(p)ppGpp——“魔斑”核苷酸在细菌中的研究进展

鸟苷四磷酸（guanosine tetraphosphate,ppGpp）/鸟苷五磷酸（guanosine pentaphosphate,pppGpp）是细菌严谨反应的信号分子,其合成和水解由Rel/SpoT同系物（RelA/SpoT homologue,RSH）家族的蛋白质合成和水解活性控制。（p）ppGpp介导的严谨反应能够提高细菌对营养匮乏的适应能力和抗生素抗性。近年来发现（p）ppGpp与细菌生长和细胞分裂、抗生素合成等都密切相关,是细胞内重要的全局调控因子。（p）ppGpp在细菌细胞中有许多靶点,使其可以调节DNA复制、转录、细胞周期、核糖体生物合成以及抗生素合成基因簇的表达。然而,（p）ppGpp如何控制转录和其他代谢过程取决于细菌种类,并在不同的微生物中通过不同的机制调节相同的过程。因此,本文通过综述（p）ppGpp的合成/水解酶的种类和调节机制,（p）ppGpp对微生物代谢调控机制、对细胞周期的影响机制,以及（p）ppGpp对抗生素合成和耐受性的调控机制,为细菌耐药性研究和细胞生理学研究奠定基础。     

信号分子ppGpp与微生物环境适应性

微生物能感知环境胁迫信号,通过触发严谨反应对生长速率进行调节,并通过一系列代谢调控,使细胞能在不利环境中生存。高度磷酸化的鸟苷四/五磷酸ppGpp/pppGpp（文中以ppGpp统称）作为信号分子对微生物生理具有广泛的调节作用,至今仍是微生物学研究热点之一。ppGpp对于微生物适应高温、高压等环境起到了积极的作用。综述了信号分子ppGpp合成降解机制及其调控微生物适应性方面的研究进展。     

细菌应激反应中(p)ppGpp代谢的调控

(p)ppGpp是介导细菌细胞对环境胁迫产生应激反应的重要胞内信号,通过控制一系列重要的细胞活动使细菌得以生存。通过对蓝细菌中(p)ppGpp代谢的研究,对(p)ppGpp作用机制、控制(p)ppGpp代谢的酶系统、环境胁迫信号传递、细胞中(p)ppGpp水平的调控及其多样性进行了总结。     

集胞藻PCC6803 中relA/spoT 同源基因syn-rsh (slr1325)的鉴定

[目的]四磷酸或五磷酸鸟苷(Guanosine 3′,5′-bispyrophosphate,(p)ppGpp)是细菌在遭遇环境胁迫时细胞产生应激反应的信号分子,(p)ppGpp由其合成酶RelA或具有合成酶或水解酶双重催化功能的RelA/SpoT合成.本文证明了集胞藻PCC6803(Synechocystis sp.)中唯一编码RelA/SpoT同源蛋白(命名为Syn-RSH)的基因slr1325(syn-rsh)的功能.[方法]通过互补试验证明syn-rsh表达产物的生物学功能;以纤维素薄层层析检测不同条件下Escherichia coli(p)ppGpp合成缺陷突变株及集胞藻PCC6803细胞中的(p)ppGpp.[结果]诱导Syn-RSH表达可使(p)ppGpp合成酶和水解酶基因缺失的E.coli突变株回复野生型表型,并在细胞中积累一定水平的ppGpp;在实验室培养条件下,集胞藻PCC6803细胞中可检测到低水平的ppGpp,氨基酸饥饿可诱导ppGpp水平升高并维持在相应水平.[结论]Syn-RSH具有(p)ppGpp合成酶和水解酶的双重功能,(p)ppGpp是集胞藻PCC6803在实验室生长条件下细胞生长所必需的.     

“严谨反应”还是“严紧反应”?

“Stringent response”是指细菌在遭受营养饥饿与环境胁迫时,由代谢酶RelA/SpoT催化合成信号分子鸟苷四/五磷酸[(p)ppGpp],从而诱导细菌细胞关闭rRNA、tRNA及核糖体蛋白基因转录,停止多种蛋白质的翻译,严控大部分代谢活动的一系列适应性基因表达过程。“Stringent response”几乎是所有细菌应对逆境的重要调节机制。目前,国内文献对“stringent response”的中文翻译存在“严谨反应”和“严紧反应”混用的现象。基于此,本文对“stringent response”的调控机制、生理功能及字面含义进行了分析,认为“stringent response”翻译为“严紧反应”更为合理、准确。     

cGAS-STING通路在细菌感染中的作用

环磷酸鸟苷-腺苷合成酶(cyclic guanosine monophosphate-adenosine monophosphate synthase,c GAS,又称为MB21D1或C6orf150)是一种胞质DNA感受器,能识别胞质中的双链DNA(double strand DNA,ds DNA),并形成2∶2型复合物,催化腺苷三磷酸(adenosine triphosphate,ATP)和鸟苷三磷酸(guanosine triphosphate,GTP)形成环鸟苷酸腺苷酸(cyclic guanosine monophosphate-adenosine monophosphate,c GAMP)。c GAMP作为第二信使激活干扰素基因刺激分子(stimulator of interferon gene,STING),进而产生Ⅰ型干扰素及其他细胞因子。近年来,多项研究表明,c GAS-STING通路在病毒感染免疫中发挥重要作用,包括人类免疫缺陷病毒(human immunodeficiency virus,HIV)、人乳头瘤病毒(human papilloma virus,HPV)和乙型肝炎病毒(hepatitis B virus,HBV)等,但关于c GAS在细菌感染中的作用的研究较少。就近年来国内外对c GAS及c GAS在细菌感染中的作用作一综述,为其在细菌感染相关的应用研究提供思路和借鉴。     

cGAS/STING通路的生物学功能及其在细菌感染中的作用北大核心

先天免疫是宿主抵御外来病原体入侵的第一道防线,而模式识别受体(pattern recognition receptors,PRRs)是介导先天免疫应答关键分子。PRRs通过识别病原体相关分子模式(pathogen associated molecular patterns,PAMPs)来激活宿主先天免疫反应。二十一世纪先天免疫领域里程碑式发现-环磷酸鸟苷腺苷合成酶(cyclic GMP-AMP synthase,cGAS),cGAS在宿主先天免疫过程中发挥重要作用,通过识别外源DNA产生第二信使2’,3’-环化鸟苷酸腺苷酸(2’,3’-cyclic guanosine monophosphate adenosine monophosphate,2’,3’-cGAMP)来介导干扰素基因刺激因子(stimulator of interferon genes,STING)的活化,从而促进下游干扰素(IFN)和其他细胞因子分泌来发挥宿主的抗病毒反应。近年研究发现,cGAS/STING通路在宿主抗细菌感染过程中发挥着重要作用,同时细菌也进化出不同机制来拮抗cGAS/STING通路。本文主要对cGAS/STING通路的生物学功能及其在细菌感染中的作用进行综述,为进一步研发新型抗菌药物提供理论参考。     

原绿球藻对环境胁迫的生理和分子响应机制

原绿球藻(Prochlorococcus)作为海洋丰度最高的浮游植物,对海洋生态系统的物质循环和能量流动起着重要的驱动作用。原绿球藻生长和光合作用活性容易受到环境胁迫的影响,进而影响整个海洋生态系统的稳定性。因此,研究原绿球藻应对环境胁迫的响应机制具有重要的生态意义。原绿球藻主要通过分化出不同的生态型来适应不同光照和营养盐的海洋环境,但仍然会很难快速适应各种突如其来的海洋环境变化。本文从原绿球藻应对环境胁迫的角度,探讨了其生理和分子响应机制的最新研究进展,包括光系统I循环电子传递在光照变化时发挥的重要作用,通过RNA快速响应而调控基因表达应对环境胁迫,以及在辅助异养细菌的保护下应对活性氧的胁迫等。本文也展望了原绿球藻对环境胁迫响应的生理和分子机制的未来研究方向,旨在为原绿球藻抗逆机制的深入研究提供参考。     

多重耐药寡养单胞菌的基因组学研究进展

寡养单胞菌(Stenotrophomonas)是一类广泛分布于自然环境中的革兰氏阴性非发酵细菌,环境适应能力较强,尤其具备高度耐受抗生素的能力,被认为是耐药基因(antimicrobial resistance,AMR)传播的“物流转运仓库”。临床上寡养单胞菌检出率逐年提高,且耐受抗生素能力及其种类呈现增加态势。本文围绕寡养单胞菌基因组编码的多样性耐药因子,结合基因组学工具开发、抗生素耐药分子机制的研究进展,针对耐药相关因子所涉及的进化/变异和序列编辑技术展开综述。寡养单胞菌的比较基因组学(comparative genomics)研究,可推动耐药菌高效监测与扩散风险防控、解析微生物适应环境关键机制和设计靶向药物。     

细菌耐药性应对策略研究进展

细菌耐药性(Antimicrobial resistance,AMR)持续增长,但新上市抗生素数量却持续下降。抗生素耐药基因(Antimicrobial resistance gene,ARG)和抗生素耐药菌感染已严重威胁人类健康。因此,需要多方面联合采取措施来应对AMR所带来的各种挑战,包括创新生物医药、改善抗生素使用和抗生素耐药监测系统、减少抗生素耐药基因产生速度、阻止健康护理相关感染和多重抗生素耐药菌传播与扩散、开发微生物学快速诊断方法与设备、减少临床和兽医抗生素滥用等。庆幸的是,AMR已受到各国政要、科学家和企业家等的高度重视与支持,相信随着新技术、新产品的不断问世和管理新措施的不断出台,AMR问题一定会得到控制和缓解。     

Stress-induced Antibiotic Susceptibility Testing on a Chip

We have developed a rapid microfluidic method for antibiotic susceptibility testing in a stress-based environment. Fluid is passed at high speeds over bacteria immobilized on the bottom of a microfluidic channel. In the presence of stress and antibiotic, susceptible strains of bacteria die rapidly. However, resistant bacteria survive these stressful conditions. The hypothesis behind this method is new: stress activation of biochemical pathways, which are targets of antibiotics, can accelerate antibiotic susceptibility testing. As compared to standard antibiotic susceptibility testing methods, the rate-limiting step - bacterial growth - is omitted during antibiotic application. The technical implementation of the method is in a combination of standard techniques and innovative approaches. The standard parts of the method include bacterial culture protocols, defining microfluidic channels in polydimethylsiloxane (PDMS), cell viability monitoring with fluorescence, and batch image processing for bacteria counting. Innovative parts of the method are in the use of culture media flow for mechanical stress application, use of enzymes to damage but not kill the bacteria, and use of microarray substrates for bacterial attachment. The developed platform can be used in antibiotic and nonantibiotic related drug development and testing. As compared to the standard bacterial suspension experiments, the effect of the drug can be turned on and off repeatedly over controlled time periods. Repetitive observation of the same bacterial population is possible over the course of the same experiment.     

大熊猫源细菌耐药性研究进展

耐药菌和耐药基因已成为一种新型环境污染物,引发世界公共卫生问题。细菌耐药性尤其是多重耐药菌在人医临床、畜禽养殖以及环境传播等多个方面得到越来越多的关注,而关于大熊猫等野生动物的耐药性研究相对较少。大熊猫(Ailuropoda melanoleuca)是世界公认的珍稀野生动物,其种群数量易受到各种疾病的威胁,尤其是肠道细菌性疾病。随着抗菌药物在疾病预防和控制中的普遍使用,由此带来的耐药性危害日益明显。本文总结了关于大熊猫源细菌耐药的国内外研究报道,对其耐药表型、耐药基因型、耐药机制及水平传播机制等方面内容进行了综述,旨在为大熊猫源细菌耐药性的研究和防控提供依据,为临床科学用药提供理论参考,从而助力大熊猫迁地保护。     

毒素-抗毒素系统介导滞留菌形成机制

滞留菌是一类处于低代谢休眠状态的抗生素耐受细菌亚群，能够在致死性压力应激后存活下来，是抗生素治疗失败和复发性感染的主要原因之一。毒素-抗毒素系统(toxin-antitoxin system, TA)作为压力应激模块普遍存在于各种细菌中，由稳定的毒素和不稳定但可以中和毒素的同源抗毒素组成。压力情况下，第二信使(p)ppGpp激活Lon,随后大多数II型TA系统被激活，诱导滞留菌形成。同样在(p)ppGpp存在的情况下，Obg刺激hokB转录，使毒素积累，抑制细菌DNA复制、转录、翻译等重要的生理过程，驱动细菌形成滞留菌。SOS反应是激活TA系统的另一个主要途径，解除了对tisB转录的抑制，使其在细胞内积累并插入细胞膜，破坏质子动力势，降低胞内ATP水平，诱使休眠和滞留菌形成。讨论TA系统介导滞留菌形成的机制有助于提出新型抗菌策略。     

Emerging and divergent roles of pyrophosphorylated nucleotides in bacterial physiology and pathogenesis

Bacteria inhabit diverse environmental niches and consequently must modulate their metabolism to adapt to stress. The nucleotide second messengers guanosine tetraphosphate (ppGpp) and guanosine pentaphosphate (pppGpp) (collectively referred to as (p)ppGpp) are essential for survival during nutrient starvation. (p)ppGpp is synthesized by the RelA-SpoT homologue (RSH) protein family and coordinates the control of cellular metabolism through its combined effect on over 50 proteins. While the role of (p)ppGpp has largely been associated with nutrient limitation, recent studies have shown that (p)ppGpp and related nucleotides have a previously underappreciated effect on different aspects of bacterial physiology, such as maintaining cellular homeostasis and regulating bacterial interactions with a host, other bacteria, or phages. (p)ppGpp produced by pathogenic bacteria facilitates the evasion of host defenses such as reactive nitrogen intermediates, acidic pH, and the complement system. Additionally, (p)ppGpp and pyrophosphorylated derivatives of canonical adenosine nucleotides called (p)ppApp are emerging as effectors of bacterial toxin proteins. Here, we review the RSH protein family with a focus on its unconventional roles during host infection and bacterial competition.     

Stress-Induced Mutagenesis in Bacteria

ABSTRACT

Bacteria spend their lives buffeted by changing environmental conditions. To adapt to and survive these stresses, bacteria have global response systems that result in sweeping changes in gene expression and cellular metabolism. These responses are controlled by master regulators, which include: alternative sigma factors, such as RpoS and RpoH; small molecule effectors, such as ppGpp; gene repressors such as LexA; and, inorganic molecules, such as polyphosphate. The response pathways extensively overlap and are induced to various extents by the same environmental stresses. These stresses include nutritional deprivation, DNA damage, temperature shift, and exposure to antibiotics. All of these global stress responses include functions that can increase genetic variability. In particular, up-regulation and activation of error-prone DNA polymerases, down-regulation of error-correcting enzymes, and movement of mobile genetic elements are common features of several stress responses. The result is that under a variety of stressful conditions, bacteria are induced for genetic change. This transient mutator state may be important for adaptive evolution.     

基于代谢组学的抗生素与细菌间作用研究进展

抗生素杀菌是一个复杂的生理过程,杀菌抗生素与靶点作用后的下游代谢变化与抗生素作用效果紧密联系,其通过干扰细菌代谢状态加速死亡进程,而细菌改变代谢状态也能影响抗生素的有效性.代谢组学通过监测细菌在抗生素作用下的变化提供全面代谢信息,我们回顾近年来基于代谢组学对抗生素与细菌间作用的研究进展,以期为开发抗生素佐剂提高抗生素效...