Genomic Instability in Gadd45a^-/- Cells is Coupled with S-Phase Checkpoint Defects

Abstract

Gadd45a is a p53-regulated gene whose protein product, like p53, is involved in maintenanceof genome stability. Specifically, deletion of Gadd45a leads to extensive aneuploidy as aconsequence of centrosome amplification and subsequent abnormal segregation of chromosomesduring mitosis. S-phase checkpoints were investigated in Gadd45a-/- cells to determine possibledefects contributing to the uncoupling of centrosome duplication and DNA replication. In thepresence of hydroxyurea, Gadd45a-/- mouse embryo fibroblasts show increased centrosomeamplification coupled with loss of a sustained S-phase checkpoint. Gadd45a deletion allowsanother form of genomic instability, gene amplification, when p21 (Cdkn1a gene product) isdeleted also. Gene amplification in Gadd45a-/-p21-/- cells correlated with loss of both G1 andS-phase checkpoints. Multiple conditions of nutrient deprivation failed to prevent DNAsynthesis in Gadd45a-/- cells. Gadd45a is therefore required for proper S-phase control andcheckpoints under multiple conditions of nutrient deprivation. It is proposed that loss of S-phasecontrol may account for both the uncoupling of DNA replication and centrosome duplication,and conferring gene amplification proficiency in cells lacking Gadd45a-/-. This is of particularimportance for solid tumors, which may lack sufficient nutrients yet are unable to elicitcheckpoints preventing genomic instability under these conditions.