Regulation of FANCD2 monoubiquitination by a kinase-phosphatase cascade

Introduction

Chk1 kinase plays pivotal roles in maintenance of genomic stability through regulating DNA replication, DNA repair, DNA recombination and checkpoint pathway. We show here Chk1 activation is required not only for proper activation of FANCD2 monoubiquitination pathway but also for its maintenance under ongoing replication stress.

Materials/methods

FANCD2 monoubiquitination was probed by immunoblotting and immunofluorescence in U2OS cells under various conditions - the proteins involved in the putative FANCD2 maintenance pathway were depleted by si- and shRNA and when required, cells were treated by inhibitors that induce replication stress and inhibit selected players in the pathway.

Results and conclusions

Mechanistically, inhibition of Chk1 kinase activity leads to failure of AuroraA-PLK1 inactivation pathway P46 Posters Posters P47 upon replication stress, which is likely to result in impairment of FANCD2 monoubiquitination. Conversely, inhibition of AuroraA kinase activity can recover FANCD2 monoubiquitination in Chk1- deficient cells, suggesting that Chk1-dependent inactivation of the AuroraA-PLK1 pathway is essential for the FA pathway. We also found that protein phosphatase 1 and protein phosphatase 2A are required for inactivation of the AuroraA-PLK1 axis under replication stress. Since it is known that AuroraA kinase is overexpressed in some tumors, our findings may provide novel insights into mechanisms of genomic instability in a subset of cancer types.