LIN28A在GT1-7细胞中过表达对性发育相关基因的影响

Effect on puberty-related genes with LIN28A overexpression in GT1-7 cells

[目的]在GT1-7细胞过表达LIN28A,研究其与性发育相关基因表达是否存在调控关系; RNA-seq发现LIN28A参与调控性发育的信号通路和基因。[方法]构建LIN28A慢病毒过表达载体,转染GT1-7细胞,Real-Time PCR检测LIN28A和性发育相关基因在mRNA水平表达变化;将过表达LIN28A的GT1-7细胞进行RNA-seq、生物信息学分析、Real-Time PCR验证,找到影响性发育的信号通路和基因。[结果]与对照相比,LIN28A在GT1-7细胞过表达18倍(P <0. 001),KISS1在mRNA水平显著升高(P <0. 01); KEGG分析,LIN28A相关的差异基因在MAPK信号通路差异显著,MAPK通路筛选到多个性发育相关基因(Fgf21、JUN、Cyp1a1、Rasgrp2),Real-Time PCR验证它们在mRNA水平差异显著(P <0. 05)。[结论]成功构建LIN28A慢病毒过表达载体,LIN28A在GT1-7细胞中过表达会促进KISS1的表达,并且LIN28A可能通过MAPK通路调控性发育。

[Objective]LIN28 A was overexpressed in GT1-7 cells to investigate whether there was a regulatory relationship between LIN28 A and gene expression related to puberty. Through RNA-seq,the signaling pathways and genes that LIN28 A regulates puberty were discovered.[Method]LIN28 A lentivirus overexpression vector was constructed. Then the vector was transfected into GT1-7 cells,and Real-Time PCR was performed to detect the mRNA expression changes of LIN28 A and puberty-related genes. GT1-7 cells overexpressing LIN28 A were verified by RNA-seq,bioinformatics analysis and Real-Time PCR to find the signaling pathways and genes affecting puberty.[Result]Compared with the control group,LIN28 A was overexpressed in GT1-7 cells 18 times( P < 0. 001),and KISS1 was significantly increased in mRNA level( P < 0. 01). KEGG analysis showed that LIN28 A-related differentially expressed genes showed significant differences in MAPK signaling pathways.MAPK pathway was used to screen out multiple genes related to puberty( Fgf21,JUN,Cyp1 a1,Rasgrp2),and Real-Time PCR verified that they had significant differences in mRNA levels( P < 0. 05).[Conclusion]LIN28 A lentivirus overexpression vector was successfully constructed. Overexpression of LIN28 A in GT1-7 cells promotes KISS1 expression,and LIN28 A may regulate pubertythrough MAPK pathway.