Loss of CHFR in Human Mammary Epithelial Cells Causes Genomic Instability by Disrupting the Mitotic Spindle Assembly Checkpoint

• Published in: Neoplasia (New York, N.Y.) Vol. 10; no. 7; pp. 643 - 652
• Main Authors: Privette, Lisa M., Weier, Jingly Fung, Nguyen, Ha Nam, Yu, Xiaochun, Petty, Elizabeth M.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01-07-2008; Elsevier
• Subjects: Aneuploidy; Aurora Kinases; Calcium-Binding Proteins - metabolism; Cell Cycle Proteins - genetics; Cell Cycle Proteins - metabolism; Cells, Cultured; Chromosome Segregation - genetics; Epithelial Cells - metabolism; Gene Deletion; Gene Expression Regulation; Genes, cdc; Genomic Instability - genetics; Genomic Instability - physiology; Humans; Mad2 Proteins; Mammary Glands, Human - metabolism; Models, Biological; Neoplasm Proteins - genetics; Neoplasm Proteins - metabolism; Poly-ADP-Ribose Binding Proteins; Protein Binding; Protein-Serine-Threonine Kinases - genetics; Protein-Serine-Threonine Kinases - metabolism; Repressor Proteins - metabolism; Spindle Apparatus - genetics; Tubulin - genetics; Tubulin - metabolism; Ubiquitin-Protein Ligases; SKY; inhibitor; HDI; CHFR
• ISSN: 1476-5586; 1476-5586; 1522-8002
• DOI: 10.1593/neo.08176

CHFR is an E3 ubiquitin ligase and an early mitotic checkpoint protein implicated in many cancers and in the maintenance of genomic stability. To analyze the role of CHFR in genomic stability, by siRNA, we decreased its expression in genomically stable MCF10A cells. Lowered CHFR expression quickly led to increased aneuploidy due to many mitotic defects. First, we confirmed that CHFR interacts with the mitotic kinase Aurora A to regulate its expression. Furthermore, we found that decreased CHFR led to disorganized multipolar mitotic spindles. This was supported by the finding that CHFR interacts with α-tubulin and can regulate its ubiquitination in response to nocodazole and the amount of acetylated α-tubulin, a component of the mitotic spindle. Finally, we found a novel CHFR interacting protein, the spindle checkpoint protein MAD2. Decreased CHFR expression resulted in the mislocalization of both MAD2 and BUBR1 during mitosis and impaired MAD2/CDC20 complex formation. Further evidence of a compromised spindle checkpoint was the presence of misaligned metaphase chromosomes, lagging anaphase chromosomes, and defective cytokinesis in CHFR knockdown cells. Importantly, our results suggest a novel role for CHFR regulating chromosome segregation where decreased expression, as seen in cancer cells, contributes to genomic instability by impairing the spindle assembly checkpoint.