Center for Biomedical Engineering and Technology, University of Maryland School of Medicine, Baltimore, MD 21201, USA.
Department of Physiology, University of Maryland School of Medicine, Baltimore, MD 21201, USA.
Hum Mol Genet. 2022 Aug 17;31(15):2639-2654. doi: 10.1093/hmg/ddac010.
XBP1 variant 1 (Xv1) is the most abundant XBP1 variant and is highly enriched across cancer types but nearly none in normal tissues. Its expression is associated with poor patients' survival and is specifically required for survival of malignant cells, but the underlying mechanism is not known. Here we report that Xv1 upregulates the polyglutamylase tubulin tyrosine ligase-like 6 (TTLL6) and promotes mitosis of cancer cells. Like the canonical XBP1, Xv1 mRNA undergoes unconventional splicing by IRE1α under endoplasmic reticulum stress, but it is also constitutively spliced by IRE1β. The spliced Xv1 mRNA encodes the active form of Xv1 protein (Xv1s). RNA sequencing in HeLa cells revealed that Xv1s overexpression regulates expression of genes that are not involved in the canonical unfolded protein response, including TTLL6 as a highly upregulated gene. Gel shift assay and chromatin immunoprecipitation revealed that Xv1s bind to the TTLL6 promoter region. Knockdown of TTLL6 caused death of cancer cells but not benign and normal cells, similar to the effects of knocking down Xv1. Moreover, overexpression of TTLL6 partially rescued BT474 cells from apoptosis induced by either TTLL6 or Xv1 knockdown, supporting TTLL6 as an essential downstream effector of Xv1 in regulating cancer cell survival. TTLL6 is localized in the mitotic spindle of cancer cells. Xv1 or TTLL6 knockdown resulted in decreased spindle polyglutamylation and interpolar spindle, as well as congression failure, mitotic arrest and cell death. These findings suggest that Xv1 is essential for cancer cell mitosis, which is mediated, at least in part, by increasing TTLL6 expression.
XBP1 变体 1（Xv1）是最丰富的 XBP1 变体，在多种癌症类型中高度富集，但在正常组织中几乎不存在。它的表达与患者预后不良相关，并且是恶性细胞存活所必需的，但潜在的机制尚不清楚。在这里，我们报告 Xv1 上调多聚谷氨酰胺酶微管酪氨酸连接酶样 6（TTLL6）并促进癌细胞有丝分裂。与经典的 XBP1 一样，Xv1 mRNA 在内质网应激下通过 IRE1α 进行非常规剪接，但也通过 IRE1β 进行组成性剪接。剪接的 Xv1 mRNA 编码 Xv1 蛋白的活性形式（Xv1s）。在 HeLa 细胞中的 RNA 测序显示，Xv1s 的过表达调节不参与经典未折叠蛋白反应的基因的表达，包括 TTLL6 作为高度上调的基因。凝胶迁移分析和染色质免疫沉淀显示 Xv1s 结合到 TTLL6 启动子区域。TTLL6 的敲低导致癌细胞死亡，但良性和正常细胞不受影响，这与敲低 Xv1 的效果相似。此外，TTLL6 的过表达部分挽救了 BT474 细胞因 TTLL6 或 Xv1 敲低而诱导的细胞凋亡，支持 TTLL6 作为 Xv1 在调节癌细胞存活中必不可少的下游效应物。TTLL6 定位于癌细胞的有丝分裂纺锤体中。Xv1 或 TTLL6 的敲低导致纺锤体多聚谷氨酰胺化和极间纺锤体减少，以及着丝粒聚集失败、有丝分裂阻滞和细胞死亡。这些发现表明 Xv1 对于癌细胞有丝分裂是必需的，至少部分通过增加 TTLL6 的表达来介导。
