RB1, development, and cancer

Abstract

The RB1 gene is the first tumor suppressor gene identified whose mutational inactivation is the cause of a human cancer, the pediatric cancer retinoblastoma. The 25 years of research since its discovery has not only illuminated a general role for RB1 in human cancer, but also its critical importance in normal development. Understanding the molecular function of the RB1 encoded protein, pRb, is a long-standing goal that promises to inform our understanding of cancer, its relationship to normal development, and possible therapeutic strategies to combat this disease. Achieving this goal has been difficult, complicated by the complexity of pRb and related proteins. The goal of this review is to explore the hypothesis that, at its core, the molecular function of pRb is to dynamically regulate the location-specific assembly or disassembly of protein complexes on the DNA in response to the output of various signaling pathways. These protein complexes participate in a variety of molecular processes relevant to DNA including gene transcription, DNA replication, DNA repair, and mitosis. Through regulation of these processes, RB1 plays a uniquely prominent role in normal development and cancer.

Figure 1. Structural organization of the Rb1 encoded protein

The schematic identifies major structural and functional elements including the “pocket,” the dual tandem cyclin folds (A1+B1, A2+B2), the unstructured tail (C), and threonine (T) and serine (S) residues whose phosphorylation regulates intermolecular and intramolecular protein interactions. The colored regions mark the positions of the five alpha helices that are characteristic of cyclin folds. Not shown are additional structural elements (i.e. alpha helices) that contribute to overall pRb structure. The schematic is approximately to scale.

Figure 2. The pRb/E2F cell proliferation switch

The pRb/E2F switch is bistable, existing primarily on one of two states, either permissive or non-permissive for cell proliferation. Bistability is established by both positive and negative feedback loops (see text). In its unphosphorylated state, pRb complexes with E2F transcription factors at gene promoters, blocks the ability of E2F to recruit transcriptional coactivators like histone acetylases (HAT), and recruits corepressors like histone deacetylases (HDAC). These coactivators and corepressors alter chromatin to a more open or condensed states, respectively. Mitogenic kinases phosphorylate pRb, altering its ability to interact with E2F and transcriptional corepressors. The resulting transcriptional derepression of genes like cyclins increases cyclin dependent kinase activity (CDK). Increased CDK activity, in turn, augments pRb phosphorylation thus creating a positive feedback loop that amplifies the mitogenic signal and triggers commitment to a round of cell division. Negative feedback is provided by the fact that the Rb1 promoter is itself regulated by E2F. As cells transit mitosis, pRb is dephosphorylated, resetting the switch to a non-proliferative state.

Figure 3. Stem/progenitor cells and sensitivity to pRb loss

The figure summarizes the hypothesis that there exists an inverse correlation between differentiation and sensitivity to pRb loss. Stem/progenitor cells are relatively undifferentiated and have a neutral chromatin state at key lineage specifying regulatory genes. Chromatin at such genes is characterized by the simultaneous presence of protein complexes and chromatin marks characteristic of both open (shapes marked +) and closed chromatin (shapes marked −). pRb is typically associated with factors that create a closed chromatin state repressive for transcription (although there are notable exceptions). Loss of pRb in such cells is expected to have a large effect on gene expression as it will tip the balance in favor gene expression. In more differentiated cells, chromatin is more rigidly committed to an open or closed state by multiple, redundant mechanisms. Loss of pRb in such cells is expected to have less effect on the expression of stably repressed genes as there are redundant mechanisms in place to maintain the closed chromatin state.