approximates that seen in the primary sequence of a classical nuclear localization sequence. Mutations in these residues prevented nuclear localization of CRABP-II, its interaction with importin alpha (a component of the nuclear translocation machinery), and its ability to enhance transcription mediated by RAR. Thus CRABP-II appears to contain a ligand-regulated switch that determines its subcellular localization. R. J. Sessler, N. Noy, A ligand-activated nuclear localization signal in cellular retinoic acid binding protein-II. Mol. Cell 18 , 343-353 (2005). [Online Journal]

Expression of estrogen receptor α, retinoic acid receptor α and cellular retinoic acid binding protein II genes is coordinately regulated in human breast cancer cells

Human breast cancer cell lines expressing the estrogen receptor alpha (ERalpha), all-trans-retinoic acid (ATRA) receptor alpha (RARalpha) and cellular retinoic acid binding protein II (CRABPII) genes are sensitive to ATRA-mediated growth inhibition. To study the relationship among ERalpha, RARalpha and CRABPII expression, the protein levels of each member were compared in five breast cancer cell lines (T47D, MCF-7, ZR-75-1, Hs587 T and MDA-MB-231 cells) and two immortalized nontumorigenic breast epithelial cell lines (MTSV1.7 and MCF-10A). ERalpha, RARalpha and CRABPII proteins were detected in T47D, MCF-7 and ZR-75-1 cells but not in other tested cell lines. RARalpha and CRABPII proteins were either reduced or undetectable in T47D/C4:2W and MCF-7/ADR cells with lost expression of ERalpha. Estradiol increased and anti-estrogens (tamoxifen and ICI 164,384) downregulated the expression of both RARalpha and CRABPII proteins in T47D and MCF-7 cells. RARalpha antagonist Ro-41-5253 inhibited CRABPII expression, but not RARalpha expression in estradiol-treated T47D and MCF-7 cells. Suppression of ERalpha by small interfering RNA (siRNA) reduced RARalpha and CRABPII gene expression and siRNA suppression of RARalpha reduced CRABPII expression while having no effect on ERalpha in T47D cells. Transient transfection of either RARalpha or ERalpha expression vectors increased CRABPII expression in MDA-MB-231 cells but only RARalpha, not ERalpha, activated hCRABPII promoter reporter. These results indicate that there is a gene activation pathway in which ERalpha drives RARalpha transcription and RARalpha drives CRABPII transcription in ERalpha-positive human breast cancer cells.

A Ligand-Activated Nuclear Localization Signal in Cellular Retinoic Acid Binding Protein-II

Primary sequences of proteins often contain motifs that serve as "signatures" for subcellular targeting, such as a nuclear localization signal (NLS). However, many nuclear proteins do not harbor a recognizable NLS, and the pathways that mediate their nuclear translocation are unknown. This work focuses on CRABP-II, a cytosolic protein that moves to the nucleus upon binding of retinoic acid. While CRABP-II does not contain an NLS in its primary sequence, such a motif could be recognized in the protein's tertiary structure. We map the retinoic acid-induced structural rearrangements that result in the presence of this NLS in holo- but not apo-CRABP-II. The signal, whose three-dimensional configuration aligns strikingly well with a "classical" NLS, mediates ligand-induced association of CRABP-II with importin alpha and is critical for nuclear localization of the protein. The ligand-controlled NLS "switch" of CRABP-II may represent a general mechanism for posttranslational regulation of the subcellular distribution of a protein.

Comprehensive survey of carapacial ridge‐specific genes in turtle implies co‐option of some regulatory genes in carapace evolution

The turtle shell is an evolutionary novelty in which the developmental pattern of the ribs is radically modified. In contrast to those of other amniotes, turtle ribs grow laterally into the dorsal dermis to form a carapace. The lateral margin of carapacial primordium is called the carapacial ridge (CR), and is thought to play an essential role in carapace patterning. To reveal the developmental mechanisms underlying this structure, we systematically screened for genes expressed specifically in the CR of the Chinese soft-shelled turtle, Pelodiscus sinensis, using microbead-based differential cDNA analysis and real-time reverse transcription-polymerase chain reaction. We identified orthologs of Sp5, cellular retinoic acid-binding protein-I (CRABP-I), adenomatous polyposis coli down-regulated 1 (APCDD1), and lymphoid enhancer-binding factor-1 (LEF-1). Although these genes are conserved throughout the major vertebrate lineages, comparison of their expression patterns with those in chicken and mouse indicated that these genes have acquired de novo expression in the CR in the turtle lineage. In association with the expression of LEF-1, the nuclear localization of beta-catenin protein was detected in the CR ectoderm, suggesting that the canonical Wnt signaling triggers carapace development. These findings indicate that the acquisition of the turtle shell did not involve the creation of novel genes, but was based on the co-option of pre-existing genes.

Mutation of Cellular Retinoic Acid-Binding Protein-II (Crabp-II) Does Not Account for Resistance to All-Trans Retinoic Acid (ATRA) in Relapse Acute Promyelocytic Leukemia (APL).

An increase in CRABP-II has frequently been invoked as a cause of resistance to ATRA in APL due to cytoplasmic sequestration and catabolism of ATRA. However, recent evidence indicates that CRABP-II has a positive rather than a negative role in ATRA activity by facilitating delivery of ATRA to retinoic acid receptor-alpha (RARα) associated with ATRA target genes in the cell nucleus or/and by serving as a co-activator of RARα-regulated transcription. This implies that if CRABP-II has a role in the development of ATRA resistance in APL, this would more likely occur through a deficiency rather than from an increase in CRABP-II. We previously reported that CRABP-II is constitutively expressed at similar levels in pretreatment and relapse APL cells (Zhou, et al, Cancer Res 58, 5770, 1998), suggesting that putative CRABP-II deficiency is not due to the loss of CRABP-II expression. To investigate the alternative possibility that CRABP-II deficiency might arise through inactivating mutations, we sequenced the entire coding region of CRABP-II from 18 cases of APL who had relapsed from prior ATRA-containing treatment regimens. In 8 cases RNA was transcribed by reverse transcriptase to cDNA and amplified by polymerase chain reaction (PCR), using primers anchored in the 5′ and 3′ untranslated region of mRNA; in 10 cases, genomic DNA was PCR amplified for sequence analysis, using primers anchored in the introns between the 4 exons of the gene. No CRABP-II mutations were identified. The samples tested included 11 first-relapse cases, 2 of whom were refractory to reinduction therapy with intravenous liposomal-ATRA, and 7 multiple relapse cases, all of whom were clinically refractory to ATRA and had, additionally, relapsed from arsenic trioxide therapy. Also, no mutations were found in 3 APL patients who had relapsed from chemotherapy-induced remissions or in 3 APL cell lines (NB4, UF-1 and AP-1060). Two heterozygous base substitutions were incidentally identified in CRABP-II intron 2 in a chemotherapy-only treated patient. These results indicate that CRABP-II mutations rarely, if ever, contribute to ATRA-resistance or disease relapse in APL.

42 IL-1 Decreases The Production of Cellular Retinoic Acid Binding Protein-I (Crabp-I) in The Lungs of Transgenic Mice: A Possible Link Between Inflammation and The Retinoic Acid Pathway in The Pathogenesis of Bronchopulmonary Dysplasia (BPD)

Background: Pulmonary inflammation, increased production of the inflammatory cytokine IL-1, and vitamin A deficiency are associated with the development of BPD. In order to determine the mechanisms by which IL-1 influences lung development, we have developed a transgenic mouse overexpressing IL-1 in the lung epithelium in an externally regulatable manner. Lung histology of these mice after perinatal induction of IL-1 production shows impaired alveolar and vascular development of the lung, similar to the histological characteristics of BPD. Retinoic acid (RA), one of the most biologically active derivatives of vitamin A, increases septation. We hypothesized that a mechanism by which IL-1 decreases septation, is inhibition of RA action. Cellular retinoic acid binding protein I (CRABP-I) is an intracellular protein that binds RA with high affinity. Lack of CRABP-I results in decreased intracellular RA concentrations when the extracellular RA level is low. OBJECTIVE: To study CRABP-I mRNA expression and protein production in the lungs of fetal and newborn IL-1 overexpressing mice and their wild-type littermates.

The effect of cellular retinoic acid binding protein-I expression on the CYP26-mediated catabolism of all- trans retinoic acid and cell proliferation in head and neck squamous cell carcinoma

The aim of this study was to confirm if catabolism of all- trans retinoic acid (RA) is enhanced by type I cellular retinoic acid binding protein (CRABP-I) expression and to investigate the effect of this enhanced catabolism on cell proliferation of the head and neck squamous cell carcinoma (HNSCC) cell line, AMC-HN-7. We also analyzed the effects of CRABP-I on RA-induced retinoic acid receptor (RAR) activity. The expression of the CRABP-I in stably transfected AMC-HN-7 cell lines (HN7-BPIa and HN7-BPIb) resulted in a lower sensitivity to administered RA compared with that of controls in a clonogenic assay. HN7-BPIs cells showed an increased amount of polar metabolites of RA in thin-layer chromatography. The transcriptional activity of the reporter plasmid RARE(DR5)-tk-CAT after the treatment of RA was lesser in HN7-BPIs than in controls. These results suggest that the increased CYP26-mediated catabolism of RA by CRABP-I transfection might decrease the amount of RA that is accessible to the nuclear receptors and make HNSCC cells resistant to RA.

Replacement of proline with valine does not remove an apparent proline isomerization-dependent folding event in CRABP I.

Site-directed mutagenesis has frequently been used to replace proline with other amino acids in order to determine if proline isomerization is responsible for a slow phase during refolding. Replacement of Pro 85 with alanine in cellular retinoic acid binding protein I (CRABP-I) abolished the slowest refolding phase, suggesting that this phase is due to proline isomerization in the unfolded state. To further test this assumption, we mutated Pro 85 to valine, which is the conservative replacement in the two most closely related proteins in the family (cellular retinoic acid binding protein II and cellular retinol binding protein I). The mutant protein was about 1 kcal/mole more stable than wild type. Retinoic acid bound equally well to wild type and P85V-CRABP I, confirming the functional integrity of this mutation. The refolding and unfolding kinetics of the wild-type and mutant proteins were characterized by stopped flow fluorescence and circular dichroism. The mutant P85V protein refolded with three kinetic transitions, the same number as wild-type protein. This result conflicts with the P85A mutant, which lost the slowest refolding rate. The P85V mutation also lacked a kinetic unfolding intermediate found for wild-type protein. These data suggest that proline isomerization may not be responsible for the slowest folding phase of CRABP I. As such, the loss of a slow refolding phase upon mutation of a proline residue may not be diagnostic for proline isomerization effects on protein folding.

Identification of proteins differentially expressed during chondrogenesis of mesenchymal cells

We performed comparative proteome analysis of mesenchymal cells and chondrocytes to identify proteins differentially expressed during chondrogenesis. Nine such proteins were identified. Type II collagen, matrilin-1, carbonic anhydrase-II (CA-II), 3′-phosphoadenosine 5′-phosphosulfate (PAPS) synthetase-2, and aldo-keto reductase were increased during chondrogenesis, whereas cellular retinoic acid binding protein-I (CRABP-I), CRABP-II, cytoplasmic type 5 actin, and fatty acid binding protein were decreased or almost disappeared. Expression of type II collagen, matrilin-1, PAPS synthetase-2, and CA-II was regulated by extracellular signal-regulated protein kinase, protein kinase C, and p38 kinase, signaling molecules known to regulate chondrogenesis.

Differential expression of CRABP-II in fibroblasts derived from dermis and subcutaneous fat

We have shown previously that fibroblasts derived from fat or dermal tissue differ in their functional properties, such as proliferation rate and contractile properties. To study these differences further, two-dimensional electrophoresis (2D PAGE) was performed on proteins isolated from cultured subcutaneous fat and dermal fibroblasts. The 2D gels were screened for proteins that were differentially expressed in all donors ( n=5). Five protein spots were subjected to further analysis by mass spectrometry. Two proteins could be identified: brain acid soluble protein 1 (BASP1) and cellular retinoic acid binding protein-II (CRABP-II). CRABP-II is of interest in terms of re-epithelialisation and was clearly expressed in dermal fibroblasts but not in fat fibroblasts. Real time PCR was performed to confirm the 2D data on CRABP-II. The CRABP-II mRNA level was significantly increased in dermal tissue and cultured dermal fibroblasts compared to fat tissue and cultured fat-derived fibroblasts, respectively. The mode of action of CRABP-II in skin is to mediate retinoic acid activity. Retinoic acid is known to inhibit migration and to stimulate differentiation of keratinocytes. The expression of CRABP-II by dermal fibroblasts implicates a role for these fibroblasts in wound re-epithelialisation, in contrast to subcutaneous fat-derived fibroblasts.

All-trans retinoic acid- and N-(4-hydroxyphenil)-retinamide-induced growth arrest and apoptosis in orbital fibroblasts in Graves’ disease

In this study, we evaluated by reverse transcription-polymerase chain reaction (RT-PCR) the expression pattern of retinoic acid receptors (RAR) α, β, and γ and cellular retinoic acid binding protein-I (CRBP-I) genes in 12 primary cultures of fibroblasts (F) from orbital tissue of Graves’ ophthalmopathy (GO) patients. We also studied the in vitro effects of all-trans retinoic acid (RA) and N-(4-hydroxyphenil)-retinamide (4HPR), a less toxic and better tolerated synthetic derivative of RA, on cell morphology, growth, apoptosis, and cyclic adenosine monophosphate (cAMP) accumulation. All primary cultures expressed RAR α, β, γ, and CRBP-I. FGO treated with RA and 4HPR (10−7 mol/L) presented morphologic changes and significantly inhibited cell growth after 72 hours. At 96 hours of drug exposure, apoptosis was detected in 15% and 50% of RA- and 4HPR (10−7 mol/L)-treated cells, and p53 protein increased in cell lysates. 4HPR induced a 70% decrease of Bcl-2 protein. After 30 minutes of RA and 4HPR (10−7 mol/L) exposure, a 20% decrease of basal cAMP accumulation was seen, and forskolin cAMP-induced increase was abolished. The expression of RAR α, β, γ, and CRBP-I in primary cultures of FGO indicates that they are targets for retinoids. Moreover, we show that RA and 4HPR are able to induce morphologic changes, inhibition of cell growth, and apoptosis in FGO exerting their effects through RAR-modulated pathways. The rapid inhibition of cAMP accumulation indicates that a novel nonclassic retinoid pathway may also be involved. Finally, the potent in vitro effects of 4HPR, a retinoid derivative with fewer adverse reactions in vivo, could justify further investigations on a clinical application of retinoids in GO.

Two novel single nucleotide polymorphisms in the promoter of the Cellular Retinoic Acid Binding Protein II gene ( CRABP-II)

The cellular retinoic acid binding protein-II (CRABP-II) is an intracellular protein involved in the transmission of the vitamin A-derived signal which regulates genes responsible for lipid metabolism and adipocyte differentiation. Cellular Retinoic Acid Binding Protein-II gene ( CRABP-II) (GDB 134819) is located on chromosome 1q21-23 and this region has been linked with related disorders such as Familial Combined Hyperlipidemia (FCHL), type 2 Diabetes Mellitus, and Lipodystrophy. In this context we hypothesized that CRABP-II is an interesting protein and aimed to provide genetic markers for future studies. In order to do that, we screened the promoter and the entire coding regions for mutations in 53 patients diagnosed with FCHL and 89 normolipidemic controls. Two new single nucleotide polymorphisms (SNPs) were identified in the promoter region a C to A change at position −515 and a T to C substitution at position −394, the latter creating a binding site for SP1. The change −515C>A was identified in a FCHL patient whereas the −394T>C was found in 3 FCHL patients and 4 normolipidemic subjects. This report provides two new polymorphisms in CRABP-II, which can be used as genetic markers for future studies of association or linkage with diseases, particularly those associated with the metabolic syndrome.

Cyclin D3 Is a Cofactor of Retinoic Acid Receptors, Modulating Their Activity in the Presence of Cellular Retinoic Acid-binding Protein II

Ligand-induced transcription activation of retinoic acid (RA) target genes by nuclear receptors (retinoic acid (RAR) and retinoid X (RXR) receptors) depends on the recruitment of coactivators. We have previously demonstrated that the small 15-kDa cellular RA-binding protein II (CRABPII) is a coactivator present in the RA-dependent nuclear complex. As identifying cell-specific partners of CRABPII might help to understand the novel control of RA signaling, we performed a yeast two-hybrid screen of a hematopoietic HL-60 cDNA library using human CRABPII as bait and have subsequently identified human cyclin D3 as a partner of CRABPII. Cyclin D3 interacted with CRABPII in a ligand-independent manner and equally bound RAR alpha, but not RXR alpha, and only in the presence of RA. We further show that cyclin D3 positively modulated RA-mediated transcription through CRABPII. Therefore, cyclin D3 may be part of a ternary complex with CRABPII and RAR. Finally, we show that cyclin D3 expression paralleled HL-60 differentiation and arrest of cell growth. These findings led us to speculate that control of cell proliferation during induction of differentiation may directly involve, at the transcriptional level, nuclear receptors, coactivators, and proteins of the cell cycle in a cell- and nuclear receptor-specific manner.

Topical retinoic acid alters the expression of cellular retinoic acid-binding protein-I and cellular retinoic acid-binding protein-II in non-lesional but not lesional psoriatic skin.

Therapeutic retinoids have profound effects on psoriatic skin pathology but their interactions with various retinoid-binding proteins in lesional vs non-lesional skin have not been investigated. Using quantitative real-time PCR the mRNA expression of cellular retinol-binding protein I (CRBPI) and retinoic acid-binding protein I/II (CRABPI/CRABPII) was studied in psoriatic and healthy control (=normal) skin after 4 days of occlusive RA/vehicle treatment (n=6). Untreated psoriatic lesions showed a markedly elevated CRABPII/CRABPI ratio, while the CRBPI level was reduced in lesional and non-lesional skin as compared to normal skin. In RA-treated normal and non-lesional skin, the mRNA expression of CRBPI was unaltered while that of CRABPI and CRABPII was reduced by approximately 80% and increased approximately 5-fold, respectively, as compared to vehicle-treated skin. In contrast, lesional skin exposed to RA showed an almost 90% increase in CRBPI transcripts but unaltered expression of CRABPI and CRABPII, yet, the mRNA expression of several inflammatory mediators, e.g. inducible nitric oxide synthase, interferon-gamma and interleukin-1beta, was clearly reduced. Immunohistochemistry localized CRABPII to suprabasal keratinocytes in normal skin and revealed markedly elevated levels in lesional skin. RA treatment induced CRABPII protein expression in normal and non-lesional skin, to similar levels as in untreated lesions. The results indicate that the effects of RA differ in normal/non-lesional psoriatic skin and lesional skin. Whether the high expression of CRABPII in psoriatic skin lesions is due to increased amounts of endogenous retinoids in lesional skin or reflects an abnormal regulation of the CRABPII gene in psoriasis remains to be studied.

Retinoid signalling and gene expression in neuroblastoma cells: RXR agonist and antagonist effects on CRABP-II and RARbeta expression.

9-cis Retinoic acid (RA) induces gene expression in neuroblastoma cells more effectively and with different kinetics than other RA isomers, and could be acting in part through Retinoid X Receptors (RXRs). The aim of this study was to characterise the effects of an RXR agonist and RXR homodimer antagonist on the induction of cellular RA binding protein II (CRABP-II) and RA receptor-beta (RARbeta) in neuroblastoma cells in response to different retinoids. The RXR agonist, LDG1069, was as effective as all-trans RA in inducing gene expression, but less effective than 9-cis RA. The RXR-homodimer antagonist, LG100754, inhibited the induction of CRABP-II mRNA in SH-SY5Y neuroblastoma cells by 9-cis RA or the RXR-specific agonist LGD1069, but had no effect when used with all-trans RA. Conversely, LG100754 did not inhibit induction of RARbeta mRNA by 9-cis or all-trans RA, or by LGD1069. RAR- and RXR-specific ligands used together induced CRABP-II and RARbeta as effectively as 9-cis RA. These results demonstrate the value of combining RXR- and RAR-specific ligands to regulate RA-inducible gene expression. The possibility that RXR-homodimers mediate, in part, the induction of CRABP-II by 9-cis RA and RXR-specific ligands is discussed.

Upregulation of cellular retinoic acid-binding protein I expression by ethanol.

Acute and chronic ethanol ingestion cause embryopathy similar to that of hyper- or hypovitaminosis A. Experimental data have suggested interaction between vitamin A and alcohol signaling pathways at the level of metabolic interference, which ultimately affects the concentration of retinoic acid (RA) in animals. The present study was set up to examine the possible effects of alcohol on cellular RA binding protein I (CRABP-I) expression during embryonic development by using transgenic mouse embryos and P19 embryonal carcinoma cells as experimental models. It was found that expression of the mouse CRABP-I gene was elevated in developing embryos at mid-gestation stages as a result of ethanol consumption by the mothers. Specific elevation of this gene was detected in the limb bud and the gut. In the P19 model, the CRABP-I gene was directly upregulated by ethanol, which was not blocked by a protein synthesis inhibitor. Furthermore, the regulation of the CRABP-I gene by ethanol was mediated by the 5' upstream regulatory region of the CRABP-I gene promoter. A potential interaction of vitamin A and ethanol at the level of CRABP-I gene expression is discussed.

Expression of cellular retinoc acid-binding protein I is specific to neurons in adult transgenic mouse brain

Cellular retinoic acid binding protein I (CRABP-I) plays a role in retinoic acid (RA) metabolism or transport. This report shows specific neuronal expression of CRABP-I in adult transgenic mouse brain using CRABP-I promotor-driven lac-Z and neuron- and astrocyte-markers. Double staining indicates that CRABP-I is expressed in neurons and large cells (>12 μm) but to much lesser degree the astrocytes. CRABP-I-lac-Z + neurons were distributed throughout the brain, but in a very discreet pattern in each brain region. CRABP-I expression in specific populations of brain neurons suggests that RA is extensively metabolized in mature brains, mostly in neurons. Additionally, the genetic basis of its specific expression in these brain areas is located in the 5′ regulatory region of this gene.