猪冷应激脂肪相关长链非编码RNA的鉴定及功能分析

doi:10.3864/j.issn.0578-1752.2026.18.016

Abstract

Abstract:

【Objective】Long non-coding RNAs (lncRNAs) are a class of RNA molecules with a length exceeding 200 nucleotides that generally do not encode proteins, and they play important regulatory roles in cold stress-induced adipose thermogenesis. This study aimed to analyze the transcriptomic data of subcutaneous adipose tissue from Tibetan pigs and Bama pigs to identify lncRNAs involved in regulating adipose metabolism under cold stress, thereby laying a foundation for further elucidation of the mechanisms underlying adipose thermogenesis in response to cold stress.【Method】Five-week-old Tibetan and Bama pigs (n = 8 per breed) were assigned to room temperature or 4 ℃ groups (four biological replicates per group). After 4 h, subcutaneous adipose tissue was collected for RNA-seq. Based on the sequencing data, lncRNAs and mRNAs were identified, and differential expression analysis was performed to compare cold stress responses between the two breeds. Target genes of differentially expressed lncRNAs were predicted via cis-regulation, followed by GO/KEGG enrichment analysis using DAVID. WGCNA was used to screen cold stress-related modules and construct co-expression networks of lncRNAs and mRNAs. Finally, RT-qPCR was performed to validate the transcriptome data of differentially expressed lncRNAs.【Result】A total of 2 988 lncRNAs were identified in the subcutaneous adipose tissue of Tibetan pigs and Bama pigs. These lncRNAs generally exhibited short transcript lengths and a small number of exons. Among them, 1 939 lncRNAs were expressed in at least one sample. Following cold stress treatment, 88 and 10 differentially expressed lncRNAs, as well as 360 and 88 differentially expressed mRNAs, were identified in the subcutaneous adipose tissue of Tibetan pigs and Bama pigs, respectively. Enrichment analysis indicated that the differentially expressed mRNAs were mainly enriched in biological processes such as HPV infection, fatty acid metabolism, and antiviral infection, as well as signaling pathways including the phosphatidylinositol 3-kinase-protein kinase B (PI3K-Akt) pathway, the peroxisome proliferator-activated receptor (PPAR) signaling pathway, and the Ras-related protein 1 (Rap1) signaling pathway. Furthermore, pairs of cis-regulated lncRNAs and protein-coding genes were screened. Among these, 108 lncRNAs were located within 10 kb upstream or downstream of 83 protein-coding genes, and 216 differentially expressed lncRNAs were located within 100 kb upstream or downstream of 289 differentially expressed genes. Finally, a co-expression network of lncRNAs and protein-coding genes was constructed using WGCNA, identifying 18 modules. Among these, the yellow module was associated with the cold stress phenotype. Key lncRNAs within this module, such as MSTRG.8042.2 and MSTRG.8043.2, showed significant associations with core lipid metabolism-related genes. It is speculated that these lncRNAs may serve as key regulatory molecules under cold stress, participating coordinately in the host's cold defense and lipid metabolism regulation. The RT-qPCR results confirmed the reliability of the transcriptome data.【Conclusion】This study identified key lncRNAs including MSTRG.8042.2 and MSTRG.8043.2 that affect cold resistance of hosts under cold stress. The results provide a reference for deciphering the molecular mechanism regulating fat metabolism upon cold stress, and offer a novel perspective for elucidating the cold adaptation mechanism in local pig breeds.

Key words: Tibetan pig, Bama pig, adipocyte, lncRNA, WGCNA

XU HaoNan, YAO Lei, PENG Cheng, JIN LinYu, CAO GuoQing, LU Chang, LI BuGao. Identification and Functional Analysis of Adipose Tissue Related Long Noncoding RNAs in Response to Cold Stress in Pigs[J].Scientia Agricultura Sinica, 2026, 59(18): 4163-4182.

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引物名称 Name	正向序列（5′→3′） Forward sequence (5′→3′)	反向序列（5′→3′） Reverse sequence (5′→3′)
MSTRG.8042.2	CTCAGAACCGTGACAAGCAC	AAAAGGCAACCCAAACTATG
MSTRG.11528.4	TGACAGTTTTCATTTTTGCACAGG	GATTGAGAGGGAGGCATGGT
MSTRG.18052.2	ACCCTTAGCAATTCCCCAGAAA	TTGTGCCTAGTCCCATCCCA
MSTRG.17619.1	GTCTTCCGAGTTGACTTGCG	GGTGGTAAACCTCCTGTCCTT
MSTRG.21051.1	GTGCTAGGGACTACGCTAAA	TACTGACACTGCCTGGTTTT
MSTRG.29467.1	TGATGCGTCCTGTTGTTGTTA	AAGGTCCTTCCTGGCGGTAT
MSTRG.13108.1	GACACAACTTTGAACGGCTGCTG	TGTCTAGTGCCCACTGCCTCTG
MSTRG.16773.1	GCTGGTTGCCGTGGTATTAGGAG	CTGGGGTTGCACACTGTGACTG
MSTRG.30235.1	AGCAGAAGGGACAGCAGTTC	GGCATTTGGGTCCTGAGTCA
MSTRG.28076.1	GGCAGAACCGTCCTGTGTAA	GAGCCCACAGAGAAGCAGTT
MSTRG.5773.1	CCCGCTGCCTGTCAGAACTTAAC	CTGAGCCACAGCAGTGCCAAC
MSTRG.16446.1	AAGATGCTGGGCTTGCGTTCG	GCTTATCCTGGGAGTGGGTGAAAC
18S	ATAAACGATGCCGACTGGCGAT	CAATCTGTCAATCCTGTCCGTGT

样本 Sample	原始序列 Raw reads	过滤后序列 Clean reads	Q20 (%)	Q30 (%)	GC含量 GC content (%)	总比对读段数 Total mapped reads	比对率 Mapping rate (%)
RT-BP1	30185424	28999302	96.40	91.99	50.30	56438442	97.31
RT-BP2	35519152	34202834	96.74	92.66	53.18	66688686	97.49
RT-BP3	28269765	27186784	96.34	91.85	50.84	52758673	97.03
RT-BP4	76021644	73196894	96.44	92.07	50.88	142206926	97.14
CE-BP1	30999664	30110740	96.97	92.57	49.82	58324503	96.85
CE-BP2	52997869	51281880	97.18	93.66	50.48	99804795	97.31
CE-BP3	38625198	37216482	96.58	92.38	49.03	72229748	97.04
CE-BP4	37319742	36246998	96.83	92.29	48.78	70079946	96.67
RT-TP1	33101118	32221914	97.10	92.86	50.09	62284960	96.65
RT-TP2	33578474	32502744	96.95	92.54	50.20	62814803	96.63
RT-TP3	31672459	30623374	96.98	92.59	49.52	59390972	96.97
RT-TP4	39844729	38634408	96.90	92.42	49.93	75198012	97.32
CE-TP1	33560307	32569774	96.92	92.47	48.63	63276557	97.14
CE-TP2	39952519	38745691	97.13	92.91	49.76	74903170	96.66
CE-TP3	33682639	32838333	97.07	92.80	49.17	63666960	96.94
CE-TP4	36206426	35228347	97.16	92.97	50.03	68180943	96.77

	长链非编码RNA lncRNAs	编码基因 Coding genes	差异表达的长链非编码RNA DElncRNAs	差异表达基因 DEGs	长链非编码RNA- 编码基因对 lncRNA-coding gene pairs	差异表达的长链非编码RNA- 差异表达基因对 DElncRNA-DEGs pairs
100 kb	1 756	5 608	216	289	7 610	120
10 kb	822	1014	108	83	1 142	43

lncRNA	P value	log₂ FC	Regulated	长度Length (bp)
MSTRG.8042	1.95E-03	3.502042800	up	1433
MSTRG.11528	5.32E-07	1.372503374	up	2364
MSTRG.18052	4.02E-09	1.940246198	up	8424
MSTRG.29467	1.80E-06	2.766504025	up	804
MSTRG.17304	3.80E-05	2.104264713	up	613
MSTRG.17619	9.56E-27	5.166865949	up	552
MSTRG.13108	7.63E-04	-1.882138759	down	571
MSTRG.16773	2.29E-03	-2.687542429	down	767
MSTRG.30235	2.04E-03	-6.41432807	down	299
MSTRG.28076	3.97E-06	-1.759364017	down	1335
MSTRG.5773	1.27E-03	-1.331161095	down	447
MSTRG.16446	2.56E-03	-1.046772722	down	767

Identification and Functional Analysis of Adipose Tissue Related Long Noncoding RNAs in Response to Cold Stress in Pigs

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