Role of Fbxw5 in the induction of centrosome abnormalities and liver tumor formation

Centrosome duplication is tightly regulated process and should occur only once per cell division. Furthermore, genomic stability requires that centrosomes and chromosomes are segregated equally to the cell poles during mitosis. However in most types of tumors the centrosome duplication cycle is corrupted and it is unclear whether centrosome abnormalities are cause or consequence of the tumor formation process. Liver tumors belongs to the most severe cancer types, moreover frequently displays centrosome amplification [1]. Previous work in our group indicates a central role for the ubiquitin E3 ligase SCFFBXW5 in the initiation of the centrosome cycle by regulating the level of SASS6. RNAi mediated destabilization of Fbxw5 induces centrosome amplification and the formation of multinucleated cells [2]. Based on these findings, we decided to investigate the role of Fbxw5 in the induction of centrosome abnormalities in the liver cell cycle and in liver tumor formation. To this end, we take advantage of well-established shRNA technology as well as CRISPR/Cas9 to silence mouse Fbxw5, either via retroviral transfer in vitro or through a transposon based system in vivo using sleeping beauty transposase. We show that knockout of Fbxw5 leads to impaired proliferation and migration in vitro. Moreover after suppression of Fbxw5 cells display multipolar spindles accompanied by a delay in the transition from M to G1 phase. Although these multipolar spindles have their origin in amplified centrosomes, which may result in genomic instability, our in vivo studies reveal that the abnormality in centrosome number is not sufficient for tumor formation in wild type, p19-/- and p53-/- backgrounds. Thus, we plan to perform screening using Fbxw5 deficient cells and genome wide CRISPR/Cas9 library to identify further players in centrosome duplication.

References:

[1] Gönczy, P., Centrosomes and cancer: revisiting a long-standing relationship. Nature Reviews Cancer 15, 639 – 652 (2015)

[2] Puklowski, A., et al., The SCF-FBXW5 E3-ubiquitin ligase is regulated by PLK4 and targets HsSAS-6 to control centrosome duplication. Nat Cell Biol, 2011. 13(8): p. 1004 – 9.