Discovery of differentially expressed genes in the intestines of Pelteobagrus vachellii within a light/dark cycle

In aquaculture, it is necessary to determine of the diurnal biological variations in the intestines to determine an appropriate feeding schedule. The present study aimed to examine the transcriptomes of the Pelteobagrus vachellii intestines at four time points (0 h, 6 h, 12 h, and 18 h) within a light/dark cycle. In comparison with the zeitgeber time 0 (ZT0) transcriptomes, we identified 37,842 unigenes with significant differential expression, including 6,638; 9,626; and 7,938 that genes upregulated, and 3,507; 4,703; and 5,412 genes that were down regulated at 4, 12, and 24 h respectively. The differentially expressed unigenes were subjected to enrichment analysis, which indicated the involvement of the major digestive pathways, including digestion of protein, lipid and carbohydrate, catabolic process (protein, carbohydrate and lipid), and circadian rhythm. We selected 73 key differentially expressed genes (DEGs) from among these pathways and identified DEGs that showed increased expression at night, including those encoding trypsin-3, chymotrypsinogen 2, amino acid transporter, maltase-glucoamylase, facilitated glucose transporter, lipase, phospholipase, fatty acid-binding protein, fatty acid synthase, long-chain fatty acid transport protein, and apolipoprotein. Moreover, DEGs involved of circadian rhythm were identified, including brain-muscle-Arnt-like 1 (BMAL1), cryptochrome-1, circadian locomoter output cycles protein kaput (CLOCK) and period circadian protein homolog 1–3. Finally, the expression levels of 12 unigenes were analyzed using quantitative real-time PCR, which were in accordance with RNA-sequencing analysis. In general, the expression of genes related to the digestion of proteins, lipids, and carbohydrates showed upregulated expression at night; however, the peak time of expression of transporters for different nutrition molecules showed more diversification within the light/dark cycle.

在水产养殖领域，需解析肠道的昼夜生物学变化规律，以制定合理的投喂方案。本研究旨在探究光暗周期内4个时间点（0 h、6 h、12 h、18 h）下瓦氏黄颡鱼（Pelteobagrus vachellii）肠道的转录组特征。与授时因子时间0（zeitgeber time 0, ZT0）的转录组相比，本研究共鉴定出37842个显著差异表达的单基因簇（unigene），其中在4 h、12 h、24 h时上调表达的基因分别为6638、9626和7938个，下调表达的基因分别为3507、4703和5412个。对上述差异表达单基因簇进行富集分析后发现，其显著富集于主要的消化通路，包括蛋白质、脂质与碳水化合物消化过程，蛋白质、碳水化合物及脂质的分解代谢过程，以及昼夜节律通路。本研究从上述通路中筛选出73个关键差异表达基因（differentially expressed genes, DEGs），并鉴定出夜间表达上调的DEGs，其编码的蛋白包括胰蛋白酶-3（trypsin-3）、胰凝乳蛋白酶原2（chymotrypsinogen 2）、氨基酸转运蛋白、麦芽糖酶-葡糖淀粉酶、易化葡萄糖转运蛋白、脂肪酶、磷脂酶、脂肪酸结合蛋白、脂肪酸合酶、长链脂肪酸转运蛋白以及载脂蛋白。此外，本研究还鉴定出参与昼夜节律调控的DEGs，包括脑-肌肉-Arnt样蛋白1（brain-muscle-Arnt-like 1, BMAL1）、隐花色素-1（cryptochrome-1）、昼夜节律运动输出周期蛋白kaput（CLOCK）以及周期节律蛋白同源物1~3。最后，本研究采用实时荧光定量PCR对12个单基因簇的表达水平进行验证，结果与RNA测序分析结果一致。总体而言，参与蛋白质、脂质与碳水化合物消化的基因在夜间呈现表达上调趋势；但不同营养物质转运蛋白的表达峰值在光暗周期内呈现出更为多样的分布特征。