RARARetinoic Acid Receptor Alpha

Taylor Beard

Big Picture• RARA encodes ligand activated transcription factor

RARα protein• RARα promotes transcription of genes involved in

differentiation and proliferation• Activated by Retinoic Acid (RA) ligand and co-

activators• Acts in a dominant negative manner in Acute

Promyelocytic Leukemia (APL) due to an oncogenic translocation mutation

• Translocation fusion protein PML-RARα inhibits differentiation of promyelocytes

• ATRA (All-Trans Retinoic Acid) treatment restores differentiation

RARA Protein Structure • N-terminal: transcription modulation region

AF-1 & AF-2

• DNA-binding domain (DBD): Zinc fingers

• Hinge region: allows rotation• Ligand binding region (LBD): ligand binding,

dimerization, and transactivation

• C-terminal: function unknown

• Phosphorylation: Ser 369 residue of LBD

Vigue (2010) Atlas of Genetics and Cytogenetics in Oncology and Haematology

RARα Function• Ligand activated transcription factor– Ligand = Retinoic Acid (RA)

• RAR/RXR heterodimers bind to Retinoic Acid Response Elements (RAREs) in target genes– heterodimer formed via binding of DBD domains– Target genes: differentiation & proliferation

• Absence of RA:– Recruit corepressors NCoR & SMRT– Histone deacetylase activity: ↑ chromatin compaction – transcriptional repression

• Presence of RA:– ligand-induced conformational changes– Dissociate corepressors & recruit co-activators (SRC/p160)– Histone acetyl- & methylation : ↓ chromatin compaction – transcription promotion

RARα Mechanism of ActionCOREPRESSORS (NCoR, SMRT)

CO-ACTIVATORS (SRC/p160, p300/CBP, CARM-1)+ RA (ligand)

Niederreither & Dolle (2008) Nature 9:541–553

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Focosi (2001)

RARA Oncogenic Gene Transformation in APL

• t(15;17) PML-RARA fusion gene- Recruits tightly bound corepressors- Physiological doses of RA inadequate for

activation• Dominant-negative inhibitor of RA signaling

pathway

• Inhibits differentiation of promyelocytes

Melnick & Licht JD (1999) Blood. 93:3167–3215

Oncogenic Activity in

APL

Goal of APL

Treatment

title

(Promyelocyte) (Mature WBC)Pitha-Rowe et al. (2003) Leukemia 17:1723–1730

PML-RARα Inhibits Differentiation

Figure 1. Expression of PML-RARα inhibits normal expression of differentiation markers in myeloid precursor cell lines.

(Control) (PML-RARα)

Grignani et al. (1993) Cell 74:423–431

ATRA Restores Differentiation

Figure 2. (below) Northern blot. ATRA treatment increases mRNA expression of differentiation transcription factor PU.1

Figure 3. (above) Western blot. ATRA treatment results in loss of PML-RARA fusion protein expression and induction of PU.1 protein expression.

Mueller et al. (2006) Blood 107:3330–3338

Acute Promyelocytic Leukemia (APL)

• A type of acute myeloid leukemia - AML M3• Deficiency of mature WBCs and excess of promyelocytes• Incidence

– Rare: 5% of all AML cases– Higher in Italy and the Americas

• Risk factors– No known environmental risk factors– Any age (mean 40-50 yrs); equal in M/F

• Symptoms: the ‘classic triad’– Fatigue– Bruising and bleeding– Infections and weight loss

• Prognosis– 5 year survival rate: 58.5%

APL Treatment• Substitution Therapy– Stabilize bleeding tendency

• All-Trans-Retinoic Acid (ATRA)– Promotes differentiation via PML-RARα cleavage

and degradation• Chemotherapy– Eliminates existing tumor cells

• Arsenic Trioxide (ATO)– Degrades PML-RARα via a different pathway than

ATRA

References• Bastien J, Rochette-Egly C. 2004. Nuclear retinoid receptors and the transcription of retinoid-target genes

Gene. Gene 328:1–16.• Focosi D. 2001. Physiology of adult homo sapiens - blood (haematology : plasma, blood cells, and

coagulation) and lymph [Internet]. Available from: http://www.ufrgs.br/imunovet/molecular_immunology/blood.html

• Grignani Francesco, Ferucci PF, Testa U, Talamo G, Fagioli M, Alcalay M, Mencarelli A, Grignani Fausto, Peschle C, Nicholetti I, et al. 1993. The acute promyelocytic leukemia-specific PML-RARα fusion protein inhibits differentiation and promotes survival of myeloid precursor cells Cell. Cell 74:423–431.

• Lingfelder E, Niederwieser D, Platzbecker U, Schlenk RF, Wörmann J. 2012. Acute Promyelocytic Leukemia (APL) [Internet]. (onkopedia, editor.). Available from: https://www.onkopedia-guidelines.info/en/onkopedia/guidelines/acute-promyelocytic-leukemia-apl/@@view/html/index.html#ID0EG

• Melnick A, Licht JD. 1999. Deconstructing a Disease: RARα, Its Fusion Partners, and Their Roles in the Pathogenesis of Acute Promyelocytic Leukemia Blood. Blood 93:3167–3215.

• Mueller BU, Pabst T, Fos J, Fey MF, Asou N, Buergi U, Tenen DG. 2006. ATRA resolves the differentiation block in t(15;17) acute myeloid leukemia by restoring PU.1 expression Blood. Blood 107:3330–3338.

• Niederreither K, Dolle P. 2008. Retinoic acid in development: towards an integrated view Nature Reviews Genetics. Nature 9:541–553.

• Pitha-Rowe I, Petty WJ, Kitaweeran S, Dmitrovsky E. 2003. Retinoid target genes in acute promyelocytic leukemia Leukemia. Leukemia 17:1723–1730.

• Vigue F. 2010. RARA (Retinoic acid receptor, alpha) [Internet]. (Atlas of Genetics and Cytogenetics in Oncology and Haematology, editor.). Available from: http://atlasgeneticsoncology.org/Genes/RARAID46.html