Multiple molecular interactions redundantly contribute to RB-mediated cell cycle control

Daniel T Passos,Matthew J Cecchini,Michael J Thwaites,Srikanth Talluri,Jasmyne Carnevale,Frederick A Dick

doi:10.1186/s13008-017-0029-6

Daniel T Passos, Matthew J Cecchini + Show 4 more

Open Access

https://doi.org/10.1186/s13008-017-0029-6

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Abstract

BackgroundThe G1-S phase transition is critical to maintaining proliferative control and preventing carcinogenesis. The retinoblastoma tumor suppressor is a key regulator of this step in the cell cycle.ResultsHere we use a structure–function approach to evaluate the contributions of multiple protein interaction surfaces on pRB towards cell cycle regulation. SAOS2 cell cycle arrest assays showed that disruption of three separate binding surfaces were necessary to inhibit pRB-mediated cell cycle control. Surprisingly, mutation of some interaction surfaces had no effect on their own. Rather, they only contributed to cell cycle arrest in the absence of other pRB dependent arrest functions. Specifically, our data shows that pRB–E2F interactions are competitive with pRB–CDH1 interactions, implying that interchangeable growth arrest functions underlie pRB’s ability to block proliferation. Additionally, disruption of similar cell cycle control mechanisms in genetically modified mutant mice results in ectopic DNA synthesis in the liver.ConclusionsOur work demonstrates that pRB utilizes a network of mechanisms to prevent cell cycle entry. This has important implications for the use of new CDK4/6 inhibitors that aim to activate this proliferative control network.

Highlights

The G1-S phase transition is critical to maintaining proliferative control and preventing carcinogenesis
A cell culture assay demonstrates molecular redundancy of RB functions in proliferative control Tumor suppression by the retinoblastoma protein has typically been associated with its ability to block cell cycle progression and repress E2F transcription factors [4]
In an attempt to describe the molecular interactions necessary for retinoblastoma protein (pRB)-mediated cell cycle arrest we investigated forms of pRB that were individually mutated at each of three distinct binding surfaces in the large pocket; the general E2F binding site (RBG), the E2F1 specific site (RBS), and the LxCxE binding cleft

Summary

Introduction

The G1-S phase transition is critical to maintaining proliferative control and preventing carcinogenesis. The retinoblastoma tumor suppressor is a key regulator of this step in the cell cycle. Uninhibited cellular division is a feature of cancer cells. The cell division cycle is frequently controlled by decisions made in the G1 phase [2]. Once through this phase, the cell is committed to DNA replication and completion of cell division. The retinoblastoma gene product (pRB) has been shown to be a key regulator of the restriction point that is responsible for controlling S-phase entry [3]. RB performs this function by directly binding the transactivation domain of E2Fs, preventing the recruitment of

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