猪免疫抑制因子MNSFβ的克隆、表达与组织分布

doi:10.3724/SP.J.1005.2013.01377

[an error occurred while processing this directive]

HEREDITAS ›› 2013, Vol. 35 ›› Issue (12): 1377-1383.doi: 10.3724/SP.J.1005.2013.01377

Cloning, eukaryotic expression and spatial expression patterns of porcine MNSFβ

WANG Jing-Na^1,2, JIANG Peng-Fei^1,2, KANG Zhan-Zhan^1,2, LI Deng-Yun^1,2, HUA Lin-Lin^1,2, ZHANG Yang^1,2, ZHANG Shu-Xia^1,2, ZHANG De-Li^1,2

1. Key Laboratory of Animal Biotechnology of National Ministry of Agriculture, Institute of Veterinary Immunology of Northwest A & F University, Yangling 712100, China;
2. Research Laboratory of Virology, Immunology & Bioinformatics, Division of Veterinary Microbiology & Virology, Department of Pre-ventive Veterinary Medicine, College of Veterinary Medicine, Northwest A & F University, Yangling 712100, China

Received:2013-05-11 Revised:2013-07-29 Online:2013-12-20 Published:2013-11-25

Abstract

Abstract:

MNSFβ(Monoclonal nonspecific suppressor factor β) is a natural immunosuppressive factor which has been reported to be involved in various biological processes, such as immune responses, cell division, stress response, cell apoptosis, and nuclear transport. However, study on porcine MNSFβ has been rarely reported. In this study, the full-length sequence of porcine MNSFβ (GenBank accession number: KF77642500) was predicted in silicon and its cDNA sequence was obtained through RT-PCR from porcine spleen. The nucleic acid and protein sequences were analyzed. Then, the gene was subcloned into pEGFP-C1 to construct a recombinant plasmid pEGFP-MNSFβ which was transfected into swine umbilical vein endothelial cells (SUVECs) using Lipofectamine 2000. The expression of GFP was detected by fluorescence microscopy, Western blot, and laser confocal fluorescence microscopy. The spatial expression patterns of porcine MNSFβ were detected by real-time qPCR. Results showed that the full length of porcine MNSFβ was 402 bp encoding 133 amino acids with only one exon. Bioinformatics analysis showed that porcine MNSFβ protein was a stable protein consisting of a ubiquitin-like domain fused to the ribosomal protein S30 with no signal peptide. The analyses of homology and phylogenetic tree of porcine MNSFβ and its homologs in other 18 species showed that the identities of MNSFβ protein sequence were higher than 91% among different species and the evolutionary distance was less than 0.05. It indicates that MNSFβ is highly con-served in the process of evolution. Fluorescence signal showed that the fusion protein GFP-MNSFβ was successfully expressed in SUVECs which was then confirmed by Western blot. Laser confocal fluorescence microscopy showed that MNSFβ was expressed in both nucleus and cytoplasm. Analysis of spatial expression patterns showed that procine MNSFβ was widely expressed in immune tissues, but not in lung, suggesting that MNSFβ may play an important role in immune response.

Key words: porcine, monoclonal nonspecific suppressor factor &beta, (MNSFβ), eukaryotic expression, spatial expres-sion patterns

WANG Jing-Na, JIANG Peng-Fei, KANG Zhan-Zhan, LI Deng-Yun, HUA Lin-Lin, ZHANG Yang, ZHANG Shu-Xia, ZHANG De-Li. Cloning, eukaryotic expression and spatial expression patterns of porcine MNSFβ[J]. HEREDITAS, 2013, 35(12): 1377-1383.

References

[1] Nakamura M, Ogawa H, Tsunematsu T. Mode of action of monoclonal-nonspecific suppressor factor (MNSF) pro-duced by murine hybridoma. Cell Immunol, 1988, 116(1): 230–239. <\p>

[2] Nakamura M, Xavier RM, Tsunematsu T, Tanigawa Y. Molecular cloning and characterization of a cDNA encod-ing monoclonal nonspecific suppressor factor. Proc Natl Acad Sci USA, 1995, 92(8): 3463–3467. <\p>

[3] Nakamura M, Tanigawa Y. Ubiquitin-like polypeptide in-hibits cAMP-induced p38 MAPK activation in Th2 cells. Immunobiology, 2004, 208(5): 439–444. <\p>

[4] Naramura M, Watanabe N. Ubiquitin-like protein MNSFβ/ endophilinⅡcomplex regulates Decin-1-mediated phagocytosis and in flammatory responses in macrophages. Biochem Biophys Res Commun, 2010, 401(2): 257–261. <\p>

[5] Suzuki K, Nakamura M, Nariai Y, Dekio S, Tanigawa Y. Monoclonal nonspecific suppressor factor β (MNSFβ) in-hibits the production of TNF-α by lipopolysaccharide-activated macrophages. Immunobiology, 1996, 195(2): 187–198. <\p>

[6] Nakamura M, Omura S. Quercetin regulates the inhibitory effect of monoclonal non-specific suppressor factor β on tumor necrosis factor-α production in LPS-stimulated macrophages. Biosci Biotechnol Biochem, 2008, 72(7): 1915–1920. <\p>

[7] Nakamura M, Watanabe J, Watanabe N. Ubiquitin-like protein MNSFβ regulates TLR-2-mediated signal trans-duction. Mol Cell Biochem, 2012, 364(1-2): 39–43. <\p>

[8] Nie GY, Li Y, Hampton AL, Salamonsen LA, Clements JA, Findlay JK. Identification of monoclonal nonspecific sup-pressor factor beta (MNSFβ) as one of the genes differen-tially expressed at implantation sites compared to interim-plantation sites in the mouse uterus. Mol Reprod Dev, 2000, 55(4): 351–363. <\p>

[9] Nakamura M, Xavier RM, Tanigawa Y. Monoclonal non-specific suppressor factor β inhibits interleukin-4 secretion by a type-2 helper T cell clone. Eur J Immunol, 1995, 25(8): 2417–2419. <\p>

[10] Nakamura M, Xavier RM, Tanigawa Y. Monoclonal non- specific suppressor factor (MNSF) inhibits the IL-4 secre-tion by bone marrow-derived mast cell (BMMC). FEBS Lett, 1994, 339(3): 239–242. <\p>

[11] 何球藻, 吴厚生, 曹雪涛. 细胞与分子免疫学. 上海: 上海科学技术文献出版社, 1997. 146–148. <\p>

[12] Nakamura M, Tanigawa Y. Noncovalent interaction of MNSFbeta, a ubiquitin-like protein, with histone 2A. Comp Biochem Physiol Part B: Biochem Mol Biol, 2005, 140(2): 207–210. <\p>

[13] Watanabe J, Nakagawa M, Watanabe N, Nakamura M. Ubiquitin-like protein MNSFβ covalently binds to Bcl-G and enhances lipopolysaccharide/interferon γ-induced apoptosis in macrophages. FEBS J, 2013, 280(5): 1281–1293. <\p>

[14] Naramura M, Yamaguchi S. The ubiquitin-like protein MNSF β regulates ERK-MAPK cascade. J Biol Chem, 2006, 281(25): 16861–16869. <\p>

[15] Cagnol S, Chambard J C. ERK and cell death: Mecha-nisms of ERK-induced cell death-apoptosis, autophagy and senescence. FEBS J, 2010, 277(1): 2–21. <\p>

[16] Jiang P, Li DY, Bi LJ, Zhang DL. BCL-G as a new candi-date gene for immune responses in pigs: Bioinformatic analysis and functional characterization. Vet Immunol Immunopathol, 2012, 150(1-2): 112–117. <\p>

[17] Nakamura M, Shimosaki S. The ubiquitin-like protein monoclonal nonspecific suppressor factor β conjugates to endophilin II and regulates phagocytosis. FEBS J, 2009, 276(21): 6355–6363. <\p>

[18] Nakamura M, Tanigawa Y. Characterization of ubiquitin- like polypeptide acceptor protein, a novel pro-apoptotic member of the Bcl2 family. Eur J Biochem, 2003, 270(20): 4052–4058.<\p>

Metrics

Comments

www.chinagene.cn
备案号：京ICP备09063187号

Cloning, eukaryotic expression and spatial expression patterns of porcine MNSFβ

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References

Related Articles 15

Recommended Articles

Metrics

Comments

[1]	Xiangwei Tang, Dan Chu, Saina Yan, Yanfei Yin, Qiao Bian, Bo Weng, Bin Chen, Maoliang Ran. MiR-191 promotes the porcine immature Sertoli cell proliferation by targeting the BDNF gene through activating the PI3K/AKT signaling pathway [J]. Hereditas(Beijing), 2021, 43(7): 680-693.
[2]	Ziying Huang, Long Li, Qianqian Li, Xiangdong Liu, Changchun Li. The effect of lncRNA TCONS_00815878 on differentiation of porcine skeletal muscle satellite cells [J]. Hereditas(Beijing), 2019, 41(12): 1119-1128.
[3]	Maoliang Ran, Lianhua Dong, Bo Weng, Rong Cao, Fuzhi Peng, Hu Gao, Hui Luo, Bin Chen. miR-362 regulates the proliferation and apoptosis of porcine immature Sertoli cells through targeting the ZNF644 gene [J]. Hereditas(Beijing), 2018, 40(7): 572-584.
[4]	Cuili Zhong, Guoling Li, Jianxin Mo, Rong Quan, Haoqiang Wang, Zicong Li, Zhenfang Wu, Xianwei Zhang. Effects of parameters, plasmid dosages and topological structures on transfection efficiency of porcine fetal fibroblasts using different electroporators [J]. Hereditas(Beijing), 2017, 39(10): 930-938.
[5]	Guoling Li, Cuili Zhong, Sheng Ni, Dewu Liu, Gengyuan Cai, Zicong Li, Huaqiang Yang, Zhenfang Wu. Establishment of porcine Xist knockout model using CRISPR/Cas9 system [J]. Hereditas(Beijing), 2016, 38(12): 1081-1089.
[6]	Lizu Li, Lixin Han, Jinkui Wang, Liang Wang, Di Liu, Xiuqin Yang. Cloning and identification of splice variants of the porcine PILRA gene [J]. HEREDITAS(Beijing), 2015, 37(9): 926-931.
[7]	Maoliang Ran, Bin Chen, Jie Yin, Anqi Yang, Zhi Li, Ming Jiang. Advances in porcine miRNAome [J]. HEREDITAS(Beijing), 2014, 36(10): 974-984.
[8]	YANG Xiu-Qin, CHEN Yue-Chan, WANG Li-ang, LI Hai-Tao, LIU Di, GUAN Qing-Zhi, FU Bo. Functional analysis of SNPs in porcine TLR4 gene [J]. HEREDITAS, 2012, 34(8): 1050-1056.
[9]	ZHANG Feng, YANG Zhen-Zhen, DOU Gong-Wei, LI Wei-Hang, LV Bei, Bolund Lars, DU Yu-Chao, TAN Ping-Ping, MA Run-Lin. Production of porcine blastocysts expressed EGFP by handmade cloning [J]. HEREDITAS, 2011, 33(5): 527-532.
[10]	. Expression analysis of porcine ATF4 gene and its association with production traits [J]. HEREDITAS, 2011, 33(12): 1347-1352.
[11]	BAO Wen-Bin, XIE Lan, BO Zhang-Yuan, SHU Jing, DU Zi-Dong, CA Jia-Jia, HUANG Xiao-Guo, SHU Guo-Jiang, TUN Ku-Long. cDNA microarray on differently expressed genes in duodenum in porcine sensitive or resistant to Escherichia coli F18 [J]. HEREDITAS, 2011, 33(1): 60-66.
[12]	HE Jian-Feng, ZHANG Yan-Ming, DUAN Hui-Juan, CAO Jin-Suo, ZHANG De-Li. In silico identification, molecular cloning and verification of a novel pig gene homologous to human BCL10 of in-nate immunity and its preliminary expression profiles in pigs [J]. HEREDITAS, 2008, 30(6): 747-754.
[13]	WU Sheng-Long, BAO Wen-Bin, JU Hui-Ping, CHEN Guo-Hong, LI Bi-Chun, CHENG Jin-Hua, ZHU Guo-Qiang. Analysis of polymorphisms in exon 14 in porcine Mx1 gene and iden-tification of new mutation points [J]. HEREDITAS, 2007, 29(6): 693-693―698.
[14]	XING Xiao-Wei , XUE Li-Qun , HUANG Sheng-Qiang , LI Shu-Juan, WANG Wei . Investigation of the biological characters of porcine endogenous retrovirus (PERV) in Shazi Ling pigs [J]. HEREDITAS, 2006, 28(7): 799-804.
[15]	LUO Gang, WEI Hong. Recombination and Fusion Expression of Porcine Defensin Gene PBD-I in E.coli [J]. HEREDITAS, 2003, 25(2): 146-150.