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在HEK293T细胞中LGR5Δ5拼接变体的功能和生物特性

Matthias Kappler¹, Laura Thielemann¹, Markus Glaß²

¹Department of Oral and Maxillofacial Plastic Surgery, Faculty of Medicine, Martin Luther University Halle-Wittenberg, 06120 Halle, Germany.

International journal of molecular sciences

|January 8, 2025

概括

这种LGR5Δ5异型,是Wnt通路调节器LGR5的变体,不恢复Wnt活性,但增强了放射敏感性. LGR5Δ5影响细胞骨和血管生成基因,与全长LGR5.5不同.

科学领域:

分子生物学分子生物学
细胞生物学细胞生物学
癌症研究癌症研究

背景情况:

氨酸丰富的重复含有G蛋白结合受体5 (LGR5) 是Wnt通路的关键调节者,在干细胞中发现,在癌症中过度表达.
之前确定的LGR5Δ5异型与口腔平细胞癌 (OSCC) 和软组织肉瘤 (STS) 的预后和转移有关.

研究的目的:

调查LGR5Δ5异形与全长LGR5 (LGR5FL) 相比在Wnt通路活性,细胞迁移和放射敏感化中的LGR5Δ5异形的独特功能.
使用HEK293T细胞模型阐明LGR5Δ5和LGR5FL功能背后的分子机制.

主要方法:

使用CRISPR/Cas9基因编辑来淘汰LGR5和LGR4.
使用HEK293T细胞进行功能测试,包括Wnt通路活性评估,细胞迁移研究和放射敏感化实验.
进行RNA测序以分析由LGR5异型诱导的基因表达变化.

主要成果:

CRISPR/Cas9对LGR5/LGR4的淘汰消除了Wnt活动,而LGR5Δ5无法恢复该活动,与LGR5FL不同.
过度表达LGR5FL减少了细胞迁移,而LGR5Δ5没有影响.
克里斯普尔/卡斯9淘汰诱导的辐射敏感化,通过LGR5FL或LGR5Δ5过度表达进一步增强.
LGR5Δ5显著增加了R-脊蛋白1 (RSPO1) 水平和调节了参与细胞骨,细胞外基因和血管生成的基因.
LGR5FL与对原蛋白和组织蛋白的调节有关.

结论:

LGR5Δ5的功能与LGR5FL不同,特别是不恢复Wnt通路的活性,但有助于放射敏感化.
LGR5Δ5在调节与细胞结构,基因相互作用和血管形成相关的基因方面发挥着作用.
LGR5FL涉及调节原蛋白和组织蛋白,这表明它在细胞过程和潜在的癌症进展中发挥着不同的作用.

关键词:

这就是CRISPR/CAS.在LGR4LGR4LGR4 在LGR5中,LGR5是指LGR5.在LGR5Δ5中.这种异形异形是什么?干细胞标志物是干细胞的标志物.

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