CENP-A overexpression promotes aneuploidy and karyotypic heterogeneity

Centromeric localization of the evolutionarily conserved histone H3 variant, CENP-A. is essential for chromosomal stability. CENP-A overexpression (OE) causesd its mislocalization to non-centromeric regions resulting in chromosomal instability (CIN) in yeast, flies and human cells. CENP-A OE and mislocalization have been observed in cancers and correlates with poor prognosis. However, the molecular consequences of CENP-A OE on CIN and aneuploidy have not been defined. Here, we overexpressed YFP-CENP-A in a pseudodiploid DLD1 cell line and showed that CENP-A OE leads to its mislocalization and CIN due to defects in kinetochore integrity and kinetochore-microtubule attachments. CENP-A OE also leads to its mislocalization and CIN in a xenograft mouse model. Under these conditions, it contributes to aneuploidy with karyotypic heterogeneity in human cells and xenograft mouse model. In summary, our studies provide a molecular link between CENP-A OE and aneuploidy, and suggest that karyotypic heterogeneity may contribute to the aggressive phenotype of CENP-A overexpressing cancers. Overall design: Three biological replicates were analyzed for each of two conditions

进化保守的组蛋白H3变体着丝粒蛋白A（CENP-A）的着丝粒定位对于维持染色体稳定性至关重要。着丝粒蛋白A过表达（overexpression，OE）会导致其错误靶向至非着丝粒区域，进而在酵母、果蝇及人类细胞中引发染色体不稳定性（chromosomal instability，CIN）。现有研究已在多种癌症中检测到CENP-A过表达与错误定位现象，且该表型与患者不良预后显著相关。然而，CENP-A过表达引发染色体不稳定性与非整倍体的具体分子机制仍未阐明。本研究在假二倍体DLD1细胞系中过表达黄色荧光蛋白（Yellow Fluorescent Protein，YFP）标记的CENP-A（YFP-CENP-A），发现CENP-A过表达会因动粒（kinetochore）完整性缺陷及动粒-微管连接异常，引发其错误定位与染色体不稳定性。在异种移植小鼠模型中，CENP-A过表达同样会诱导其错误定位与染色体不稳定性。在此条件下，CENP-A过表达会促使人类细胞与异种移植小鼠模型出现核型异质性相关的非整倍体。综上，本研究揭示了CENP-A过表达与非整倍体之间的分子关联，并提示核型异质性可能是CENP-A过表达癌症侵袭性表型的潜在诱因。
实验整体设计：两种实验条件各设置3次生物学重复并开展分析。