PTD-FNK蛋白影响猪睾丸支持细胞抗热应激能力的转录组学分析

Transcriptomic analysis of the effect of PTD-FNK protein on heat stress resistance of boar Sertoli cells

旨在通过转录组测序和生物信息学分析评价PTD-FNK蛋白对猪睾丸支持细胞抵抗热应激能力的作用机制。将原代培养的睾丸支持细胞分为对照组、热应激(HS)组和热应激+PTD-FNK蛋白(HS+PR)组,HS+PR组添加0.1 nmol/L的PTD-FNK蛋白溶液作用1 h,对照组和HS组分别用等体积的PBS作用1 h,而后HS组和HS+PR组43℃热处理1 h,对照组37℃处理1 h。分别提取3组细胞总RNA,通过转录组测序进行分析,共获得76.75 Gb的有效数据(clean reads),各样品有效数据均达到7.11 Gb以上,测序质量较好。对照组与HS组相比共有270个差异表达基因(DEGs),其中上调基因193个,下调基因77个;HS组与HS+PR组相比共有3649个DEGs,其中上调基因1430个,下调基因2219个。基因本体(GO)注释结果显示,对照组与HS组相比以及HS组与HS+PR组相比,DEGs主要富集在细胞过程、细胞成分、结合、生物调节、代谢过程上,京都基因与基因组百科全书(KEGG)通路主要富集在磷脂酰肌醇-3-激酶-蛋白激酶B(PI3K-Akt)信号通路、丝裂原活化蛋白激酶(MAPK)信号通路以及癌症通路等。综合基因表达水平的分析,挖掘到包括分化抑制剂3(ID3)、组蛋白2A变异体(H2AX)、细胞周期蛋白依赖性激酶抑制剂1C(CDKN1C)、肿瘤抑制蛋白p53(TP53)、诱导DNA损伤的转录因子3(DDIT3)、重组酶(RAD51)等调控细胞增殖与凋亡的DEGs。本研究为分析PTD-FNK蛋白的作用提供了新的参考和理论依据。

This study was to analyze the mechanism of the PTD-FNK protein in the anti-heat stress ability of Sertoli cells in boar by transcriptome sequencing and bioinformatics analysis.Sertoli cells in primary culture were obtained and randomly divided into the control group,the heat stress group(HS)and the heat stress+protein group(HS+PR).The HS+PR group was treated with 0.1 nmol/L PTD-FNK protein solution,and the control group and the HS group were treated with an equal volume of PBS,respectively.After 1 h of treatment,the cells in the HS group and the HS+PR group were heat-treated at 43℃for 1 h,while the cells in the control group were treated at 37℃for 1 h.Then,the total RNA of the three groups were extracted respectively and a total of 76.75 Gb clean reads were obtained after analysis by transcriptome sequencing.76.75 Gb of the clean reads of the samples with each being of more than 7.11 Gb,which indicated that high quality sequencing information was obtained in this experiment.Compared with the control group,there were 270 differentially expressed genes(DEGs)in the HS group,including 193 up-regulated genes and 77 down-regulated genes.Compared with the HS group,the HS+PR group had a total of 3649 DEGs,including 1430 up-regulated genes and 2219 downregulated genes.The GO annotation analysis showed that the DEGs in the control group,as compared with those in the HS group,and those the HS group,as compared with the HS+PR group,were mainly concentrated in the cellular process,in the cellular components,and in the binding,biological regulation,and metabolic processes.The KEGG pathways were mainly concentrated on the phosphatidylinositol-3-kinase/Akt(PI3K-Akt)signaling pathway,the milasin activates protein kinase(MAPK)signaling pathway and the pathways in cancer.By the analysis of the gene expression levels,the DEGs,including the inhibitor of differentiation 3(ID3),the histone family2A variant(H2AX),cyclin dependent kinase inhibitor 1C(CDKN1C),tumor protein p53(TP53),DNA damage inducing transcript 3(DDIT3)and recombinase(RAD51)that regulated cell proliferation and apoptosis,were obtained.These results provided new reference and theoretical basis for the role analyzing of the PTD-FNK protein.