More epigenetic hits than meets the eye: microRNAs and genes associated with the tumorigenesis of retinoblastoma

Abstract

Retinoblastoma (RB), a childhood neoplasia of the retinoblasts, can occur unilaterally or bilaterally, with one or multiple foci per eye. RB is associated with somatic loss of function of both alleles of the tumor suppressor gene RB1. Hereditary forms emerge due to germline loss of function mutations in RB1 alleles. RB has long been the prototypic "model" cancer ever since Knudson's "two-hit" hypothesis. However, a simple two-hit model for RB is challenged by an increasing number of studies documenting additional hits that contribute to RB development. Here we review the genetics and epigenetics of RB with a focus on the role of small non-coding RNAs (microRNAs) and on novel findings indicating the relevance of DNA methylation in the development and prognosis of this neoplasia. Studies point to an elaborated landscape of genetic and epigenetic complexity, in which a number of events and pahtways play crucial roles in the origin and prognosis of RB. These include roles for microRNAs, inprinted loci, and parent-of-origin contributions to RB1 regulation and RB progression. This complexity is also manifested in the structure of the RB1 locus itself: it includes numerous repetitive DNA segments and retrotransposon insertion elements, some of which are actively transcribed from the RB1 locus. Altogether, we conclude that RB1 loss of function represents the tip of an iceberg of events that determine RB development, progression, severity, and disease risk. Comprehensive assessment of personalized RB risk will require genetic and epigenetic evaluations beyond RB1 protein coding sequences.

Keywords: Rb1; childhood cancer; imprinting; methylation; retinoblastoma; risk assessment; tumor suppressor; two-hit hypothesis.

Figure 1

Biogenesis of microRNAs (miRNAs). miRNA genes are transcribed and the resulting primary transcripts (pri-miRNAs) are polyadenylated at the 3′ end and capped at the 5′ end. Pri-miRNA molecules are recognized by the Drosha-DGCR8 complex and trimmed in a precursor-miRNA (pre-miRNA) that is transported to the cytoplasm by Exportin-5. In the cytoplasm, Dicer processes the pre-miRNAs and one miRNA duplex is released from each pre-miRNA. The two strands of the duplex are separated from each other by the Dicer-TRBP complex and one of the strands (called mature miRNA) is incorporated into the RNA-induced silencing complex (RISC) that will target specific mRNAs in a sequence-dependent manner. The other strand, which is not incorporated into RISC, is called the miRNA strand and is degraded.

Figure 2

An overview to cell cycle control by microRNAs and some key proteins. S indicates the S-phase; M indicates the mitosis; G1 and G2 indicate transition phases of the cell cycle whereas G0 indicates quiescent cells.

Figure 3

Chromosome 13 and the RB1 gene (not drawn to scale). (A) Chromosome 13 and the q14.2 band highlighted in a red square. Inside q14.2 band are RB1 gene (highlighted in a green square) and flanking genes. (B) RB1 gene showing exons 1–27. Black boxes are untranslated regions (UTR) 5′ and 3′. Two promoter regions investigated in methylation studies are shown inside the 5′UTR. A large number of MSP studies investigated the essential promoter region, while the MLPA studies investigated part of these two regions. These regions concentrate a large number of CpG island, like CpG 106, CpG 42, and CpG 85. (C) A pair of horizontal lines showing the location of LINE1 repeats in RB1 gene oriented in the sense direction (top line) and antisense direction (bottom line). Each black bar represents one LINE1, orange bars represent two LINE1 repeats. The bar thickness is proportional to the size of the sequence. (D) A pair of horizontal lines showing the location of ALU repeats in RB1 gene oriented in the sense direction (top line) and antisense direction (bottom line). Each black bar represents one ALU repeat, orange bars represent two ALU repeats.