Inactivation of NIPA Results in Premature Mitotic Entry and Subsequent Mitotic Catastrophe.

Ubiquitin-mediated destruction of regulatory proteins marks a vital means of controlling cell cycle progresssion. The E3 ubiquitin-ligases are prominent in this process as they determine specificity of the ubiquitination process and thus regulate proteasomal degradation of target proteins. Recently, a class of E3 ligases referred to as SCF complexes has been identified. The substrate binding specifity within this class of E3 ligases is mediated by a class of molecules termed F-box proteins. We previously reported the cloning of NIPA (nuclear interaction partner of ALK) in complex with constitutively-active oncogenic fusions of ALK, which contribute to the development of certain lymphomas and sarcomas. Subsequently we characterized NIPA as a human F-box protein that defines an oscillating ubiquitin ligase (SCF-NIPA) which targets nuclear cyclin B1 in interphase. We have now determined the consequence of inactivating NIPA with regard to cell cycle regulation using an RNAi approach. Kinetic analysis of cell cycle phase transition times revealed a premature onset of Cdk1/cyclin B1 kinase activity and early mitotic entry in cells treated with NIPA siRNA. Cyclin B1 was shown to accumulate within the nucleus in these cells correlative to a reduced ubiquitination activity of the SCF-NIPA complex. Subsequent to premature mitotic entry, NIPA inactivated cells arrested in prometaphase and mitotic catastrophe was observed thereafter. We searched for relevant proteins involved in this process and found Survivin, a member of the inhibitor of apoptosis proteins (IAP) to be strongly downregulated in NIPA siRNA treated cells. Survivin has been shown to function as a mitotic checkpoint molecule that induces mitotic catastrophe subsequent to aberant mitosis when inactivated or downregulated. While the precise functional relationship between Survivin and NIPA is still under investigation, the current data distinguishes NIPA as a central molecule in timing mitotic entry. Given this function, NIPA directly influences the fidelity of DNA replication and segregation. Interference with NIPA function may therefore be an oncogenic principle that favours genomic instability in tumor cells.