Control of chromosome stability by the \u3b2-TrCP\u2013REST\u2013Mad2 axis

Abstract

REST/NRSF (repressor-element-1-silencing transcription factor/ neuron-restrictive silencing factor) negatively regulates the tran- scription of genes containing RE1 sites1,2. REST is expressed in non-neuronal cells and stem/progenitor neuronal cells, in which it inhibits the expression of neuron-specific genes. Overexpression of REST is frequently found in human medulloblastomas and neuroblastomas3\u20137, in which it is thought to maintain the stem character of tumour cells. Neural stem cells forced to express REST and c-Myc fail to differentiate and give rise to tumours in the mouse cerebellum3. Expression of a splice variant of REST that lacks the carboxy terminus has been associated with neuronal tumours and small-cell lung carcinomas8\u201310, and a frameshift mutant (REST-FS), which is also truncated at the C terminus, has oncogenic properties11. Here we show, by using an unbiased screen, that REST is an interactor of the F-box protein b-TrCP. REST is degraded by means of the ubiquitin ligase SCFb-TrCP dur- ing the G2 phase of the cell cycle to allow transcriptional derepres- sion of Mad2, an essential component of the spindle assembly checkpoint. The expression in cultured cells of a stable REST mutant, which is unable to bind b-TrCP, inhibited Mad2 expres- sion and resulted in a phenotype analogous to that observed in Mad21/2 cells. In particular, we observed defects that were con- sistent with faulty activation of the spindle checkpoint, such as shortened mitosis, premature sister-chromatid separation, chro- mosome bridges and mis-segregation in anaphase, tetraploidy, and faster mitotic slippage in the presence of a spindle inhibitor. An indistinguishable phenotype was observed by expressing the oncogenic REST-FS mutant11, which does not bind b-TrCP. Thus, SCFb-TrCP-dependent degradation of REST during G2 permits the optimal activation of the spindle checkpoint, and consequently it is required for the fidelity of mitosis. The high levels of REST or its truncated variants found in certain human tumours may contri- bute to cellular transformation by promoting genomic instability