富营养饮食重塑代谢性疾病易感猪关键组织mRNA剪接模式研究

Study on the mRNA Splicing Pattern Reshaped by the Nutritious Diet in Key Tissues of Metabolic Disease Susceptible Pigs

旨在探讨富营养饮食对代谢性疾病易感猪关键组织可变剪接(alternative splicing,AS)和剪接因子(splicing factors,SFs)的影响。本研究以健康状态、月龄和体重相同的12头代谢性疾病易感公猪为材料,随机将其分为两组,其中8头猪饲喂高脂高能量饲料2个月作为富营养饮食组(TG-HFHSD),4头猪饲喂正常饲料2个月作为对照饮食组(TG-CD)。对TG-HFHSD和TG-CD的脂肪、肝、肌肉和下丘脑组织进行RNA-seq测序,利用rMATS软件鉴定组间差异AS,对差异剪接基因(differential splicing genes,DSGs)进行GO和KEGG富集分析,并利用DESeq2软件和皮尔逊相关性系数分析肝组织中的SFs。结果显示,脂肪、肝、肌肉和下丘脑中分别鉴定到48279、39536、47888和52210个AS事件,均以SE类型为主,占比在77.09%~78.29%之间。脂肪、肝、肌肉和下丘脑中的DSGs数分别为528、1070、570和600个,其中肝增加的DSGs部分主要是RI和MXE两种类型,分别为177和471个,而其它组织仅为12~28个和99~110个。GO和KEGG富集显示,肝DSGs除富集到细胞结构相关的过程外,还显著富集到可变剪接及mRNA代谢、脂质代谢、DNA损伤及修复、补体与凝血级联等过程。肝中筛选到56个差异表达的SFs,其中CLK1、PGC-1ɑ、USP4等与糖脂代谢和代谢性疾病密切相关。本研究表明,富营养饮食显著增加了肝DSGs,通过剪接调控、糖脂代谢以及DNA损伤来影响肝代谢性疾病的发生,将为肝代谢性疾病早期发生机制研究提供参考。

The objective of this study was to investigate the effects of nutritious diet on alternative splicing(AS)and splicing factors(SFs)in key tissues of metabolic disease susceptible pigs.Twelve metabolic disease susceptible male pigs with the same health status,age and body weight were randomly divided into 2groups.Eight pigs were fed a high fat and energy diet for 2months as the nutrient-rich diet group(TG-HFHSD),and 4pigs were fed a normal diet for 2months as the control diet group(TG-CD).The adipose,liver,muscle,and hypothalamus tissues of TG HFHSD and TG-CD were sequenced by RNA-seq,and the differential AS between the groups was identified by rMATS software,GO and KEGG enrichment analysis of differential splicing genes(DSGs),and SFs in liver was analyzed by DESeq2software and pearson correlation coefficient.The results showed that 48279,39536,47888and 52210AS events were identified in adipose,liver,muscle,and hypothalamus,respectively,all of which were mainly of SE type,accounting for 77.09%-78.29%.The number of DSGs in adipose,liver,muscle,and hypothalamus were 528,1070,570,and 600,respectively.The increased DSGs in the liver mainly came from RI and MXE with 177and 471,respectively,while only 12-28and 99-110for other tissues.The enrichment of GO and KEGG showed that,in addition to the processes related to cell structure,the DSGs in liver,was also significantly enriched in processes such as alternative splicing and mRNA metabolism,lipid metabolism,DNA damage and repair,complement and coagulation cascades.Fifty-six differentially expressed SFs were screened from liver,among which CLK1,PGC-1ɑ,USP4and others were closely related to glucolipid metabolism and metabolic diseases.The results of this study showed that nutritious diet significantly increased DSGs in liver and affected the occurrence of liver metabolic diseases through splicing regulation,glucolipid metabolism and DNA damage,which will provide reference for the study of the early pathogenesis of liver metabolic diseases.