Encyclopedia of Signaling Molecules

2018 Edition
| Editors: Sangdun Choi

Retinoic Acid Receptors (RARA, RARB, and RARC)

Reference work entry
DOI: https://doi.org/10.1007/978-3-319-67199-4_385

Synonyms

 NR1B1;  NR1B2;  NR1B3;  RARα;  RARβ;  RARγ

Historical Background

The retinoic acid receptors (RARs) are ligand-dependent transcription factors that belong to the NR1B subtype of the nuclear receptor (NRs) superfamily and have broad roles in development, cell growth and survival, vision, spermatogenesis, inflammation, and neural patterning. These receptors act in trans mainly as heterodimers with retinoid X receptors (RXRs). The actions of RARs are stimulated by the binding of cognate natural ligands (all trans retinoic acid and 9-cis retinoic acid) as well as a number of synthetic ligands. In the presence or absence of ligand, RAR/RXR heterodimers associate with retinoic acid DNA response elements (RAREs) present in the promoter or enhancer regions of target genes. When no ligand is present, corepressor proteins and histone deacetylases interact with the receptor DNA complexes and prevent transcription from occurring. When retinoid ligands are present, they bind and activate the RARs by initiating a conformation more favorable to the association of coactivator proteins and subsequent recruitment of histone acetyltransferases (HATs) and the components of the basal transcriptional machinery that initiate transcription of the RARE (Fig. 1). These consensus site DNA sequences generally consist of two directly repeated half sites of AGGTCA separated by two or five base pair spacers (DR2 or DR5 elements). Due to their broad action in diverse cell and tissue populations, RARs are essential signaling proteins for basic and clinical research.
Retinoic Acid Receptors (RARA, RARB, and RARC), Fig. 1

Activation model of RARs. In the presence or absence of ligand, RAR/RXR heterodimers associate with retinoic acid response elements (RAREs) present in the promoter or enhancer regions of target genes. When no ligand is present, corepressor proteins and histone deacetylases interact with the receptor DNA complexes and prevent transcription from occurring. When retinoid ligands are present, they bind and activate the RARs by initiating a conformation more favorable to the association of coactivator proteins and subsequent recruitment of histone acetyltransferases (HATs) and the components of the basal transcriptional machinery to initiate transcription

Role of RARs in Embryonic Development

Research from the 1950s implicated vitamin A deficiencies as a cause for a number of congenital malformations and defects observed in the development of animals. These results were known long before the discovery of retinoic acids as the most biologically active forms of vitamin A. Later research then investigated the role of retinoic acid in early embryonic development and in the early 1990s led to the discovery of the retinoic acid receptors (RARs). Since then much work has been done to determine the role that RARs play in embryonic development.

Genetic studies were conducted in the early 1990s to investigate the role of all three RAR isoforms in embryonic development by generating single and double RAR mutant mice. While mice with a mutation deleting only one RAR isoform contained certain developmental abnormalities, they were still viable, indicating a certain redundancy among isoforms (Mark et al. 2009). However, RAR double null mutants, containing genetic deletions of any two of the RAR isoforms, died in utero or at birth due to severe developmental deficits mainly in the vertebrae, brain, and limbs indicating the importance of the expression of the RARs on the formation of these structures. The same deficits were observed by a number of investigators where various components of retinoic acid signaling were knocked out such as retinaldehyde-synthesizing enzyme RDH10 (Sandell et al. 2007), RA-synthesizing enzyme RALDH2 (Halilagic et al. 2007), or RALDH3 (Dupe et al. 2003). In addition, treatment of wild-type animals with synthetic pan-specific RAR antagonists also produced the same defects (Kochhar et al. 1998; Wendling et al. 2001). This demonstrated that not only expression of RARs was essential for normal development but that the receptors required activation by retinoic acid ligands to mediate their important roles in developmental signaling programs.

Using combined strategies of selective RAR isoform knockouts and mutations in the RA signaling pathways, much has been learned about the crucial roles that RARs play in specific stages of organ and brain development. For instance, RARs are involved in formation of a number of limb structures and control the antero-posterior axis of the limbs (Dupe et al. 1999). Moreover, RAR-mediated RA signaling is responsible for both neurogenesis as well as the anterior-posterior patterning of the developing central nervous system through a complex mechanism of gene activation and repression by RARs (Maden 2002) (Fig. 2). Several studies have shown that RARs are required for the formation of a number of eye structures as well as histogenesis and physiological apoptosis in the retina. Moreover, it has been discovered that RARs play important roles in cardiac development, respiratory system development, as well as the formation of important structures in the kidneys and urogenital tract.
Retinoic Acid Receptors (RARA, RARB, and RARC), Fig. 2

RAR-mediated RA signaling mediates neurogenesis and anterior-posterior patterning of central nervous system. Diagram showing RAR-mediated up- and downregulation of genes involved in both neurogenesis and anterior-posterior which are vital in the development of neurons, spinal cord, and hindbrain

While much has been learned about the important roles played by the RARs in embryonic development, research is now shifting toward elucidating the roles played by these receptors during the postnatal development. To do this, new strategies have emerged to allow for the selective mutation of the retinoid receptors in specific cell types so as to further understand the functions of these receptors in the postnatal animal as it develops and grows (Metzger and Chambon 2001; Metzger et al. 2003).

Role of RARs in Regulating Cell Proliferation and Cancer

In addition to the effects that retinoic acids and the RARs have on developmental pathways, a large amount of evidence has emerged implicating RARs in the control of cell-cycle pathways and cellular proliferation. In normal cells, retinoic acids generally inhibit cell-cycle progression by instituting a block in the G1 phase of the cell cycle (Mongan and Gudas 2007). Of all the RAR isoforms, these effects are mostly mediated by RARβ2 following binding and activation by retinoic acids (Faria et al. 1999). Several studies have shown in a number of cell types that activation of RARβ2 leads to the transactivation of several genes involved in cell-cycle arrest such as p21CIP1 and p27KIP1 (Li et al. 2004; Suzui et al. 2004). In addition to activating cell-cycle arrest proteins, RARs also mediate both the downregulation of mRNA expression as well as protein ubiquitination and degradation for both the Cyclin D and E families which prevents progression of the cell cycle from the G1 to S phase (Tang and Gudas 2011). Moreover, RARs induce apoptosis following binding of retinoids in a number of cell types as a guard against tumor formation. Retinoic acid binds to RARα and induces apoptosis in both acute lymphoblastic leukemia cells as well as myeloid leukemia cell lines (Chikamori et al. 2006; Luo et al. 2009). In addition, it has been reported that RARγ induces apoptosis upon binding to retinoids in both skin keratinocytes as well as pancreatic adenocarcinoma cells (Hatoum et al. 2001; Pettersson et al. 2002). Finally, RARβ2 has been implicated in the induction of apoptosis in breast cells. Taken together, these observations have provided clear evidence that RARs play key roles in the regulation of cell-cycle progression and cell growth as well as apoptosis and therefore when RAR-mediated signaling pathways are disturbed, there can be major implications for the development and progression of cancer.

One of the most well-studied examples of aberrant RAR signaling leading to cancer is acute promyelocytic leukemia (APL). It has been shown in a number of reports that APL is the result of a genetic rearrangement of the RARα gene that fuses it to the promyelocytic leukemia gene (PML) or other PML-related genes (Tang and Gudas 2011). These PML/RARα fusion proteins lead to dramatic increases in expression of both HDACs and DNA methyltransferases that cause reductions in gene expression for retinoid-regulated genes such as those involved in differentiation (Fig. 3). As previously stated, under normal conditions retinoid-regulated genes regulate cellular proliferation mainly through inhibiting cell-cycle progression and promoting cellular differentiation. When the expression of these genes is reduced, the regulatory controls on these functions are lost resulting in uncontrolled proliferation. In addition to inducing epigenetic silencing, the PML/RARα fusion protein can also repress important RAR target genes such as RARβ2 itself by binding the RAR response element (RARE) on the RARβ2 promoter and recruiting corepressors such as NuRD that keep RARβ2 from being expressed.
Retinoic Acid Receptors (RARA, RARB, and RARC), Fig. 3

Model of aberrant repression of RAR gene activation by the PML/RARα fusion protein. Schematic comparing the ligand-dependent activation of RAR response genes in normal tissues expressing the wild-type RARα/RXRα heterodimer compared to the APL model where that is replaced by the PML/RARα fusion protein. The PML/RARα fusion protein forms a dominant negative repressive complex onto the RAREs due to the lack of RXRα and the enhanced recruitment of HDACs to the PML portion of the fusion protein. Reversing this phenomonon requires HDAC inhibitors or excess amounts of retinoic acids

The loss of retinoic acid signaling is not restricted to APL and in fact reduction in expression of both RARα and RARβ2 has been shown to occur in several types of cancer such as embryonal carcinomas, acute myeloid leukemia, and breast cancer (Mongan and Gudas 2007; Altucci et al. 2007). In contrast to APL where RARα is mutated into the PML/RARα fusion protein, most cancer cell types do not contain mutated RARs but rather have dramatically downregulated expression of these receptors. Understanding the underlying mechanisms behind the silencing of RARs in cancer cells has been a major focus of recent research and several mechanisms have been uncovered. For example, it was reported that the RARβ2 promoter in many cancer cell types is silenced by hypermethylation at CpG regions of its promoter. In addition, corepressors such as PRAEME, meningioma 1, acinus-S′, HACE1, and SMRT have all been shown to inhibit expression of either RARβ2 or RAR response genes either due to overexpression of these corepressors or a greater affinity for the RARβ2 response elements concomitant to changes caused by aberrant AKT signaling in a variety of cancer cell types (Tang and Gudas 2011). In contrast, repression of RAR coactivator expression has also been observed to downregulate expression of RARs in neuroblastoma cells. Taken together, these reports indicate that a multitude of mechanisms are at work in various cancer cell types that all result in the repression of RARs and their downstream target genes. Regardless of mechanism, the end result is an absence of RAR-mediated balances between differentiation and proliferation and demonstrates the vital roles RARs play in the progression of cancer.

Development of RAR Ligands for Use as Therapeutics

Given the correlation between the reduction in RAR-mediated retinoic acid signaling and the progression of a number of cancers, therapies have been developed to treat cancer patients with natural retinoids such as all-trans retinoic acid (ATRA) to induce differentiation and cell growth arrest. This strategy has been extremely successful in the treatment of APL as pharmacological doses of retinoic acid stimulate irreversible differentiation of leukemic cells into granulocytes. Moreover, it has been reported that pharmacological doses of retinoic acid also trigger growth arrest and differentiation of leukemia stem cells known as leukemia-inducing cells (LICs) (Tang and Gudas 2011). When combined with other apoptosis-inducing chemotherapeutic drugs such as anthracyclins, retinoic acid treatment reverses gene silencing and leads to induced cell death of the cancer cells curing 70–80% of APL patients (Altucci et al. 2007). This treatment is successful since the expression of the PML/RXRα fusion protein is high and the RARα portion of this protein contains a functioning ligand-binding domain and coactivator recruitment site that allows for the retinoic acid-mediated activation of a number of RARα genes that stimulate differentiation.

Differentiation therapy involving treatment with natural retinoids has been developed for many cancers such as breast, ovarian, renal, head and neck, melanoma, and prostate. However, the success of this approach has been much less successful in these other types of cancers where the expression of RAR genes themselves are downregulated by events such as DNA methylation of their promoters as previously discussed. Combination therapies have been adopted with some success to overcome these limitations with the coadministration of HDAC inhibitors and DNMTase inhibitors in addition to retinoic acid (Tang et al. 2009). This strategy first reverses the repressive effects of protein acetylation and DNA methylation on RAR gene expression and then once expressed, provides the natural agonist to activate RAR target genes to induce growth arrest and apoptosis (Fig. 4).
Retinoic Acid Receptors (RARA, RARB, and RARC), Fig. 4

Schematic model of the benefit of combination therapy to reverse the repression of RARβ2 expression in various cancers. (a) Normal tissue where RAR/RXRs regulate expression of RARβ2 which is vital for the balance between cell proliferation and differentiation and inducing apoptosis when necessary (b) Tumor tissue where hypermethylation of CpG islands on the promoter of RARβ2 prevent its expression and in turn repress genes involved in regulation of cellular proliferation and apoptosis leading to the cancer phenotype. (c) Treatment of tumors with combinations of retinoic acid and other inhibitors such as DNA methyltransferase inhibitors (DNMTase inhibitor) that first remove the detrimental hypermethylation and then restore normal expression of RARβ2 which is activated by retinoic acids

While cell differentiation therapies using high levels of natural retinoids such as ATRA have proven to be very successful in the treatments of some cancers, there are significant drawbacks to their therapeutic use. Retinoids are powerful teratogens that at pharmacological concentrations can induce congenital defects and toxicity in all vertebrate species. In addition, there are some cancers such as prostate cancer where ATRA and other synthetic retinoid agonists are not effective in inducing growth arrest and/or apoptosis. Moreover, a common feature of many cancers is the development of resistance to the growth inhibitory effects of retinoids limiting the utility of these therapies. Even APL, which responds well to differentiation therapy, has several variants that display retinoid resistance and does not respond to this therapy. For these reasons, efforts have been underway to develop new types of synthetic ligands for RAR that can promote the positive effects of retinoids without the detrimental side effects.

A number of synthetic retinoids have been developed as potential therapeutics for a variety of cancers. These are often referred to as atypical retinoids or retinoid-related molecules because they are based on the retinoic acid structure and have been shown to bind and transactivate RARs. Many of these compounds have been approved for the treatment of a number of diseases such as cancer, acne, and psoriasis (Altucci et al. 2007). The majority of these atypical retinoids are RAR agonists; however, there have been some RAR antagonists that have also been synthesized. In some cancers such as prostate cancer, pan-specific antagonists of RAR such as AGN194310 demonstrated much more significant anti-proliferative and pro-apoptotic effects than any RAR natural or synthetic agonist. In fact a number of synthetic molecules known as the retinoid-related molecules such as MX781, AGN 194310, and ST1926 have demonstrated potent anti-proliferative activities against large panels of human tumor cells (de Lera et al. 2007). Until recently, all of the synthetic retinoid-related molecules reported that directly bind and modulate RAR activity share structural similarities to the natural agonist retinoic acid. This means that while some have proven efficacious in the treatments of a number of important cancers, they could still be susceptible to the same limitations regarding retinoid resistance as the natural retinoids. Interestingly, a recent report has identified the first synthetic non-retinoid, non-acid RAR modulator that binds and activates all three isoforms of RAR (Busby et al. 2011). Synthetic structures such as these may provide the basis for novel chemical scaffolds of non-retinoid, non-acid RAR modulators that may be developed that are potent and efficacious toward restoring RAR signaling while at the same time overcome the challenges of toxicity and resistance seen with use of natural retinoids such as ATRA.

Summary

The retinoic acid receptors (RARs) are ligand-dependent transcription factors that belong to the NR1B subtype of the nuclear receptor (NR) superfamily. RARs are ligand-dependent transcription factors that bind to retinoids, the most potent biologically active forms of vitamin A, and heterodimerize with the rexinoid X receptor (RXR) to regulate many genes involved in the regulation of cellular growth and differentiation. RARs play significant roles in a number of developmental cascades from formation of limbs and organs to the central nervous system. In addition, all three RAR isoforms are instrumental in the control of a cellular growth through the inhibition of the cell cycle. That combined with the activation of genes involved in differentiation provides multiple pathways that RARs regulate cellular growth. Given these critical roles in cellular growth, it is not surprising that a great deal of evidence has emerged that either mutations or reductions in RAR expression are correlated with a number of cancers. This has led to the development of differentiation therapies alone or in combination with other types of drugs to restore RAR-mediated retinoic acid signaling in a number of cancers. Due to the potential toxicity and emergence of retinoid resistance in some cancers, synthetic retinoid-related molecules have been developed including one novel non-acid non-retinoid chemical scaffold that may provide safer, more efficacious ways to treat cancer by restoring normal RAR-mediated RA signaling. Further understandings of the roles of the various RAR isoforms in the progression of cancer and how to modulate the activities of RARs may provide important clues to develop novel therapies to treat cancer.

References

  1. Altucci L, Leibowitz MD, Ogilvie KM, de Lerea AR, Gronemeyer H. RAR and RXR modulation in cancer and metabolic disease. Nat Rev Drug Discov. 2007;6(10):793–810.PubMedCrossRefGoogle Scholar
  2. Busby SA, Kumar N, Kuruvilla DS, Istrate MA, Conkright JJ, Wang Y, Kamenecka TM, Cameron MD, Roush WR, Burris TP, Griffin PR. Identification of a novel non-retinoid pan inverse agonist of the retinoic acid receptors. ACS Chem Biol. 2011;6(6):618–27.PubMedPubMedCentralCrossRefGoogle Scholar
  3. Chikamori K, Hill JE, Grabowski DR, Zarkhin E, Grozav AG, et al. Downregulation of topoisomerase IIβ in myeloid leukemia cell lines leads to activation of apoptosis following all-trans retinoicacid–induced differentiation/growth arrest. Leukemia. 2006;20:1809–18.PubMedCrossRefGoogle Scholar
  4. de Lera AR, Bourguet W, Altucci L, Gronemeyer H. Design of selective nuclear receptor modulators: RAR and RXR as a case study. Nat Rev Drug Discov. 2007;6:811–20.PubMedCrossRefGoogle Scholar
  5. Dupe V, Ghyselinck NB, Thomazy V, Nagy L, Davies PJ, Chambon P, Mark M. Essential roles of retinoic acid signaling in interdigital apoptosis and control of BMP-7 expression in mouse autopods. Dev Biol. 1999;208:30–43.PubMedCrossRefGoogle Scholar
  6. Dupe V, Matt N, Garnier JM, Chambon P, Mark M, Ghyselinck NB. A newborn lethal defect due to inactivation of retinaldehyde dehydrogenase type 3 is prevented by maternal retinoic acid treatment. Proc Natl Acad Sci U S A. 2003;100:14036–41.PubMedPubMedCentralCrossRefGoogle Scholar
  7. Faria TN, Mendelsohn C, Chambon P, Gudas LJ. The targeted disruption of both alleles of RARbeta(2) in F9 cells results in the loss of retinoic acid-associated growth arrest. J Biol Chem. 1999;274(38):26783–8.PubMedCrossRefGoogle Scholar
  8. Halilagic A, Ribes V, Ghyselinck NB, Zile MH, Dolle P, Studer M. Retinoids control anterior and dorsal properties in the developing forebrain. Dev Biol. 2007;303:362–75.PubMedCrossRefGoogle Scholar
  9. Hatoum A, El-Sabban ME, Khoury J, Yuspa SH, Darwiche N. Overexpression of retinoic acid receptors α and γ into neoplastic epidermal cells causes retinoic acid–induced growth arrest and apoptosis. Carcinogenesis. 2001;22:1955–63.PubMedCrossRefGoogle Scholar
  10. Kochhar DM, Jiang H, Penner JD, Johnson AT, Chandraratna RA. The use of a retinoid receptor antagonist in a new model to study vitamin A-dependent developmental events. Int J Dev Biol. 1998;42:601–8.PubMedGoogle Scholar
  11. Li R, Faria TN, Boehm M, Nabel EG, Gudas LJ. Retinoic acid causes cell growth arrest and an increase in p27 in F9 wild type but not in F9 retinoic acid receptor β2 knockout cells. Exp Cell Res. 2004;294:290–300.PubMedCrossRefGoogle Scholar
  12. Luo P, Lin M, Lin M, Chen Y, Yang B, He Q. Function of retinoid acid receptor α and p21 in all-trans-retinoic acid–induced acute T-lymphoblastic leukemia apoptosis. Leuk Lymphoma. 2009;50:1183–9.PubMedCrossRefGoogle Scholar
  13. Maden M. Retinoid signaling in the development of the central nervous system. Nat Rev Neurosci. 2002;3:843–53.PubMedCrossRefGoogle Scholar
  14. Mark M, Ghyselinck NB, Chambon P. Function of retinoic acid receptors during embryonic development. Nucl Recept Signal. 2009;7:1–15. doi: 10.1621/nrs.07002.Google Scholar
  15. Metzger D, Chambon P. Site- and time-specific gene targeting in the mouse. Methods. 2001;24:71–80.PubMedCrossRefGoogle Scholar
  16. Metzger D, Indra AK, Li M, Chapellier B, Calleja C, Ghyselinck NB, Chambon P. Targeted conditional somatic mutagenesis in the mouse: temporally-controlled knock out of retinoid receptors in epidermal keratinocytes. Methods Enzymol. 2003;364:379–408.PubMedGoogle Scholar
  17. Mongan NP, Gudas LJ. Diverse actions of retinoid receptors in cancer prevention and treatment. Differentiation. 2007;75(9):853–70.PubMedCrossRefGoogle Scholar
  18. Pettersson F, Dalglesh AG, Bissonnette RP, Colston KW. Retinoids cause apoptosis in pancreatic cancer cells via activation of RARγ and altered expression of Bcl-2/Bax. Br J Cancer. 2002;87:555–61.PubMedPubMedCentralCrossRefGoogle Scholar
  19. Sandell LL, Sanderson BW, Moiseyev G, Johnson T, Mushegian A, Young K, Rey JP, Ma JX, Staehling-Hampton K, Trainor PA. RDH10 is essential for synthesis of embryonic retinoic acid and is required for limb, craniofacial, and organ development. Genes Dev. 2007;21:1113–24.PubMedPubMedCentralCrossRefGoogle Scholar
  20. Suzui M, Shimizu M, Masuda M, Lim JT, Yoshimi N, Weinstein IB. Acyclic retinoid activates retinoic acid receptor βand induces transcriptional activation of p21CIP1 in HepG2 human hepatoma cells. Mol Cancer Ther. 2004;3:309–16.PubMedGoogle Scholar
  21. Tang XH, Gudas LJ. Retinoids, retinoic acid receptors and cancer. Annu Rev Pathol Mech Dis. 2011;6:345–64.CrossRefGoogle Scholar
  22. Tang XH, Albert M, Scognamiglio T, Gudas LJ. A DNA methyltransferase inhibitor and alltransretinoic acid reduce oral cavity carcinogenesis induced by the carcinogen 4-nitroquinoline 1-oxide. Cancer Prev Res. 2009;2:1100–10.CrossRefGoogle Scholar
  23. Wendling O, Ghyselinck NB, Chambon P, Mark M. Roles of retinoic acid receptors in early embryonic morphogenesis and hindbrain patterning. Development. 2001;128:2031–8.PubMedGoogle Scholar

Copyright information

© Springer International Publishing AG 2018

Authors and Affiliations

  1. 1.Department of Molecular TherapeuticsThe Scripps Research InstituteJupiterUSA