猪白色与米色脂肪细胞的三维基因组结构解析

doi:10.11843/j.issn.0366-6964.2025.11.008

畜牧兽医学报 ›› 2025, Vol. 56 ›› Issue (11): 5433-5448.doi: 10.11843/j.issn.0366-6964.2025.11.008

猪白色与米色脂肪细胞的三维基因组结构解析

蒋一帆¹(), 李昊星²^,³^,⁴(), 林育¹, 张治华²^,³^,⁴, 王彦芳¹^,*()

1. 中国农业科学院北京畜牧兽医研究所, 北京 100193
2. 国家生物信息中心, 北京 100101
3. 中国科学院北京基因组研究所, 北京 100101
4. 中国科学院大学生命科学学院, 北京 100049

收稿日期:2025-04-08 出版日期:2025-11-23 发布日期:2025-11-27
通讯作者: 王彦芳 E-mail:jiang092122@163.com;1076599497@qq.com;wangyanfang@caas.cn
作者简介:蒋一帆(1999-)，女，重庆人，硕士，主要从事猪遗传育种研究，E-mail：jiang092122@163.com
李昊星(1996-)，男，云南昆明人，博士，主要从事生物信息学研究，E-mail: 1076599497@qq.com
第一联系人：
蒋一帆和李昊星为同等贡献作者
基金资助:
国家重点研发计划(2020YFA0509503)；国家杰出青年科学基金(32025034)；中国农业科学院科技创新工程(ASTIP-IAS05)；中央级公益性科研院所基本科研业务费专项资金项目(2024-YWF-ZYSQ-06)

Three-Dimensional Genome Architecture of Porcine White and Beige Adipocytes

JIANG Yifan¹(), LI Haoxing²^,³^,⁴(), LIN Yu¹, ZHANG Zhihua²^,³^,⁴, WANG Yanfang¹^,*()

1. Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing 100193, China
2. China National Center for Bioinformation, Beijing 100101, China
3. Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 100101, China
4. College of Life Sciences, University of Chinese Academy of Sciences, Beijing 100049, China

Received:2025-04-08 Online:2025-11-23 Published:2025-11-27
Contact: WANG Yanfang E-mail:jiang092122@163.com;1076599497@qq.com;wangyanfang@caas.cn

摘要/Abstract

摘要：

旨在从三维基因组层面探究猪白色与米色脂肪细胞的差异，并鉴定出两种脂肪细胞的特异性启动子-增强子互作(promoter-enhancer interactions，PEIs)，筛选出促进猪米色脂肪细胞分化的关键增强子。本研究分化了猪白色与米色脂肪细胞，利用高通量染色体构象捕获(high-throughput chromosome conformation capture, Hi-C)技术比对了两种脂肪细胞在染色质区室、拓扑关联结构域(TADs)以及染色质环(loops)的差异，对猪白色与米色脂肪细胞的三维基因组构象进行详细解析；结合全基因组H3K27ac与H3K4me3的ChIP-seq数据鉴定两种脂肪细胞的PEIs，筛选促进猪米色脂肪细胞分化的潜在增强子，构建双荧光素酶载体验证增强子活性。Hi-C数据展示了两种脂肪细胞的三维基因组结构差异，有5%的基因组区域存在染色质区室状态的不同，鉴定到白色脂肪细胞特异性TADs 1 923个，特异性loops 12 920个；米色脂肪细胞特异性TADs 2 010个，特异性loops 9 526个。ChIP-seq结果显示鉴定到白脂特异性PEIs 795个，米脂特异性PEIs 737个。结合转录组数据，筛选出在米脂中特异性高表达且存在特异性PEIs基因的增强子作为促进米色脂肪分化的关键增强子。进一步，构建了TMEM100基因525 bp与AMCF-Ⅱ基因200 bp潜在增强子片段的双荧光素酶报告质粒，荧光素酶活性检测结果显示，两个基因的潜在增强子片段对报告质粒荧光活性有显著上调作用(P < 0.05)。本研究构建了猪白色与米色脂肪细胞的三维基因组图谱，揭示了两种脂肪细胞间的染色质空间结构差异，初步验证了TMEM100与AMCF-Ⅱ基因的增强子活性，提示TMEM100与AMCF-Ⅱ基因的增强子可能通过PEIs调控进而促进猪米色脂肪细胞分化。

关键词: 猪, 米色脂肪细胞, Hi-C, 三维基因组, 增强子

Abstract:

This study aimed to investigate the differences between porcine white and beige adipocytes at the three-dimensional (3D) genome level, identify cell type-specific promoter-enhancer interactions (PEIs) in both adipocyte types, and screen for critical enhancers that promote the differentiation of porcine beige adipocytes. This study differentiated porcine white and beige adipocytes, and employed high-throughput chromosome conformation capture (Hi-C) to compare differences in chromatin compartments, topologically associating domains (TADs), and chromatin loops between the two adipocyte types. We comprehensively resolved the 3D genome architecture of porcine adipocytes. Using genome-wide H3K27ac and H3K4me3 ChIP-seq data, the study identified promoter-enhancer interactions (PEIs) in both adipocyte types and screened for potential enhancers that promote the differentiation of pig beige adipocytes. The activity of these enhancers was further validated through the construction of dual-luciferase reporter vectors. The Hi-C data revealed divergent 3D genome architectures between the two adipocyte types, with chromatin compartment switching occurring in 5% of genomic regions. A total of 1 923 white adipose-specific TADs (topologically associating domains) and 12 920 white adipose-specific loops were identified, while 2 010 beige adipose-specific TADs and 9 526 beige adipose-specific loops were also detected. ChIP-seq results revealed the identification of 795 white adipose-specific PEIs and 737 beige adipose-specific PEIs. By integrating transcriptomic data, we identified gene enhancers that are specifically highly expressed in beige adipocytes and exhibit cell type-specific PEIs, which were prioritized as critical enhancers promoting beige adipocyte differentiation. Furthermore, dual-luciferase reporter plasmids carrying 525 bp (TMEM100 gene) and 200 bp (AMCF-Ⅱ gene) candidate enhancer fragments were constructed. Luciferase activity assays demonstrated that these putative enhancers upregulated luciferase activity significantly (P < 0.05). This study constructed three-dimensional genome maps of porcine white and beige adipocytes, revealing differences in chromatin spatial structures between the two types of adipocytes. We validated the enhancer activity of the TMEM100 and AMCF-Ⅱ genes preliminarily, suggesting that the enhancers of both genes may promote the differentiation of porcine beige adipocytes through PEIs regulation.

Key words: pig, beige adipocyte, Hi-C, 3D genome, enhancer

中图分类号:

S828.2

蒋一帆, 李昊星, 林育, 张治华, 王彦芳. 猪白色与米色脂肪细胞的三维基因组结构解析[J]. 畜牧兽医学报, 2025, 56(11): 5433-5448.

JIANG Yifan, LI Haoxing, LIN Yu, ZHANG Zhihua, WANG Yanfang. Three-Dimensional Genome Architecture of Porcine White and Beige Adipocytes[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(11): 5433-5448.

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基因 Gene	上游引物(5′→3′) Forward primer	下游引物(5′→3′) Reverse primer
18S rRNA	GTAACCCGTTGAACCCCATT	CCATCCAATCGGTAGTAGCG
FABP4	GGCCAAACCCAACCTGATCA	CATCCCACTTCTGCACCTGT
PPARg	GTCATGGGTGAAACTCTGGGA	TGTCAACCATGGTCACCTCTTG
UCP3	CAACAGGAAGTACAGCGGGA	GTGATGTTGGGCAGAATTCCTTT
CIDEA	GTCAAGGCCACCATGTACGA	AGCATTCGGAGCATGTACGT
PDK4	GGCAGCAGTGGTCCAAGAT	CGACTGTAGCCCTCATTGCA
LEPTIN	TGGCCCTATCTGTCCTACGT	TGGACAGACTCAGGACAGGA
ADCY5	CAGACATCAACGCCAAGCAG	AAGCCCTCGATGTCAGCAAA
TMEM100	ACCTGTGCTGTGTTGTGTTTG	TATCGCCCATTCACTGCACA
AMCF-Ⅱ	AGGCGGCTGTAGTGAGAGA	CCATTCTTCAGGGTGGCTATCA

功能元件名称及基因组定位 Name and genomic localization of functional elements	上游引物(5′→3′) Forward primer	下游引物(5′→3′) Reverse primer
TMEM100 Promoter (chr12:3212541-32016094)	CTATCTGATGCCCTAGACCT	CAAATTGTAAATCAACTATACTCC
TMEM100 Enhancer 200 bp (chr12:31945275-31945473)	GGGAAGAAGACAGGGTGGAA	CTGTGTCTTAGTTTATTGCCAT
TMEM100 Enhancer 525 bp (chr12:31945100-31945623)	GCCTCCAAATCTTCTTTTGAG	TGGGAGTCACCAGTGTGGG
AMCF-Ⅱ Promoter (chr8:70006225-70008224)	TTGGTGCCCAAACTAATTCA	AAGGCGCGGAGATTTGAAG
AMCF-Ⅱ Enhancer 250 bp (chr8:70102263-70102550)	GACTGCACTTTCTTAGGTTTA	TTGTTTTAGCCAAAGGAATCT
AMCF-Ⅱ Enhancer 500 bp (chr8:70102150-70102650)	TTTTCCATCTGTAATGGAGCC	ACCTAGGGATTTAGCACAGG

基因ID Gene ID	基因名称 Gene name	差异倍数 log₂FC	基因ID Gene ID	基因名称 Gene name	差异倍数 log₂FC
ENSSSCG00000009169	SLC39A8	5.523	ENSSSCG00000009120	ZGRF1	2.249
ENSSSCG00000004597	AQP9	4.990	ENSSSCG00000007603	NPTX2	2.209
ENSSSCG00000012129	GLRA2	4.873	ENSSSCG00000060144	CCNA2	2.146
ENSSSCG00000060220	NA	4.625	ENSSSCG00000011624	MCM2	2.111
ENSSSCG00000059641	NA	4.278	ENSSSCG00000003888	STIL	2.105
ENSSSCG00000009543	MYO16	4.259	ENSSSCG00000012150	REPS2	2.073
ENSSSCG00000053166	NA	3.858	ENSSSCG00000039760	PRR16	2.046
ENSSSCG00000037821	RGS5	3.823	ENSSSCG00000059773	NA	2.043
ENSSSCG00000053781	NA	3.376	ENSSSCG00000004157	IL20RA	2.007
ENSSSCG00000025240	DDIT4L	3.353	ENSSSCG00000035355	F2R	1.895
ENSSSCG00000013273	CHST1	3.052	ENSSSCG00000009446	PCDH17	1.779
ENSSSCG00000012540	SLC25A53	2.898	ENSSSCG00000015959	RAPGEF4	1.770
ENSSSCG00000015961	CDCA7	2.726	ENSSSCG00000041777	K7GN43	1.734
ENSSSCG00000030016	PDE9A	2.655	ENSSSCG00000056445	NA	1.715
ENSSSCG00000026527	HCN1	2.471	ENSSSCG00000010647	ADRB1	1.706
ENSSSCG00000025965	SPDL1	2.356	ENSSSCG00000039027	TRO	1.701
ENSSSCG00000024325	SGK2	2.323	ENSSSCG00000023434	PPM1L	1.541
ENSSSCG00000015136	UBASH3B	2.294	ENSSSCG00000017254	MAP2K6	1.517
ENSSSCG00000034229	MIS18A	2.274	ENSSSCG00000010912	KIF14	1.509

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猪白色与米色脂肪细胞的三维基因组结构解析

Three-Dimensional Genome Architecture of Porcine White and Beige Adipocytes

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