秀丽隐杆线虫病原逃避行为调控的分子机制研究进展
邹伟1, 王琦1, 林容2, 黄晓玮2
1.昆明医科大学 公共卫生学院,云南 昆明 650500
2.云南大学 省部共建云南生物资源保护与利用国家重点实验室,云南 昆明 650091
通信作者:黄晓玮(1973-),女,山东人,研究员,主要从事病原与宿主相互用用关系研究.E-mail:xwhuang@ynu.edu.cn.

作者简介:邹 伟(1988-),男,云南人,讲师,主林从事病原与宿主相互作用关系研究.E-mial:280018988@qq.com.

摘要

线虫( Caenorhabditis elegans)生活在复杂生态系统的环境中,具备对环境刺激变化的敏锐感知和应对能力.除常规非生物或生物逆境,如不适宜的气味、温度和盐离子可使线虫产生逃避反应外,病原细菌侵染也能诱发其逃避行为,从而有效帮助线虫辨别病原细菌和细菌食物、迅速逃离危险,是线虫提高生存率的重要策略.现有的研究证明,病原菌铜绿假单胞PA14( Pseudomonas aeruginosa,简写为PA14)、沙雷式菌( Serentia marcescens)、苏云金芽孢杆菌( Bacillus thuringiensis)等都会诱发逃避.根据逃避行为的方式和作用机制不同可将其分为两类:一类是应对病原伤害形成的本能逃避行为;第二类是学习记忆产生的学习型逃避行为.该文将针对秀丽隐杆线虫的两类病原逃避行为的分子机制进行综述.

关键词: 秀丽隐杆线虫; 病原细菌; 逃避行为; 分子机制
中图分类号:Q442 文献标志码:A 文章编号:0258-7971(2018)06-1277-06
Molecular mechanisms of the avoidance behavior induced by pathogenic bacteria in Caenorhabditis elegans
ZOU Wei1, WANG Qi1, LIN Rong2, HUANG Xiao-wei2
1.School of Public Health,Kunming Medical University,Kunming 650500,China
2.State Key Laboratory for Conservation and Utilization of Bio-resources,Yunnan University,Kunming 650091,China
Abstract

Since Caenorhabditis elegans lives in a complex ecosystem,it has evolved the power to perceive and response to a variety of environmental stimuli effectively.Besides the usual abiotic and biological stress,such as unsuitable odors,high or low temperatures,hyperhaline stimulation and so on,infection of pathogenic bacteria can also induce the behavior of avoidance in C. elegans.The avoidance triggered by pathogens helps the worms distinguish the harmful bacteria from the food bacteria and escape from the fatal dangers quickly,which hence enhances the possibilities of C. elegansto survive.It has been reported that the bacterial pathogens,at least including Pseudomonasaeruginosa PA14, Serentia marcescens and Bacillus thuringiensis,can induce the avoidance.Furthermore,the aversive behavior is mainly divided into two categories based on the different mechanisms:an instinctive escape response and the avoidance depending on learning.In this paper,the related signaling pathways involved in those two types of avoidance were both reviewed.

Keyword: Caenorhabditis elegans; pathogenic bacteria; avoidance behavior; molecular mechanism

自然界中逃避行为广泛存在, 该行为对人类及动物的生存和环境的适应都具有重要意义.研究发现黑鲫鱼闻到同类受伤皮肤产生的气味后, 会迅速潜入水底并找到安全的掩体躲避, 这种快速的逃避危险的行为是由大脑产生的一元胺类五羟色胺和多巴胺诱发[1].模式生物秀丽隐杆线虫(Caenorhabditis elegans)逃避行为的研究则更为广泛和深入, 不适宜的气味、温度、盐离子和病原细菌都能诱发线虫的逃避行为.此外, 由于以土壤中的细菌为食, 在长期的生存斗争中, 线虫还进化出有效区分病原细菌和有益细菌食物, 敏锐的感知并逃离病原菌的能力[2, 3].依据秀丽隐杆线虫对病原细菌逃避行为形成方式的不同, 病原逃避行为分为本能逃避和学习型的逃避两类.五羟色胺TPH-1、TGF-β 信号通路配体DBL-1、DAF-7和胰岛素信号肽INS-6、INS-7 参与调控线虫学习型病原逃避.TAX-4/TAX-2环鸟甘酸门控通道(TAX-4/TAX-2 cGMP-gated channel)、胰岛素受体DAF-2、神经肽受体NPR-1和泛素化E3连接酶HECW-1则参与本能逃避行为调控.

1 依赖于学习的病原逃避行为

大肠杆菌OP50(以下简称OP50)是线虫C.elegans的食物, 而铜绿假单胞菌PA14(以下简称PA14)则是线虫的致病菌.然而当线虫首次同时嗅到OP50和PA14的气味时, 线虫更喜欢PA14[4, 5], 且被其吸引.研究发现这种偏好行为主要由AWB神经元的神经肽NLP-9、受体NPR-18以及AWC神经元的神经肽NLP-1、受体Gα ODR-3介导[5, 6].此外, 线虫偏好会随着对食物质量的好坏感知而发生改变, 将线虫放入PA14菌斑中训练4h后, 会对PA14的伤害产生记忆, 再次嗅到PA14气味, 发生气味偏好转变, 表现为对PA14的逃避反应.Zhang等报道这种病原逃避行为与线虫对病原菌的伤害学习相关[7], 线虫暴露于病原菌后, ADF神经元中的TPH-1表达量增加, 合成血清素进而加强线虫对病原的学习能力激活逃避行为.反之, 当敲除血清素合成基因(cat-1cat-4 , 尤其是tph-1基因)和受体mod-1后, 降低了线虫的病原学习能力从而抑制逃避行为[7].进一步研究发现, ADF神经元中合成血清素与中间神经元AIZ、AIY受体MOD-1结合, 激活下游信号通路调控线虫逃避行为[5, 7](图1).

图1 秀丽隐杆线虫依赖于血清素产生的学习型病原逃避行为示意图Fig.1 The model of C.elegans for pathogen avlidance based on serotonim

其次, tir-1基因也参与了依赖于血清素TPH-1介导的学习型逃避行为.tir-1同源于哺乳动物的SARM基因, 作用于保守信号途径NSY-1-SEK-1-PMK-1 MAPK 的上游[8].在线虫抵抗病原菌侵染的过程中, TIR-1-NSY-1-SEK-1信号通路在不同组织中具有不同的作用.在肠道中, TIR-1-NSY-1-SEK-1信号通路调控天然免疫; 而神经元中, TIR-1-NSY-1-SEK-1信号通路作用于tph-1基因上游调控病原逃避行为[9, 10](图1).

另一方面, 细胞重要代谢通路监测途径也作用于血清素上游参与病原逃避行为调控.致病菌合成释放大量毒素和致病因子作用于线虫的转录、翻译和线粒体呼吸传递链等信号通路, 消弱其代谢功能和抵抗力.线虫通过监测细胞中核糖体、溶酶体和线粒体的反应变化激活解毒功能、逃避行为和天然免疫系统以抵抗病原菌的侵染.在病原逃避行为调控的过程中, 线虫感应到细胞核心代谢通路受损后, 激活JNK MAPK途径(MLK-1、MEK-1和KGB-1) 传递应激信号到神经系统, 进而整合到ADF神经元, 增加合成血清素促进病原逃避行为 [7, 11](图1).

此外, TGF-β 途径和胰岛素信号肽ISN-6、INS-7则独立于血清素途径参与学习型病原逃避行为调控.TGF-β 途径包含DBL-1和DAF-7两个配体, DAF-7参与滞育起始和新陈代谢调控, DBL-1参与形态发育和天然免疫调控[12, 13].当线虫经过PA14训练后, 由G-CaMP通道介导抑制AVA神经元的活性, 合成释放DBL-1, 结合到ASI神经元和真皮的受体SMAD-6, 对病原气味感知进行可塑性调控, 从而激活学习型逃避行为[14](图2).与TGF-β 途径的配体DBL-1参与学习型逃避行为的调控不同, DAF-7只在趋化神经元ASI中表达作为应对选择细菌食物好坏的策略[15, 16].PA14产生的2种次生代谢产物吩嗪酰胺和绿脓菌素螯铁蛋白激活ASJ神经元G蛋白信号通路, G蛋白的亚基GPA-2和GPA-3激活DAF-7的表达, DAF-7分泌后与中间神经元RIM/RIC上的受体DAF-1结合, 调控趋氧性行为从而促进线虫对PA14的逃避[17](图2).而在胰岛素信号途径中, ASI神经元合成类胰岛素信号肽INS-6通过抑制URX神经元中INS-7的表达, 进而拮抗RIA神经元中的DAF-2表达激活线虫病原逃避行为(图2)[18].

图2 TGF-β 途径、胰岛素信号途径介导的学习型病原逃避行为Fig.2 The model of C.elegans for pathogen adoidance based on TGF-β and Insulin signaling pathuany

2 本能的病原逃避行为

TAX-4/TAX-2环鸟甘酸门控通道、胰岛素受体DAF-2、神经肽受体NPR-1、泛素化E3连接酶HECW-1、和TGF-β 途径的DAF-7/DAF-1则参与线虫本能的病原逃避行为调控.相较于依赖学习的病原逃避行为, 即需要病原细菌有预暴露产生记忆而发生逃避, 沙雷式菌的次级代谢产物W2则可以直接使线虫产生本能的逃避反应.研究发现线虫的这种逃避反应依赖于AWB神经元 [19], G蛋白ODR-3、受体激酶GRK-2、TAX-4/TAX-2环鸟甘酸门控通道以及Toll-like受体Tol-1参与了该逃避行为的调控[8, 19](图3).

图3 秀丽隐杆线虫病原本能逃避行为调控通路示意图Fig.3 The moldel of C.elegans for pathogen adoidance independent with learniny

此外, 当线虫被革兰氏阳性细菌苏云金芽孢杆菌侵染时, 血清素信号途径可产生2条不同的应对策略:逃避和咽喉运动.研究发现daf-2突变引起线虫咽喉运动减慢从而减弱对病原物质的吸入, 同时加快线虫逃离苏云金芽孢杆菌的速度.这2种行为的应答十分迅速, 属于线虫对病原菌本能的应激反应[20](图3).

神经肽受体NPR-1和泛素化E3连接酶HECW-1则是另一条调控线虫本能的病原逃避行为通路.npr-1基因调控了多种食物依赖性行为, 如聚集、趋氧和运动等行为[21, 22, 23].近年来研究发现NPR-1 215F菌株CB4856 (Hawaiian strain)与野生型菌株N2相比, npr-1基因的活性降低, 该菌株花费更多的时间停留在致病细菌中, 因与致病菌接触时间延长而使致死率增加[21, 24, 25].泛素化E3连接酶HECW-1可抑制该行为, 表现为hecw-1突变株线虫逃避反应增强.此外, 通过在不同的神经元中对HECW-1进行回复发现HECW-1作用于NPR-1上游并在OLL神经元发挥功能[24](图3).

3 结论与展望

动物肠道对病原细菌十分易感, 铜绿假单胞菌和沙门氏菌能在线虫肠道繁殖, 并在数天内将其杀死, 线虫通过启动天然免疫和逃避行为进行自我保护.其中胰岛素信号途径同时参与了天然免疫和病原逃避行为的调控.在天然免疫调控过程中, AGE-1激活DAF-2调控下游DAF-16的转录, 使DAF-16入核调控应对应激反应的基因表达[26, 27].对于病原逃避行为调控, 胰岛素途径在不同的神经元中功能不同.例如, ISN-1在中间神经元AIA中表达, 激活ASER神经元的受体DAF-2调控线虫对盐离子的逃避行为[28], ISN-1作用于中间神经元AIY、AIZ和RIA的受体AGE-1调控线虫逃避行为[29].此外, 在AWC神经元中AGE-1调控线虫对气味的趋避行为[30].

多种神经递质和一元胺参与线虫对病原细菌的逃避行为调控, 目前报道较多的是ADF神经元产生的血清素和中间神经元上的受体NPR-1.其它的一元胺, 章鱼胺参与了线虫饥饿行为调控[31], 多巴胺参与线虫对高温的物理性逃避行为[32, 33].此外, 五羟色胺和多巴胺被认为是食物信号, 在培养基中的五羟色胺等同于食物信号, 相反的多巴胺为饥饿信号.线虫对温度、气味、和盐离子的可塑性逃避行为多与饥饿相关, 因此五羟色胺和多巴胺间接调控了线虫对环境因子的可塑逃避行为.综上所述多条信号通路同时参与了线虫对病原细菌和环境因子的逃避行为调控.

自然环境中的线虫面对复杂的生活环境, 进化出复杂的适应机制.当线虫遭遇高温、高盐、饥饿、病原菌侵染等不利因素时, 正是在多条信号通路的协同作用下提高线虫的应激能力、天然免疫和激活逃避行为, 增加了线虫不利条件下的生存机率.而对其中所涉及通路尤其是保守通路的研究, 相信对理解高等生物的环境适应亦具有指导意义.

The authors have declared that no competing interests exist.

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