鸡节律相关基因家族生物信息学及表达分析

doi:10.16431/j.cnki.1671-7236.2025.12.020

摘要/Abstract

摘要： 【目的】试验旨在系统分析鸡节律相关基因家族，探究鸡在持续黑暗条件下松果体内源性节律基因的表达规律，并分析鸡松果体部分节律基因在胚胎后期到育雏初期的表达模式。【方法】通过查询InterPro数据库，获取多种节律相关基因家族序列，构建系统进化树，分析其蛋白理化性质、保守结构域、顺式作用元件等。基于转录组数据，采用RAIN算法与PSEA分析内源性节律基因的表达特征，并采用实时荧光定量PCR分析不同阶段燕山红玉鸡与巴布考克B380鸡松果体中节律基因的表达模式。【结果】共鉴定到7个节律相关基因家族(PER、CRY、CLOCK、BMAL、ROR、TIMELESS、OPN4)。系统进化树显示，CLOCK与BMAL基因在进化中较为接近。染色体定位分析发现，各基因家族成员分散分布在不同染色体上，仅发现OPN4与RRH基因存在串联重复。蛋白理化性质分析显示，PER3和TIMELESS蛋白分子质量＞130 ku，其他家族成员蛋白分子质量＜100 ku。OPN4及其亚型OPN4-1为疏水性且含7次跨膜结构域(7tm_GPCRs)，而其他蛋白均为亲水性且无跨膜结构域。RAIN算法结果显示，CRY1与CRY2基因的表达相位存在明显差异，且二者启动子区转录因子结合位点及密度存在差异。PSEA分析结果发现，在持续黑暗的条件下，主观夜高表达基因CRY2、PER3参与昼夜节律调控及损伤DNA结合、能量代谢、脂质代谢等通路；主观日高表达基因CLOCK、ARNTL、CRY1则参与节律基因转录激活、DNA修复及损伤响应等通路。实时荧光定量PCR结果显示，CRY2、PER2和CLOCK基因表达水平在出雏第1天主观日2 h较孵化第19天显著上调(P＜0.05)，随后在出雏第10天较出雏第1天显著下调(P＜0.05)；而TIMELESS和OPN4基因表达水平均较前一阶段持续显著上调(P＜0.05)。【结论】鸡节律相关基因家族在染色体上分布较为分散，OPN4与RRH基因存在串联重复，CRY基因表达相位差异可能受启动区调控元件影响。出雏第1天，光照刺激可能显著上调雏鸡松果体CRY2、PER2等核心节律基因表达；至出雏第10天，生物钟调控网络趋于成熟。

关键词: 鸡; 节律相关基因家族; 生物信息学; 表达

Abstract: 【Objective】 This experiment aimed to systematically analyze the circadian rhythm-related gene family in chickens,explore the expression patterns of endogenous rhythm genes in the pineal gland of chickens under the constant darkness conditions,and analyze the expression patterns of partial rhythm genes in pineal gland of chickens from late embryonic stage to early chick rearing stage. 【Method】 Various rhythm-related gene family sequences were obtained from InterPro database,the phylogenetic tree was constructed,and the physicochemical properties,conserved domains,and cis-regulatory elements were analyzed.Based on transcriptome data,the RAIN algorithm and PSEA were used to analyze the expression characteristics of endogenous rhythm genes.Finally,Real-time quantitative PCR was used to analyze the expression patterns of rhythm genes in pineal gland of Yanshan Hongyu chickens and Babcock B380 chickens at different stages. 【Result】 Seven rhythm-related gene families (PER,CRY,CLOCK,BMAL,ROR,TIMELESS,and OPN4) were identified.Phylogenetic tree showed that CLOCK and BMAL genes were relatively close in evolution.Chromosome localization analysis revealed that the members of each gene family were dispersed across different chromosomes,and OPN4 gene was tandemly repeated with RRH gene.Protein physicochemical analysis showed that the molecular weight of PER3 and TIMELESS proteins exceeded 130 ku,while the other family members were smaller than 100 ku.OPN4 and its subtype OPN4-1 were hydrophobic and contained 7 transmembrane domain (7tm_GPCRs),while the other proteins were hydrophilic and lack transmembrane domains.RAIN algorithm results showed significant phase differences in the expression of CRY1 and CRY2 genes,with differences in the transcription factor binding sites and density in their promoter regions.PSEA analysis results found that under the constant darkness conditions,the genes CRY2 and PER3,which exhibited high expression during the subjective night,were involved in circadian rhythm regulation and pathways related to DNA damage binding,energy metabolism,and lipid metabolism.The genes CLOCK,ARNTL,and CRY1,which exhibited high expression during the subjective day,participated in circadian gene transcription activation,DNA repair,and damage response pathways.Real-time quantitative PCR results showed that at subjective day 2 h on incubation days 19,and on day 1 and days 10 post-hatching,the expression of CRY2,PER2,and CLOCK genes firstly significantly increased and then significantly decreased (P＜0.05),while the expression of TIMELESS and OPN4 genes continuously increased (P＜0.05). 【Conclusion】 The circadian rhythm-related gene familiy in chickens was widely dispersed on chromosomes,and there was tandem duplication between OPN4 and RRH genes.The phase difference of CRY gene expression might be influenced by regulatory elements in their promoter regions.On day 1 post-hatching,light stimulation might significantly upregulate the expression of core rhythm genes such as CRY2 and PER2 in pineal gland of chickens.By days 10 post-hatching,the circadian regulatory network tended to mature.

Key words: chickens; circadian rhythm-related gene family; bioinformatics; expression

中图分类号:

S814.8

葛华梁, 任敏鹏, 张夕霏, 杨姣, 耿立英, 张传生, 李祥龙. 鸡节律相关基因家族生物信息学及表达分析[J]. 中国畜牧兽医, 2025, 52(12): 5749-5761.

GE Hualiang, REN Minpeng, ZHANG Xifei, YANG Jiao, GENG Liying, ZHANG Chuansheng, LI Xianglong. Bioinformatics and Expression Analysis of Circadian Rhythm-related Gene Family in Chickens[J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(12): 5749-5761.

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