Genes and pathways related to piglet diarrhea in miRNAs-MUC13 regulation system

doi:10.3969/j.issn.1004-1524.20230088

Acta Agriculturae Zhejiangensis ›› 2023, Vol. 35 ›› Issue (12): 2785-2793.DOI: 10.3969/j.issn.1004-1524.20230088

• Animal Science • Previous Articles Next Articles

Genes and pathways related to piglet diarrhea in miRNAs-MUC13 regulation system

LU Fuzeng(), HUA Weidong, CHU Xiaohong, DAI Lihe, CHEN Xiaoyu, ZHANG Lifeng, WANG Zhigang, XU Ruhai()

Institute of Animal Husbandry and Veterinary Medicine, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China

Received:2023-01-29 Online:2023-12-25 Published:2023-12-27

Abstract

Abstract:

In order to discover the molecular regulatory mechanism related to piglet diarrhea, the genes and pathways related to piglet diarrhea in the microRNAs-Mucin13 (miRNAs-MUC13) regulatory system were explored and verified. NCBI blastn tool was used to compare the nucleic acid sequences of mouse MUC13 gene and pig gene bank, and found that the similarity between them was 74%-87%, with high homology. Taking the mouse as a model, the miRDB, mirTarBase and TargetScan databases were selected to predict the miRNA sequence that regulated the MUC13 gene. The target genes of these miRNA sequences were predicted through the StarBase database, and 36 target gene intersections were obtained. The interaction network of piglet diarrhea genes was built around MUC13 and these 36 target genes by using Cytoscape software. The network contains 19 screened target genes, and 4 molecular complex groups were obtained based on MCOMD algorithm analysis. The gene and pathway enrichment analysis of these 4 molecular complexes were carried out through DAVID platform. The results showed that the target genes SRF and STAG2 and their involved pathways were related to piglet diarrhea. Using Escherichia coli strain 200 and piglet intestinal epithelial cell line (IPEC-1) as the test materials, the piglet diarrhea model was designed and verified by the in vitro cell test. The real-time fluorescence quantitative PCR (qRT-PCR) results showed that the relative expressions of MUC13, SRF, IL-6, MAPK, STAG2, NF-κB, TLR7 in the test group were significantly higher than those of the control group (P<0.05), so it was speculated that SRF and STAG2 genes were involved in the regulation of piglet diarrhea through MAPK, IL 6 and Toll-like receptor signal pathway in the miRNAs-MUC13 regulation system. This study explored 4 molecular complex groups and 2 core genes (SRF and STAG2) around the miRNAs-MUC13 regulatory system, and carried out experimental verification, which provided theoretical reference and scientific basis for targeted treatment of piglet diarrhea.

Key words: Mucin 13, microRNA, piglet diarrhea, gene network

CLC Number:

S828

LU Fuzeng, HUA Weidong, CHU Xiaohong, DAI Lihe, CHEN Xiaoyu, ZHANG Lifeng, WANG Zhigang, XU Ruhai. Genes and pathways related to piglet diarrhea in miRNAs-MUC13 regulation system[J]. Acta Agriculturae Zhejiangensis, 2023, 35(12): 2785-2793.

Figures/Tables 7

References 30

[1]	孙宏伟, 王泽岩, 任少敏, 等. 猪主要腹泻病的发病机制综述[J]. 中国动物检疫, 2016, 33(3): 63-66.
	SUN H W, WANG Z Y, REN S M, et al. Summary on the pathogenesis of porcine primary diarrhea diseases[J]. China Animal Health Inspection, 2016, 33(3): 63-66. (in Chinese with English abstract)
[2]	任军, 晏学明, 艾华水, 等. 仔猪断奶前腹泻抗病基因育种技术的创建及应用[J]. 猪业科学, 2012, 29(1): 44-48.
	REN J, YAN X M, AI H S, et al. Establishment and application of disease-resistant gene breeding technology for diarrhea in piglets before weaning[J]. Swine Industry Science, 2012, 29(1): 44-48. (in Chinese)
[3]	方宇瑜, 吴艳, 高硕, 等. 苏淮猪群体抗腹泻基因MUC13和FUT1多态性分析及其抗腹泻选育方案研究[J]. 畜牧与兽医, 2015, 47(12): 12-17.
	FANG Y Y, WU Y, GAO S, et al. Polymorphism analysis of anti-diarrhea genes MUC13 and FUT1 in Suhuai pigs and its selective breeding on anti-diarrhea traits[J]. Animal Husbandry & Veterinary Medicine, 2015, 47(12): 12-17. (in Chinese with English abstract)
[4]	ZHANG X Q, LI C, ZHANG B Z, et al. Differential expression and correlation analysis of miRNA-mRNA profiles in swine testicular cells infected with porcine epidemic diarrhea virus[J]. Scientific Reports, 2021, 11: 1868.
[5]	WU Z C, QIN W Y, WU S, et al. Identification of microRNAs regulating Escherichia coli F18 infection in Meishan weaned piglets[J]. Biology Direct, 2016, 11(1): 1-19.
[6]	SHANNON P, MARKIEL A, OZIER O, et al. Cytoscape: a software environment for integrated models of biomolecular interaction networks[J]. Genome Research, 2003, 13(11): 2498-2504.
[7]	TUIKKALA J, VÄHÄMAA H, SALMELA P, et al. A multilevel layout algorithm for visualizing physical and genetic interaction networks, with emphasis on their modular organization[J]. BioData Mining, 2012, 5: 2.
[8]	HUANG D W, SHERMAN B T, LEMPICKI R A. Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources[J]. Nature Protocols, 2009, 4(1): 44-57.
[9]	DAWIDOWSKA M, JAKSIK R, DROBNA M, et al. Comprehensive investigation of miRNome identifies novel candidate miRNA-mRNA interactions implicated in T-cell acute lymphoblastic leukemia[J]. Neoplasia, 2019, 21(3): 294-310.
[10]	WEBER M J. New human and mouse microRNA genes found by homology search[J]. Genome, 2006, 49(10): 1283-1286.
[11]	KIM J, CHO I S, HONG J S, et al. Identification and characterization of new microRNAs from pig[J]. Mammalian Genome, 2008, 19(7/8): 570-580.
[12]	王伟, 滚双宝, 王鹏飞, 等. 猪miR-204组织表达与重要靶基因筛选[J]. 浙江农业学报, 2020, 32(9): 1564-1573.
	WANG W, GUN S B, WANG P F, et al. Tissue expression and significant target genes analysis of swine miR-204[J]. Acta Agriculturae Zhejiangensis, 2020, 32(9): 1564-1573. (in Chinese with English abstract)
[13]	朱静静, 周晓龙, 汪涵, 等. 靶向猪内质网应激通路的microRNAs预测与验证[J]. 中国农业科学, 2020, 53(15): 3169-3179.
	ZHU J J, ZHOU X L, WANG H, et al. Prediction and verification of microRNAs targeting porcine endoplasmic reticulum stress pathway[J]. Scientia Agricultura Sinica, 2020, 53(15): 3169-3179. (in Chinese with English abstract)
[14]	楚丹, 冉茂良, 卞桥, 等. miR-139进化分析、靶基因功能预测以及在猪睾丸组织中的表达分析[J]. 中国畜牧杂志, 2022, 58(4): 77-83.
	CHU D, RAN M L, BIAN Q, et al. Evolution analysis, target gene function prediction and expression analysis of miR-139 in pig testis[J]. Chinese Journal of Animal Science, 2022, 58(4): 77-83. (in Chinese)
[15]	张名媛, 郭晓萍, 孙晴超, 等. 广西巴马小型猪miR-148b慢病毒制备及靶基因通路预测分析[J]. 南方农业学报, 2019, 50(7): 1596-1604.
	ZHANG M Y, GUO X P, SUN Q C, et al. Construction of miR-148b lentivirus vector and prediction of target gene pathways in Guangxi Bama mini-pig[J]. Journal of Southern Agriculture, 2019, 50(7): 1596-1604. (in Chinese with English abstract)
[16]	HUANG Y A, LIN L Y, YU X T, et al. Functional involvements of heterogeneous nuclear ribonucleoprotein A1 in smooth muscle differentiation from stem cells in vitro and in vivo[J]. Stem Cells, 2013, 31(5): 906-917.
[17]	BORENSZTEIN M, MONNIER P, COURT F, et al. Myod and H19-Igf2 locus interactions are required for diaphragm formation in the mouse[J]. Development, 2013, 140(6): 1231-1239.
[18]	TRITSCH E, MALLAT Y, LEFEBVRE F, et al. An SRF/miR-1 axis regulates NCX1 and Annexin A5 protein levels in the normal and failing heart[J]. Cardiovascular Research, 2013, 98(3): 372-380.
[19]	VULTAGGIO A, NENCINI F, PRATESI S, et al. The TLR7 ligand 9-benzyl-2-butoxy-8-hydroxy adenine inhibits IL-17 response by eliciting IL-10 and IL-10-inducing cytokines[J]. The Journal of Immunology, 2011, 186(8): 4707-4715.
[20]	VULTAGGIO A, NENCINI F, FITCH P M, et al. Modified adenine (9-benzyl-2-butoxy-8-hydroxyadenine) redirects Th2-mediated murine lung inflammation by triggering TLR7[J]. The Journal of Immunology, 2009, 182(2): 880-889.
[21]	ZHU H, LU W, LAURENT C, et al. Genes encoding catalytic subunits of protein kinase A and risk of spina bifida[J]. Birth Defects Research. Part A, Clinical and Molecular Teratology, 2005, 73(9): 591-6.
[22]	LEE N, BATT M K, CRONIER B A, et al. Ciliary neurotrophic factor receptor regulation of adult forebrain neurogenesis[J]. The Journal of Neuroscience: the Official Journal of the Society for Neuroscience, 2013, 33(3): 1241-1258.
[23]	YANG C X, ZHANG K, ZHANG A X, et al. Co-expression network modeling identifies specific inflammation and neurological disease-related genes mRNA modules in mood disorder[J]. Frontiers in Genetics, 2022, 13: 865015.
[24]	SHENG Y, TRIYANA S, WANG R, et al. MUC1 and MUC13 differentially regulate epithelial inflammation in response to inflammatory and infectious stimuli[J]. Mucosal immunology, 2013, 6(3): 557-568.
[25]	BEUTLER B, POLTORAK A. Sepsis and evolution of the innate immune response[J]. Critical Care Medicine, 2001, 29: S2-S6.
[26]	BEUTLER B. Inferences, questions and possibilities in Toll-like receptor signalling[J]. Nature, 2004, 430(6996): 257-263.
[27]	ABREU M T, VORA P, FAURE E, et al. Decreased expression of toll-like receptor-4 and MD-2 correlates with intestinal epithelial cell protection against dysregulated proinflammatory gene expression in response to bacterial lipopolysaccharide[J]. The Journal of Immunology, 2001, 167(3): 1609-1616.
[28]	MOYNAGH P N. TLR signalling and activation of IRFs: revisiting old friends from the NF-κB pathway[J]. Trends in Immunology, 2005, 26(9): 469-476.
[29]	HIMMEL M E, HARDENBERG G, PICCIRILLO C A, et al. The role of T-regulatory cells and Toll-like receptors in the pathogenesis of human inflammatory bowel disease[J]. Immunology, 2008, 125(2): 145-153.
[30]	SZEBENI B, VERES G, DEZSÕFI A, et al. Increased expression of Toll-like receptor (TLR) 2 and TLR4 in the colonic mucosa of children with inflammatory bowel disease[J]. Clinical and Experimental Immunology, 2008, 151(1): 34-41.

基因名称 Gene name	上游引物 Forward primer (5'→3')	下游引物 Reverse primer (5'→3')	基因编号 Gene number	熔解温度 Melting temperature/℃	产物大小 Product size/bp
MUC13	GTGATGCAGATGCCAGAGGT	AAAGAAAGCTTGTGCGGCAG	NM_001105293.1	60	135
SRF	GCACTGATTCAGACCTGCCT	CGCAGGTTTCGGTGTATCCT	XM_001929265.3	60	162
IL-6	CCTCCAGGAACCCAGCTATG	GTGGCATCACCTTTGGCATC	NM_214399.1	60	139
MAPK	ACAGGGTTCCTGACGGAGTA	ATGTGGTTCAGCTGGTCGAG	NM_001198922.1	60	179
STAG2	ACAACATTCATGTGACGGGC	AGACAATTCCGAAGTCGGGC	XM_003360436.1	60	133
NF-κB	ATGGAGTACCCTGAGGCTATAA	GTCCGCAATGGAGGAGAAG	EU399817.1	60	138
TLR7	CCAGCTGAAGACTGTCCCTG	GAGCTGAGGTCCAGATGTCG	NM_001097434.1	60	141
GAPDH	CGATGGTGAAGGTCGGAGTG	TGCCGTGGGTGGAATCATAC	NM_001206359.1	60	159

基因名称 Gene name	上游引物 Forward primer (5'→3')	下游引物 Reverse primer (5'→3')	基因编号 Gene number	熔解温度 Melting temperature/℃	产物大小 Product size/bp
MUC13	GTGATGCAGATGCCAGAGGT	AAAGAAAGCTTGTGCGGCAG	NM_001105293.1	60	135
SRF	GCACTGATTCAGACCTGCCT	CGCAGGTTTCGGTGTATCCT	XM_001929265.3	60	162
IL-6	CCTCCAGGAACCCAGCTATG	GTGGCATCACCTTTGGCATC	NM_214399.1	60	139
MAPK	ACAGGGTTCCTGACGGAGTA	ATGTGGTTCAGCTGGTCGAG	NM_001198922.1	60	179
STAG2	ACAACATTCATGTGACGGGC	AGACAATTCCGAAGTCGGGC	XM_003360436.1	60	133
NF-κB	ATGGAGTACCCTGAGGCTATAA	GTCCGCAATGGAGGAGAAG	EU399817.1	60	138
TLR7	CCAGCTGAAGACTGTCCCTG	GAGCTGAGGTCCAGATGTCG	NM_001097434.1	60	141
GAPDH	CGATGGTGAAGGTCGGAGTG	TGCCGTGGGTGGAATCATAC	NM_001206359.1	60	159

基因名称 Gene name	基因编号 Gene number	基因名称 Gene name	基因编号 Gene number
Stag2	NM_001077712	Gnaq	NM_008139
Fam108b	NM_146096	Dag1	NM_010017
Nfe2l1	NM_001130451	Mtap2	NM_008632
Ier5	NM_010500	Nrxn3	NM_172544
Rnf38	NM_001038993	Tardbp	NM_001003899
Zzz3	NM_198416	Prkacb	NM_011100
Tspan7	NM_019634	Neurod1	NM_010894
Ankrd40	NM_027799	Zfp608	NM_175751
Adipor2	NM_197985	Nrxn1	NM_177284
Srf	NM_020493	Dnajb12	NM_019965
Fubp1	NM_057172	Cd24a	NM_009846
Arl8b	NM_026011	Neurod2	NM_010895
Gabrb2	NM_008070	Klf12	NM_010636
Tnrc6b	NM_177124	Calm1	NM_009790
Ivns1abp	NM_001039512	Vsnl1	NM_012038
Gabra2	NM_008066	Nufip2	NM_001024205
Rtn1	NM_001007596	Gal3st3	NM_001024717
Zfhx2	NM_001039198	Zmynd11	NM_144516

基因名称 Gene name	基因编号 Gene number	基因名称 Gene name	基因编号 Gene number
Stag2	NM_001077712	Gnaq	NM_008139
Fam108b	NM_146096	Dag1	NM_010017
Nfe2l1	NM_001130451	Mtap2	NM_008632
Ier5	NM_010500	Nrxn3	NM_172544
Rnf38	NM_001038993	Tardbp	NM_001003899
Zzz3	NM_198416	Prkacb	NM_011100
Tspan7	NM_019634	Neurod1	NM_010894
Ankrd40	NM_027799	Zfp608	NM_175751
Adipor2	NM_197985	Nrxn1	NM_177284
Srf	NM_020493	Dnajb12	NM_019965
Fubp1	NM_057172	Cd24a	NM_009846
Arl8b	NM_026011	Neurod2	NM_010895
Gabrb2	NM_008070	Klf12	NM_010636
Tnrc6b	NM_177124	Calm1	NM_009790
Ivns1abp	NM_001039512	Vsnl1	NM_012038
Gabra2	NM_008066	Nufip2	NM_001024205
Rtn1	NM_001007596	Gal3st3	NM_001024717
Zfhx2	NM_001039198	Zmynd11	NM_144516

分类 Category	所属通路 Pathway affiliated	基因数量 Gene number	所含基因名称 Name of gene contained	P值 P value	发现错误率 False discovery rate
分子复合物1 Molecular Complex 1	KEGG通路:MAPK信号通路 KEGG_PATHWAY: MAPK signaling pathway	11	AKT1, TRP53, FOS, MAPK1, ELK4, MAPK3, ELK1, MAPK8, PAK1, SRF, MYC	2.51×10^-8	2.55×10^-5
	BIOCART:MAL在Rho介导的SRF 激活中的作用 BIOCARTA: Role of MAL in Rho-mediated activation of SRF	6	MAPK1, MAPK3, RHOA, MAPK8, MAL, SRF	3.50×10^-6	0.004 047
	BIOCARTA:PDGF信号通路 BIOCARTA: PDGF signaling pathway	6	FOS, MAPK3, ELK1, MAPK8, SRF, PIK3R1	2.74×10^-5	0.031 626
	BIOCARTA:EGF信号通路 BIOCARTA:EGF signaling pathway	6	FOS, MAPK3, ELK1, MAPK8, SRF, PIK3R1	2.74×10^-5	0.031 626
	KEGG通路:细胞周期 KEGG_PATHWAY: Cell cycle	5	TRP53, CDKN1A, SMAD3, MYC, STAG2	0.001 506	1.518 954
	BIOCARTA:IL-6信号通路 BIOCARTA: IL-6 signaling pathway	4	FOS, MAPK3, ELK1, SRF	0.002 880	3.275 084
	KEGG通路:Toll样受体信号通路 KEGG_PATHWAY: Toll-like receptor signaling pathway	7	AKT1, FOS, MAPK1, MAPK3, MAPK8, PIK3R1, STAG2	3.63×10^-5	0.036 899
分子复合物2 Molecular Complex 2	KEGG通路:MAPK信号通路 KEGG_PATHWAY: MAPK signaling pathway	3	BDNF, NTRK2,PFN2	0.015 087	11.720 810
分子复合物3 Molecular Complex 3	KEGG通路:ABC转运器 KEGG_PATHWAY: ABC transporters	3	ABCB1B, ABCC2, ABCG2	6.01×10^-5	0.006 015
分子复合物4 Molecular Complex 4	KEGG通路:p53信号通路 KEGG_PATHWAY: p53 signaling pathway	3	BBC3, DDB2, GADD45A	0.001 039	0.735 622

Genes and pathways related to piglet diarrhea in miRNAs-MUC13 regulation system

RichHTML

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

Figures/Tables 7

References 30

Related Articles 5

Recommended Articles

Metrics

Comments

[1]	DING Zhaoxue, WANG Jiajie, SHEN Zhonghao, ZHOU Xiaolong, YANG Songbai, JIN Hangfeng, ZHAO Ayong, WANG Han. Construction of PK15 cells with porcine miR-22 upstream sequence mutation [J]. Acta Agriculturae Zhejiangensis, 2022, 34(9): 1849-1855.
[2]	SHAO Jiahao, WANG Jie, JIA Xianbo, CHEN Shiyi, LAI Songjia. Research progress of microRNA in rabbits [J]. , 2020, 32(1): 176-182.
[3]	MU Tong, WANG Guo-mei, YANG Qi-rui, JIN Li, ZHANG Hai-long, ZHANG Li, TIAN Xiao-jing, LIU Li-xia. Polymorphism of SLA-DQB exon 2 and its association with diarrhea in Yantai black pigs [J]. , 2016, 28(10): 1671-1677.
[4]	CHEN Li1, SHEN Jun\\|da1, LI Guo\\|qin1, FAN Yi\\|fei2, MA Jian\\|sheng2, LU Li\\|zhi1，* . Expression and regulation of Tyr, Tyrp1 and C\\|kit gene in feather bulbs of spot\\|billed ducks [J]. , 2015, 27(5): 729-.
[5]	LU Fu\\|zeng;CHU Xiao\\|hong;DAI Li\\|he;CHEN Shao\\|meng;FU Chun\\|quan;HU Jin\\|ping;XYU Ru\\|hai;*. Construction of piglet diarrhea regulation network involved with MUC13 gene [J]. , 2013, 25(6): 0-1224.