布鲁氏菌Ⅳ型分泌系统效应蛋白VceC对山羊滋养层细胞内质网应激和性腺激素分泌的影响

doi:10.11843/j.issn.0366-6964.2023.03.032

Abstract

Abstract: We aimed to study the effects of Brucella Type IV secretes system effector protein VceC on endoplasmic reticulum stress and gonadal hormone secretory in goat trophoblast cells, and to reveal its role in Brucella infection of host cells, which is of great significance for clarifying the mechanism of intracellular survival of Brucella and miscarriage in animals. In this experiment the eukaryotic expression vector pEGFP-C1-VceC of VceC was constructed and transfected it into GTC, then the main factors of ERS, GRP78 and CHOP, were detected by Western blot method. Then the related molecules of unfolded protein reaction (UPR) were further detected by qRT-PCR and Western blot methods. The concentration of Progesterone and Estrogen secreted by GTC were detected by ELISA. The results showed that the VceC eukaryotic expression vector pEGFP-C1-VceC was constructed and transfected it into GTC successfully. The protein expression of main factors of ERS, GRP78 were significantly increased (P<0.01) at 12 and 24 h after transfection of pEGFP-C1-VceC into GTC, while CHOP protein expression was significantly decreased (P<0.05) at 24 h after transfection. According to the qRT-PCR results, the mRNA expression levels of IRE1 and XBP-1 were significantly increased (P<0.05) at 24 h after transfection of pEGFP-C1-VceC, while PERK, ATF6 and ATF4 mRNA had little change (P>0.05). According to the Western blot results, the protein expression of IRE1 was significantly increased at 12 h after transfection of pEGFP-C1-VceC (P<0.01). In addition, the content of progesterone in cell culture supernatant of the VceC transfection group decreased significantly (P<0.01) at 12 and 24 h than those in the GTC group and pEGFP-C1 transfected GTC group, while the content of Estrogen had little change (P>0.05). The results suggest that VceC can activate the ERS of GTC by activating the IRE1 pathway of unfolded protein response. Moreover, VceC protein also changes the Progesterone/Estrogen ratio in GTC.

Key words: Brucella, type IV secretion system, VceC effector protein, goat trophoblastic cells, endoplasmic reticulum stress, gonadal hormone

CLC Number:

S855.12

XIANG Caixia, WANG Xiangguo, LI Junmei, ZHI Feijie, FANG Jiaoyang, ZHENG Weifang, CHEN Jialu, JIN Yaping, WANG Aihua. The Influence of Brucella Type IV Secretes System Effector Protein VceC on Endoplasmic Reticulum Stress and Gonadal Hormone Secretory in Goat Trophoblast Cells[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(3): 1210-1220.

References 38

[1]	陆承平.兽医微生物学[M]. 5版.北京:中国农业出版社, 2013.LU C P. Veterinary Microbiology[M]. 5th Edition. Beijing:China Agriculture Press, 2013.(in Chinese)
[2]	ATLURI V L, XAVIER M N, DE JONG M F, et al. Interactions of the human pathogenic Brucella species with their hosts[J]. Annu Rev Microbiol, 2011, 65:523-541.
[3]	CASWELL C C. Assessment of survival and replication of Brucella spp. in murine peritoneal macrophages[M]//ALLEN I C. Mouse Models of Innate Immunity. New York:Humana Press, 2019, 1960:181-189.
[4]	FRANCO M P, MULDER M, SMITS H L. Persistence and relapse in brucellosis and need for improved treatment[J]. Trans R Soc Trop Med Hyg, 2007, 101(9):854-855.
[5]	BYNDLOSS M X, TSAI A Y, WALKER G T, et al. Brucella abortus infection of placental trophoblasts triggers endoplasmic reticulum stress-mediated cell death and fetal loss via type IV secretion system-dependent activation of CHOP[J]. mBio, 2019, 10(4):e01538-19.
[6]	ZHOU Y C, BU Z Y, QIAN J, et al. The UTP-glucose-1-phosphate uridylyltransferase of Brucella melitensis inhibits the activation of NF-κB via regulating the bacterial type IV secretion system[J]. Int J Biol Macromol, 2020, 164:3098-3104.
[7]	DE JONG M F, SUN Y H, DEN HARTIGH A B, et al. Identification of VceA and VceC, two members of the VjbR regulon that are translocated into macrophages by the Brucella type IV secretion system[J]. Mol Microbiol, 2008, 70(6):1378-1396.
[8]	DE JONG M F, STARR T, WINTER M G, et al. Sensing of bacterial type IV secretion via the unfolded protein response[J]. mBio, 2013, 4(1):e00418-12.
[9]	ZIMMERMANN R. Components and mechanisms of import, modification, folding, and assembly of immunoglobulins in the endoplasmic reticulum[J]. J Clin Immunol, 2016, 36 Suppl 1:5-11.
[10]	WANG M, KAUFMAN R J. Protein misfolding in the endoplasmic reticulum as a conduit to human disease[J]. Nature, 2016, 529(7586):326-335.
[11]	PLUQUET O, POURTIER A, ABBADIE C. The unfolded protein response and cellular senescence. A review in the theme:cellular mechanisms of endoplasmic reticulum stress signaling in health and disease[J]. Am J Physiol Cell Physiol, 2015, 308(6):C415-C425.
[12]	GORMAN A M, HEALY S J M, JÄGER R, et al. Stress management at the ER:regulators of ER stress-induced apoptosis[J]. Pharmacol Ther, 2012, 134(3):306-316.
[13]	CLÁUDIO N, DALET A, GATTI E, et al. Mapping the crossroads of immune activation and cellular stress response pathways[J]. EMBO J, 2013, 32(9):1214-1224.
[14]	TODD D J, LEE A H, GLIMCHER L H. The endoplasmic reticulum stress response in immunity and autoimmunity[J]. Nat Rev Immunol, 2008, 8(9):663-674.
[15]	MCCONKEY C A, DELORME-AXFORD E, NICKERSON C A, et al. A three-dimensional culture system recapitulates placental syncytiotrophoblast development and microbial resistance[J]. Sci Adv, 2016, 2(3):e1501462.
[16]	SFERRUZZI-PERRI A N, CAMM E J. The programming power of the placenta[J]. Front Physiol, 2016, 7:33.
[17]	MARTÍNEZ R, DUNNER S, TORO R, et al. Effect of polymorphisms in the Slc11a1 coding region on resistance to brucellosis by macrophages in vitro and after challenge in two Bos breeds (Blanco Orejinegro and Zebu)[J]. Genet Mol Biol, 2010, 33(3):463-470.
[18]	BOSCHIROLI M L, OUAHRANI-BETTACHE S, FOULONGNE V, et al. The Brucella suis virB operon is induced intracellularly in macrophages[J]. Proc Natl Acad Sci U S A, 2002, 99(3):1544-1549.
[19]	CELLI J, DE CHASTELLIER C, FRANCHINI D M, et al. Brucella evades macrophage killing via VirB-dependent sustained interactions with the endoplasmic reticulum[J]. J Exp Med, 2003, 198(4):545-556.
[20]	SALCEDO S P, MARCHESINI M I, DEGOS C, et al. BtpB, a novel Brucella TIR-containing effector protein with immune modulatory functions[J]. Front Cell Infect Microbiol, 2013, 3:28.
[21]	SENGUPTA D, KOBLANSKY A, GAINES J, et al. Subversion of innate immune responses by Brucella through the targeted degradation of the TLR signaling adapter, MAL[J]. J Immunol, 2010, 184(2):956-964.
[22]	MYENI S, CHILD R, NG T W, et al. Brucella modulates secretory trafficking via multiple type IV secretion effector proteins[J]. PLoS Pathog, 2013, 9(8):e1003556.
[23]	DÖHMER P H, VALGUARNERA E, CZIBENER C, et al. Identification of a type IV secretion substrate of Brucella abortus that participates in the early stages of intracellular survival[J]. Cell Microbiol, 2014, 16(3):396-410.
[24]	DUDEK J, BENEDIX J, CAPPEL S, et al. Functions and pathologies of BiP and its interaction partners[J]. Cell Mol Life Sci, 2009, 66(9):1556-1569.
[25]	MUNRO S, PELHAM H R B. An Hsp70-like protein in the ER:identity with the 78 kd glucose-regulated protein and immunoglobulin heavy chain binding protein[J]. Cell, 1986, 46(2):291-300.
[26]	GIORGI C, BONORA M, SORRENTINO G, et al. p53 at the endoplasmic reticulum regulates apoptosis in a Ca2+-dependent manner[J]. Proc Natl Acad Sci U S A, 2015, 112(6):1779-1784.
[27]	OYADOMARI S, MORI M. Roles of CHOP/GADD153 in endoplasmic reticulum stress[J]. Cell Death Differ, 2004, 11(4):381-389.
[28]	QIN Q M, PEI J W, ANCONA V, et al. RNAi screen of endoplasmic reticulum-associated host factors reveals a role for IRE1α in supporting Brucella replication[J]. PLoS Pathog, 2008, 4(7):e1000110.
[29]	SMITH J A, KHAN M, MAGNANI D D, et al. Brucella induces an unfolded protein response via TcpB that supports intracellular replication in macrophages[J]. PLoS Pathog, 2013, 9(12):e1003785.
[30]	TAGUCHI Y, IMAOKA K, KATAOKA M, et al. Yip1A, a novel host factor for the activation of the IRE1 pathway of the unfolded protein response during Brucella infection[J]. PLoS Pathog, 2015, 11(3):e1004747.
[31]	HE Y Q, REICHOW S, RAMAMOORTHY S, et al. Brucella melitensis triggers time-dependent modulation of apoptosis and down-regulation of mitochondrion-associated gene expression in mouse macrophages[J]. Infect Immun, 2006, 74(9):5035-5046.
[32]	TOLOMEO M, DI CARLO P, ABBADESSA V, et al. Monocyte and lymphocyte apoptosis resistance in acute and chronic brucellosis and its possible implications in clinical management[J]. Clin Infect Dis, 2003, 36(12):1533-1538.
[33]	CHEN F, HE Y Q. Caspase-2 mediated apoptotic and necrotic murine macrophage cell death induced by rough Brucella abortus[J]. PLoS One, 2009, 4(8):e6830.
[34]	JI L, BRKIĆ J, LIU M, et al. Placental trophoblast cell differentiation:physiological regulation and pathological relevance to preeclampsia[J]. Mol Aspects Med, 2013, 34(5):981-1023.
[35]	HALASZ M, SZEKERES-BARTHO J. The role of progesterone in implantation and trophoblast invasion[J]. J Reprod Immunol, 2013, 97(1):43-50.
[36]	WANG X G, LIN P F, LI Y, et al. Brucella suis vaccine strain 2 induces endoplasmic reticulum stress that affects intracellular replication in goat trophoblast cells in vitro[J]. Front Cell Infect Microbiol, 2016, 6:19.
[37]	王相国.内质网应激在布鲁氏菌体外感染山羊子宫内膜与滋养层细胞中的作用[D].杨凌:西北农林科技大学, 2016.WANG X G. The Role of endoplasmic reticulum stress in Brucella-infected goat endometrial and trophoblast cells in vitro[D].Yangling:Northwest A&F University, 2016.(in Chinese)
[38]	MARTIROSYAN A, PÉREZ-GUTIERREZ C, BANCHEREAU R, et al. Brucella β 1, 2 cyclic Glucan is an activator of human and mouse dendritic cells[J]. PLoS Pathog, 2012, 8(11):e1002983.

The Influence of Brucella Type IV Secretes System Effector Protein VceC on Endoplasmic Reticulum Stress and Gonadal Hormone Secretory in Goat Trophoblast Cells

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References 38

Related Articles 15

Recommended Articles

Metrics

Comments

[1]	XU Zhenyu, DENG Xiaoyu, WANG Yueli, SUN Can, WU Aodi, CAO Jian, YI Jihai, WANG Yong, WANG Zhen, CHEN Chuangfu. Biological Characteristics of Brucella abortus A19ΔBtpA Deletion Strain and Its Immunogenicity Study [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(5): 2135-2145.
[2]	WU Shangjie, LUAN Yuanyuan, WANG Mingkun, ZHANG Hechun, YU Bo, MA Yuehui, JIANG Lin, HE Xiaohong. Advances of Disease-Resistant Breeding on Ovine Brucellosis [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(3): 882-893.
[3]	GAO Kangkang, YI Yanyan, ZHAO Yiteng, LIN Pengfei, CHEN Huatao, JIN Yaping. Protective Effect of Endoplasmic Reticulum Stress Preadaptation on LPS-Induced Inflammatory Response in Goat Endometrial Epithelial Cells [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(8): 3546-3556.
[4]	WANG Chongnian, YU Jialin, GONG Zhaoqian, WU Xiaoling, DENG Guangcun. Regulation of BCG-induced Autophagy in Macrophages RAW264.7 by PLIN2 [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(5): 2134-2146.
[5]	ZHAI Yunyi, YUAN Ye, LI Junmei, TIAN Lulu, DIAO Ziyang, LI Bin, CHEN Jialu, ZHOU Dong, JIN Yaping, WANG Aihua. Preparation and Preliminary Application of Monoclonal Antibody to Brucella Outer Membrane Protein 16 [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(5): 2083-2091.
[6]	YANG Mengran, YANG Chenglin, WU You, WANG Siqi, KONG Xiangyi, NING Can, XIAO Wenguang, FAN Hui, WU Jing, YUAN Zhihang. Study on the Mechanism of Citrinin Induced Intestinal Barrier Dysfunction in Mice [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(11): 4805-4816.
[7]	LI Yang, ZHOU Dong, YIN Yanlong, ZHANG Guangdong, XIANG Caixia, ZHI Feijie, BAI Furong, LIN Pengfei, JIN Yaping, WANG Aihua. Effects of Brucella Outer Membrane Protein 16 on Apoptosis and Immune Activity of RAW264.7 Cells [J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(8): 2642-2651.
[8]	ZHANG Kaizhao, HU Hui, XU Zekai, WANG Shiqian, CUI Hongjie, HUANG Xiaohong. Toxicity of Zearalenone on Chicken Embryo Fibroblasts [J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(5): 1615-1625.
[9]	YANG Qin, DENG Xiaoyu, XIE Shanshan, YI Jihai, WANG Yong, ZHANG Qian, WANG Zhen, CHEN Chuangfu. Effects of Brucella bovis Type IV Secretion System on Endoplasmic Reticulum Stress and Apoptosis of Macrophages [J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(4): 1192-1200.
[10]	XI Jing, WANG Yueli, DENG Xiaoyu, YANG Qin, LI Peidong, ZHANG Jiangwei, SUN Tianhao, ZHU Liangquan, YI Jihai, CHEN Chuangfu. Effect of STAT6 Mediated Macrophage Polarization on Intracellular Survival of Brucella [J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(1): 263-271.
[11]	MEI Li, WANG Yingchao, CHENG Rujia, YU Guoji, FAN Xuezheng, GAO Xiaolong, GAO Min, QIN Yuming, LI Xiaoying, LI Qiaoling, ZHU Liangquan, FENG Xiaoyu. A Droplet Digital PCR Method for Detection of Brucella [J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(6): 1753-1759.
[12]	WANG Zongjie, ZHANG Ruixue, LIU Shouqin, JIN Yaping, LIN Pengfei. The Apoptotic Effect of Zearalenone on Goat Endometrial Stromal Cells [J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(2): 535-542.
[13]	JIANG Hui, FENG Yu, LI Xiaoying, FAN Xuezheng, PENG Xiaowei, DING Jiabo. Comparative Research of Four High Throughput Antibody Detection Methods for Brucella [J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(11): 3208-3214.
[14]	NIE Xueyi, LIU Lei, ZHENG Xuedi, YANG Yi, XU Jinrui, WANG Yujiong. Role of Endoplasmic Reticulum Stress on Pyroptosis of BCG Infected Macrophages [J]. Acta Veterinaria et Zootechnica Sinica, 2021, 52(10): 2842-2851.
[15]	WANG Shuli, ZHANG Huiru, BI Yanqi, WANG Dejuan, CHEN Lin, ZHANG Xiaoting, LI Zhiqiang. Analysis of Immune Responses Induced by Brucella Transcriptional Regulatory Factor HFQ [J]. Acta Veterinaria et Zootechnica Sinica, 2020, 51(8): 1977-1984.