LINC-PINT 通过与 XRCC6 结合并影响其功能在鼻咽癌中发挥抑癌作用。

LINC-PINT plays an anti-tumor role in nasopharyngeal carcinoma by binding to XRCC6 and affecting its function.

机构信息

Hunan Provincial Key Laboratory of the Fundamental and Clinical Research on Functional Nucleic Acid, Changsha Medical University, Changsha 410219, China; The First Clinical College, Changsha Medical University, Changsha 410219, China; Hunan Provincial Key Laboratory of the Research and Development of Novel Pharmaceutical Preparations, Changsha Medical University, Changsha 410219, China.

Hunan Provincial Key Laboratory of the Fundamental and Clinical Research on Functional Nucleic Acid, Changsha Medical University, Changsha 410219, China.

出版信息

Pathol Res Pract. 2024 Aug;260:155460. doi: 10.1016/j.prp.2024.155460. Epub 2024 Jul 18.

DOI:10.1016/j.prp.2024.155460

PMID:39032384

Abstract

BACKGROUND

LINC-PINT was downregulated in nasopharyngeal carcinoma (NPC) and correlated with treatment efficiency of NPC. However, the underlying mechanism of LINC-PINT in NPC has not yet been fully explored.

METHOD

We used CellTiter luminescent assay, clone formation assay, Hoechst staining, and SYTO-9/PI staining to examine cell viability and cell apoptosis regulated by LINC-PINT in NPC cells. Xenograft tumor model, HE staining, Ki67 staining, and TUNEL assay were conducted to assess the role of LINC-PINT in vivo. Bioinformatics and RNA immunoprecipitation assay was performed to identify the binding protein of LINC-PINT. Fluorescence in situ hybridization and immunofluorescence were utilized to measure the colocalization of XRCC6 with LINC-PINT and DNA-PKcs. Mito-Tracker red CMXRos staining was used to label mitochondria in cells specifically.

RESULT

We found LINC-PINT was downregulated in many tumors (including NPC) and associated with poor prognosis. The cell viability was significantly inhibited and cell apoptosis was remarkably promoted in LINC-PINT overexpressed cells in contrast to control cells. The growth of tumor xenografts was significantly suppressed and the tumor weight was significantly decreased in LINC-PINT overexpression group compared to the control group. Correspondingly, the positive Ki67 foci was decreased while TUNEL foci was increased in LINC-PINT overexpression group. Mechanically, we verified XRCC6 as a new binding protein of LINC-PINT through RNA binding domains prediction, RIP and colocalization of LINC-PINT and XRCC6. By binding to XRCC6, LINC-PINT interfered the formation of DNA-PK complex, regulated mitochondria accumulation status and affected the modification of apoptosis proteins, leading to more cell apoptosis.

CONCLUSION

Our study provided the first evidence that LINC-PINT promotes cell apoptosis in NPC by binding to XRCC6 and affecting its function.

摘要

背景

LINC-PINT 在鼻咽癌（NPC）中下调，并与 NPC 的治疗效率相关。然而，LINC-PINT 在 NPC 中的潜在机制尚未完全探索。

方法

我们使用细胞活力发光检测法、克隆形成实验、Hoechst 染色和 SYTO-9/PI 染色来检查 LINC-PINT 对 NPC 细胞活力和细胞凋亡的调节作用。通过异种移植肿瘤模型、HE 染色、Ki67 染色和 TUNEL 染色来评估 LINC-PINT 在体内的作用。通过生物信息学和 RNA 免疫沉淀实验来鉴定 LINC-PINT 的结合蛋白。荧光原位杂交和免疫荧光技术用于测量 XRCC6 与 LINC-PINT 和 DNA-PKcs 的共定位。Mito-Tracker red CMXRos 染色用于特异性标记细胞中的线粒体。

结果

我们发现 LINC-PINT 在许多肿瘤（包括 NPC）中下调，与不良预后相关。与对照组相比，LINC-PINT 过表达细胞的细胞活力显著抑制，细胞凋亡显著促进。与对照组相比，LINC-PINT 过表达组肿瘤异种移植的生长明显受到抑制，肿瘤重量明显降低。相应地，LINC-PINT 过表达组的阳性 Ki67 焦点减少，而 TUNEL 焦点增加。通过 RNA 结合结构域预测、RIP 和 LINC-PINT 和 XRCC6 的共定位，我们验证了 XRCC6 是 LINC-PINT 的一个新的结合蛋白。通过与 XRCC6 结合，LINC-PINT 干扰了 DNA-PK 复合物的形成，调节了线粒体积累状态，并影响了凋亡蛋白的修饰，导致更多的细胞凋亡。