A truncated and catalytically inactive isoform of KDM5B histone demethylase accumulates in breast cancer cells and regulates H3K4 tri-methylation and gene expression

KDM5B histone demethylase is overexpressed in many cancers and plays an ambivalent role in oncogenesis, depending on the specific context. This ambivalence could be explained by the expression of KDM5B protein isoforms with diverse functional roles, which could be present at different levels in various cancer cell lines. We show here that one of these isoforms, namely KDM5B-NTT, accumulates in breast cancer cell lines due to remarkable protein stability relative to the canonical PLU-1 isoform, which shows a much faster turnover. This isoform is the truncated and catalytically inactive product of an mRNA with a transcription start site downstream of the PLU-1 isoform, and the consequent usage of an alternative ATG for translation initiation. It also differs from the PLU-1 transcript in the inclusion of an additional exon (exon-6), previously attributed to other putative isoforms. Overexpression of this isoform in MCF7 cells leads to an increase in bulk H3K4 methylation and induces derepression of a gene cluster, including the tumor suppressor Cav1 and several genes involved in the interferon-alpha and -gamma response. We discuss the relevance of this finding considering the hypothesis that KDM5B may possess regulatory roles independent of its catalytic activity. Overall design: Comparative gene expression profiling analysis of RNA-seq data for MCF7 and MDA-MB-231 cells over-expressing the KDM5B-NTT vs control samples.

KDM5B组蛋白去甲基化酶（KDM5B histone demethylase）在多种癌症中呈过表达状态，且在肿瘤发生过程中发挥着依赖特定情境的双向调控作用。这种功能两面性可通过表达具有不同功能的KDM5B蛋白同工型（isoform）得以解释，不同癌细胞系中此类同工型的表达水平存在差异。本研究证实，其中一类同工型KDM5B-NTT相较于经典PLU-1同工型（canonical PLU-1 isoform）具有显著更高的蛋白质稳定性，因此在乳腺癌细胞系中发生积累，而经典PLU-1同工型的蛋白周转速度更快。该同工型是由一段转录起始位点位于PLU-1同工型下游的信使RNA（mRNA）翻译而来的截短且催化活性失活的产物，其翻译起始使用了可变ATG起始密码子。此外，相较于PLU-1转录本，该同工型的mRNA还包含一段额外的外显子6（exon-6），该外显子此前曾被归因于其他推定同工型。在MCF7细胞中过表达该同工型，会导致整体H3K4甲基化水平升高，并诱导包括肿瘤抑制基因Cav1以及多个参与干扰素-α和干扰素-γ应答的基因在内的基因簇发生去抑制。本研究结合“KDM5B可能具有不依赖其催化活性的调控功能”这一假说，讨论了上述发现的研究意义。整体实验设计：对过表达KDM5B-NTT的MCF7与MDA-MB-231细胞的RNA测序（RNA-seq）数据开展比较基因表达谱分析，并以相应对照样本作为参照。