BACKGROUND & AIMS: :Understanding the regulation of human food intake in response to an acute exercise session is of importance for interventions with athletes and soldiers, as well as overweight individuals. However, the influence of hot and cold environments on this crucial function for the regulation of body mass and motor performance has not been summarized. The purpose of this review was to exhaustively search the literature on the effect of ambient temperature during an exercise session on the subsequent subjective feeling of appetite, energy intake (EI) and its regulation. In the absence of stress due to environmental temperature, exercise-induced energy expenditure is not compensated by EI during an ad libitum meal following the session, probably due to decreased acylated ghrelin and increased peptide tyrosine tyrosine (PYY), glucagon-like peptide 1 (GLP-1), and pancreatic polypeptide (PP) levels. No systematic analysis has been yet made for major alterations of relative EI in cold and hot environments. However, observed eating behaviors are altered (proportion of solid/liquid food, carbohydrate/fat) and physiological regulation appears also to be altered. Anorexigenic signals, particularly PYY, appear to further increase in hot environments than in those that are thermoneutral. Ghrelin and leptin may be involved in the observed increase in EI after exercise in the cold, in parallel with increased energy expenditure. The potential influence of ambient thermal environment on eating behaviors after an exercise session should not be neglected.

背景与目标： : 了解对急性运动过程中人类食物摄入量的调节对于对运动员和士兵以及超重个体的干预至关重要。但是，尚未总结冷热环境对调节体重和运动性能的关键功能的影响。这篇综述的目的是详尽地检索有关运动过程中环境温度对随后的主观食欲，能量摄入 (EI) 及其调节的影响的文献。在没有由于环境温度引起的压力的情况下，运动引起的能量消耗在饮食后的饮食过程中未被EI补偿，这可能是由于酰化ghrelin降低和肽酪氨酸 (PYY)，胰高血糖素样肽1 (GLP-1)，和胰多肽 (PP) 水平。尚未对冷热环境中相对EI的主要变化进行系统分析。但是，观察到的饮食行为发生了变化 (固体/液体食物，碳水化合物/脂肪的比例)，生理调节似乎也发生了变化。在炎热的环境中，厌食信号 (尤其是PYY) 似乎比在热中性环境中进一步增加。Ghrelin和leptin可能与寒冷运动后观察到的EI增加有关，同时增加了能量消耗。不应忽略运动后周围热环境对饮食行为的潜在影响。

BACKGROUND & AIMS: :Our knowledge on the physiological function of the insect Neuropeptide F (NPF) mostly comes from studies in the fruit fly, Drosophila melanogaster, where NPF was shown to regulate diverse processes, such as feeding, learning and responding to stress. In the desert locust, Schistocerca gregaria, only a truncated form of the "full-length" NPF (the biologically active "trNPF") has been isolated. In this study, we investigated whether this peptide is involved in the regulation of feeding in this orthopteran species. In the S. gregaria EST-database, an NPF-precursor encoding transcript was found. Alignment with other insect NPF-precursors showed relatively highest sequence conservation within the trNPF region (and the flanking dibasic cleavage site), as compared to other regions of the NPF-precursor. Quantitative real-time RT-PCR revealed that the Schgr-NPF-precursor encoding transcript occurs throughout the central nervous system with relatively high transcript levels in the brain, optic lobes and suboesophageal ganglion. It was also detected at relatively high levels in the midgut, which suggests that the encoded peptide also functions in the digestive system. Moreover, Schgr-NPF-transcript levels were notably higher in starved animals than in animals fed ad libitum, while transcript levels were also shown to be regulated after the consumption of a meal. Injection of locust trNPF in adults stimulated food intake, while RNAi knockdown reduced food intake. Furthermore, injection of trNPF in adults stimulated weight increase, while RNAi knockdown reduced weight gain. This effect of trNPF on body weight gain may result from its stimulatory effect on food intake. Taken together, we provide clear evidence for an important role of trNPF in the regulation of feeding in the desert locust, S. gregaria.

背景与目标： : 我们对昆虫神经肽F (NPF) 的生理功能的了解主要来自果蝇果蝇的研究，其中NPF被证明可以调节多种过程，例如进食，学习和应对压力。在沙漠蝗虫中，仅分离出 “全长” NPF (具有生物活性的 “trNPF”) 的截短形式。在这项研究中，我们调查了该肽是否参与了该直翅目物种的进食调节。在S. gregaria EST数据库中，发现了NPF前体编码转录本。与NPF前体的其他区域相比，与其他昆虫NPF前体的比对在trNPF区域 (和侧翼的二元切割位点) 内显示出相对最高的序列保守性。实时定量rt-pcr显示，Schgr-NPF前体编码转录本发生在整个中枢神经系统中，在大脑，视神经叶和食管下神经节中具有相对较高的转录本水平。在中肠中也检测到相对较高的水平，这表明编码的肽也在消化系统中起作用。此外，饥饿动物的Schgr-NPF转录水平明显高于随意喂养的动物，而食用餐后转录水平也受到调节。成人注射刺槐trNPF刺激了食物摄入，而RNAi敲低则减少了食物摄入。此外，在成年人中注射trNPF可刺激体重增加，而RNAi敲低可减少体重增加。trNPF对体重增加的这种影响可能是由于其对食物摄入的刺激作用所致。总而言之，我们为trNPF在沙漠蝗虫 (S. gregaria) 的摄食调节中的重要作用提供了明确的证据。

BACKGROUND & AIMS: Hormonal fluctuations associated with the menstrual cycle influence appetite control and eating behaviour. Energy intake varies during the reproductive cycle in humans and animals, with a periovulatory nadir and a luteal phase peak. Patterns of macronutrient selection show less consistency but a number of studies report carbohydrate cravings in the premenstrual phase, particularly in women with premenstrual syndrome. The cyclical nature of food cravings are frequently, but not invariably, associated with depression. Fluctuations in appetite, cravings and energy intake during the menstrual cycle may occur in parallel with cyclical rhythms in serotonin, which can be accompanied by affective symptoms. The premenstrual phase can be considered as a time when women are especially vulnerable to overconsumption, food craving and depression; this is often associated with low serotonin activity.

背景与目标： 与月经周期相关的荷尔蒙波动会影响食欲控制和饮食行为。在人类和动物的生殖周期中，能量摄入有所不同，具有排卵周期的最低点和黄体相峰。大量营养素选择的模式显示出较少的一致性，但许多研究报告称在经前期阶段对碳水化合物的渴望，尤其是在患有经前期综合征的女性中。食物渴望的周期性经常但并非总是与抑郁症有关。月经周期中食欲，渴望和能量摄入的波动可能与血清素的周期性节律同时发生，这可能伴有情感症状。经前期可以被认为是女性特别容易过度消费，食物渴望和抑郁的时期; 这通常与血清素活性低有关。

BACKGROUND & AIMS: OBJECTIVE:The intracellular signals that contribute to granulocyte colony-stimulating factor (G-CSF) receptor induced stem cell mobilization are poorly characterized. METHODS:We show enhanced G-CSF induced mobilization of stem cells in mice deficient in expression of Src family kinases (SFK-/-), which is associated with hypersensitivity of SFK-/- bone marrow cells to G-CSF as well as sustained activation of signal transducer and activator of transcription-3. RESULTS:A proteome map of the bone marrow fluid derived from wild-type and SFK-/- mice revealed a significant global reduction in the number of proteins in SFK-/- mice compared to controls, which was associated with elevated matrix metalloproteinase-9 levels, reduced stromal-derived factor-1 expression, and enhanced breakdown of vascular cell adhesion molecule-1. Transplantation of wild-type or SFK-/- stem cells into wild-type mice and treatment with G-CSF recapitulated the G-CSF-induced increase in stem cell mobilization noted in SFK-/- nontransplanted mice; however, the increase was significantly less. G-CSF treatment of SFK-/- mice engrafted with wild-type stem cells also demonstrated a modest increase in stem cell mobilization compared to controls, however, the observed increase was greatest in mice completely devoid of SFKs. CONCLUSIONS:These data suggest an involvement of both hematopoietic intrinsic and microenvironmental factors in Src kinase-mediated mobilization of stem cells and identify Src kinases as potential targets for modulating stem cell mobilization.

背景与目标：

BACKGROUND & AIMS: :Class-switch recombination (CSR) is essential for humoral immunity. However, the regulation of CSR is not completely understood. Here we demonstrate that phosphatidylinositol 3-kinase (PI3K) actively suppressed the onset and frequency of CSR in primary B cells. Consistently, mice lacking the lipid phosphatase, PTEN, in B cells exhibited a hyper-IgM condition due to impaired CSR, which could be restored in vitro by specific inhibition of PI3Kdelta. Inhibition of CSR by PI3K was partially dependent on the transcription factor, BLIMP1, linking plasma cell commitment and cessation of CSR. PI3K-dependent activation of the serine-threonine kinase, Akt, suppressed CSR, in part, through the inactivation of the Forkhead Box family (Foxo) of transcription factors. Reduced PI3K signaling enhanced the expression of AID (activation-induced cytidine deaminase) and accelerated CSR. However, ectopic expression of AID could not fully overcome inhibition of CSR by PI3K, suggesting that PI3K regulates both the expression and function of AID.

背景与目标： : 类别转换重组 (CSR) 对于体液免疫至关重要。然而，对企业社会责任的监管还没有完全理解。在这里，我们证明了磷脂酰肌醇3-激酶 (PI3K) 可以有效抑制原代b细胞中CSR的发作和频率。一致地，由于CSR受损，b细胞中缺乏脂质磷酸酶PTEN的小鼠表现出高IgM状态，可以通过特异性抑制PI3Kdelta在体外恢复。PI3K对CSR的抑制部分取决于转录因子BLIMP1，该转录因子将浆细胞的承诺与CSR的停止联系起来。丝氨酸-苏氨酸激酶Akt的PI3K-dependent激活部分通过转录因子的叉头盒家族 (Foxo) 的失活来抑制CSR。减少的PI3K信号增强了AID (激活诱导的胞苷脱氨酶) 的表达并加速了CSR。但是，AID的异位表达不能完全克服PI3K对CSR的抑制，这表明PI3K调节了AID的表达和功能。

BACKGROUND & AIMS: A variety of experimental models indicate that programmed cell death, or apoptosis, of lymphocytes is a key mechanism in the homeostatic regulation of immunity. Apoptosis is important in early B- and T-cell development to delete cells with nonfunctional antigen receptors, and is also critical for censoring self-reactive cells at the immature lymphocyte stage and at various stages after lymphocytes reach maturity. In this article we focus on the role of the apoptosis regulatory gene bcl-x in controlling survival during lymphocyte development and following B- and T-cell activation. Interesting parallels are observed for bcl-x expression between the B- and T-lineages. The available data also indicate that bcl-x and bcl-2 are expressed in reciprocal patterns during the lifespan of a lymphocyte, suggesting unique regulatory roles for these two survival proteins.

背景与目标： 多种实验模型表明，淋巴细胞的程序性细胞死亡或凋亡是免疫稳态调节的关键机制。凋亡在早期b细胞和T细胞发育中很重要，可以删除具有无功能抗原受体的细胞，并且对于在未成熟淋巴细胞阶段和淋巴细胞成熟后的各个阶段审查自我反应细胞也至关重要。在本文中，我们专注于凋亡调节基因bcl-x在控制淋巴细胞发育期间以及b细胞和T细胞活化后的存活中的作用。在B谱系和T谱系之间观察到bcl-x表达的有趣的相似之处。现有数据还表明，bcl-x和bcl-2在淋巴细胞的寿命期间以相互模式表达，这表明这两种存活蛋白具有独特的调节作用。

BACKGROUND & AIMS: :In rat frontal cortex, extracellular levels of glutamate are raised by the anti-psychotic drug clozapine. We have recently shown that a significant reduction in the levels of the glutamate transporter GLT-1 may be one of the mechanisms responsible for this elevation. Here we studied whether GLT-1 down-regulation induced by chronic clozapine treatment is associated with changes in the expression of synaptophysin, synaptosome-associated protein of 25 kDa (SNAP-25) and vesicular glutamate transporter 1 (VGLUT1), three major presynaptic proteins involved in neurotransmitter release. Quantitative high-resolution confocal microscopy studies in vivo showed that GLT-1 down-regulation is closely associated with a significant increase in synaptophysin, but not SNAP-25 and VGLUT1, expression. This was confirmed in vitro studies, and in western blotting studies of synaptophysin, SNAP-25 and VGLUT1. In addition, our results show that, following clozapine treatment, synaptophysin expression increases in the very cortical regions in which GLT-1 expression is down-regulated. These findings suggest that part of the effects of clozapine may be exerted via an action on the presynaptic machinery involved in neurotransmitter release.

背景与目标： : 在大鼠额叶皮层中，抗精神病药物氯氮平可提高谷氨酸的细胞外水平。我们最近表明，谷氨酸转运蛋白GLT-1水平的显着降低可能是导致这种升高的机制之一。在这里，我们研究了由慢性氯氮平治疗引起的GLT-1下调是否与突触素，突触体相关蛋白25 kDa (SNAP-25) 和囊泡谷氨酸转运蛋白1 (VGLUT1) 的表达变化有关神经递质释放的三种主要突触前蛋白。体内定量高分辨率共聚焦显微镜研究表明，GLT-1下调与突触素的显着增加密切相关，但与SNAP-25和VGLUT1的表达无关。这在体外研究以及突触素，SNAP-25和vglut1的蛋白质印迹研究中得到了证实。此外，我们的结果表明，氯氮平治疗后，突触素表达在GLT-1表达下调的皮质区域增加。这些发现表明，氯氮平的部分作用可能是通过对参与神经递质释放的突触前机制的作用来发挥的。

BACKGROUND & AIMS: The transcription factor E2F-1 interacts stably with cyclin A via a small domain near its amino terminus and is negatively regulated by the cyclin A-dependent kinases. Thus, the activities of E2F, a family of transcription factors involved in cell proliferation, are regulated by at least two types of cell growth regulatorsthe retinoblastoma protein family and the cyclin-dependent kinase family. To investigate further the regulation of E2F by cyclin-dependent kinases, we have extended our studies to include additional cyclins and E2F family members. Using purified components in an in vitro system, we show that the E2F-1-DP-1 heterodimer, the functionally active form of the E2F activity, is not a substrate for the active cyclin D-dependent kinases but is efficiently phosphorylated by the cyclin B-dependent kinases, which do not form stable complexes with the E2F-1-DP-1 heterodimer. Phosphorylation of the E2F-1-DP-1 heterodimer by cyclin B-dependent kinases, however, did not result in down-regulation of its DNA-binding activity, as is readily seen after phosphorylation by cyclin A-dependent kinases, suggesting that phosphorylation per se is not sufficient to regulate E2F DNA-binding activity. Furthermore, heterodimers containing E2F-4, a family member lacking the cyclin A binding domain found in E2F-1, are not efficiently phosphorylated or functionally down-regulated by cyclin A-dependent kinases. However, addition of the E2F-1 cyclin A binding domain to E2F-4 conferred cyclin A-dependent kinase-mediated down-regulation of the E2F-4-DP-1 heterodimer. Thus, both enzymatic phosphorylation and stable physical interaction are necessary for the specific regulation of E2F family members by cyclin-dependent kinases.

背景与目标： 转录因子E2F-1通过其氨基末端附近的小结构域与细胞周期蛋白A稳定相互作用，并被细胞周期蛋白a依赖性激酶负调控。因此，E2F (参与细胞增殖的转录因子家族) 的活性受至少两种类型的细胞生长调节剂 (视网膜母细胞瘤蛋白家族和细胞周期蛋白依赖性激酶家族) 调节。为了进一步研究细胞周期蛋白依赖性激酶对E2F的调节，我们已将研究扩展到包括其他细胞周期蛋白和E2F家族成员。在体外系统中使用纯化的组分，我们表明E2F-1-DP-1异二聚体，即E2F活性的功能活性形式，不是活性细胞周期蛋白D依赖性激酶的底物，而是被细胞周期蛋白B依赖性激酶有效磷酸化，其不与E2F-1-DP-1异二聚体形成稳定的复合物。然而，细胞周期蛋白B依赖性激酶对E2F-1-DP-1异二聚体的磷酸化并未导致其DNA结合活性的下调，正如在细胞周期蛋白A依赖性激酶磷酸化后容易看到的那样，这表明磷酸化本身不足以调节e2fdna结合活性。此外，含有E2F-4的异二聚体 (缺乏E2F-1中发现的细胞周期蛋白a结合结构域的家族成员) 不能被细胞周期蛋白A依赖性激酶有效地磷酸化或功能下调。然而，将E2F-1细胞周期蛋白A结合结构域添加到E2F-4赋予细胞周期蛋白A依赖性激酶介导的E2F-4-DP-1异二聚体的下调。因此，酶促磷酸化和稳定的物理相互作用对于细胞周期蛋白依赖性激酶对E2F家族成员的特异性调节都是必需的。

BACKGROUND & AIMS: :Auto-anti-idiotypic mechanisms can regulate the protective immune response against Schistosoma mansoni. Anti-idiotypic responses were stimulated by immunization of mice either with nonspecifically induced lymphoblasts, produced with Con A, or with Ag-induced lymphoblasts bearing specific idiotypic receptors. The effect of the induced anti-idiotypic response upon clonotypic cellular reactivity was assessed in vitro through the suppression of antigen-mediated blast transformation by cloned T cells and in vivo by suppression of resistance to S. mansoni and delayed-type hypersensitivity responses against specific Ag. Differential regulation of humoral immune responses was studied at the levels of specific epitopic recognition, the expression of specific Id, and the production of anti-idiotypic responses directed against mAb bearing specific Id. Anti-idiotypic sensitization resulted in variable (10 to 90%) suppression of the immune response to discrete antigenic epitopes, the expression of specific idiotypic phenotypes, and anti-idiotypic, antiparatopic responses against T cell clonotypes and antibody idiotypic phenotypes. In vitro admixture and in vivo challenge studies resulted in consonant differential suppression. Thus idiotypic regulation can mold the fine specificities of the protective immune response to S. mansoni at the clonal level and may provide an approach to optimize the expression and assessment of resistance.

背景与目标： : 自身抗独特型机制可以调节曼氏血吸虫的保护性免疫反应。通过用Con A产生的非特异性诱导的淋巴母细胞或带有特定独特型受体的Ag诱导的淋巴母细胞免疫小鼠来刺激抗独特型反应。诱导的抗独特型反应对克隆型细胞反应性的影响在体外通过克隆的T细胞抑制抗原介导的胚转化进行评估，并在体内通过抑制对曼氏杆菌的抗性和对特异性Ag的延迟型超敏反应进行评估。在特定的表位识别，特定Id的表达以及针对带有特定Id的mAb的抗独特型反应的产生的水平上研究了体液免疫反应的差异调节。抗独特型致敏作用导致对离散抗原表位的免疫应答、特定独特型表型的表达以及针对T细胞克隆型和抗体独特型表型的抗独特型、抗独特型应答的可变 (10至90%) 抑制。体外混合和体内激发研究导致辅音差异抑制。因此，独特型调节可以在克隆水平上塑造对曼氏杆菌的保护性免疫反应的精细特异性，并可能提供一种优化表达和评估抗性的方法。

BACKGROUND & AIMS: :The size of the pool of growing follicles was normal after prolonged down-regulation, as indicated by normal AMH levels 4 and 8 weeks after goserelin administration. However, there was a profound down-regulation of LH levels; therefore we suggest administration of exogenous LH to proceed to IVF or alternatively stimulation of endogenous LH secretion with daily administration of GnRH agonist. These need to be assessed prospectively.

背景与目标： : 长时间下调后，卵泡生长池的大小正常，如戈舍瑞林给药后4周和8周正常的AMH水平所示。但是，LH水平有明显的下调; 因此，我们建议给予外源性LH进行IVF或通过每天给予GnRH激动剂来刺激内源性LH分泌。这些需要进行前瞻性评估。

BACKGROUND & AIMS: :P strains of Drosophila are distinguished from M strains by having P elements in their genomes and also by having the P cytotype, a maternally inherited condition that strongly represses P-element-induced hybrid dysgenesis. The P cytotype is associated with P elements inserted near the left telomere of the X chromosome. Repression by the telomeric P elements TP5 and TP6 is significantly enhanced when these elements are crossed into M' strains, which, like P strains, carry P elements, but have little or no ability to repress dysgenesis. The telomeric and M' P elements must coexist in females for this enhanced repression ability to develop. However, once established, it is transmitted maternally to the immediate offspring independently of the telomeric P elements themselves. Females that carry a telomeric P element but that do not carry M' P elements may also transmit an ability to repress dysgenesis to their offspring independently of the telomeric P element. Cytotype regulation therefore involves a maternally transmissible product of telomeric P elements that can interact synergistically with products from paternally inherited M' P elements. This synergism between TP and M' P elements also appears to persist for at least one generation after the TP has been removed from the genotype.

背景与目标： : 果蝇的P株与M株的区别在于其基因组中具有P元素，并且还具有P细胞型，这是一种强烈抑制P元素诱导的杂种发育不全的母系遗传性疾病。P细胞类型与插入X染色体左端粒附近的P元件有关。当端粒P元素TP5和TP6杂交成M' 菌株时，它们的抑制作用显着增强，M' 菌株与P菌株一样，带有P元素，但几乎没有抑制发育不全的能力。端粒和m'p元素必须在雌性中共存，以增强这种抑制能力。但是，一旦建立，它就会独立于端粒P元素本身而通过母系传播给直接的后代。携带端粒P元素但不携带m'p元素的雌性也可能独立于端粒P元素向其后代传递抑制发育不全的能力。因此，细胞类型调节涉及端粒P元件的母体可传播产物，该产物可以与父系遗传的m'p元件的产物协同相互作用。TP和m'p元素之间的这种协同作用在TP从基因型中去除后似乎也持续了至少一代。

BACKGROUND & AIMS: :'Citizen participation' includes various participatory techniques and is frequently viewed as an unproblematic and important social good when used as part of the regulation of the innovation and implementation of science and technology. This is perhaps especially evident in debates around 'anticipatory governance' or 'upstream engagement'. Here, we interrogate this thesis using the example of the European Union's regulation of emerging health technologies (such as nanotechnology). In this case, citizen participation in regulatory debate is concerned with innovative objects for medical application that are considered to be emergent or not yet concrete. Through synthesising insights from law, regulatory studies, critical theory, and science and technology studies, we seek to cast new light on the promises, paradoxes, and pitfalls of citizen participation as a tool or technology of regulation in itself. As such we aim to generate a new vantage point from which to view the values and sociotechnical imaginaries that are both 'designed-in' and 'designed-out' of citizen participation. In so doing, we show not only how publics (do not) regulate technologies, but also how citizens themselves are regulated through the techniques of participation.

背景与目标： : “公民参与” 包括各种参与技术，当被用作科学技术创新和实施监管的一部分时，通常被视为毫无问题且重要的社会利益。这在围绕 “预期治理” 或 “上游参与” 的辩论中可能尤其明显。在这里，我们以欧盟对新兴卫生技术 (例如纳米技术) 的监管为例来质疑本论文。在这种情况下，公民参与监管辩论与医疗应用的创新对象有关，这些对象被认为是紧急的或尚未具体的。通过综合法律，监管研究，批判理论以及科学和技术研究的见解，我们寻求对公民参与本身作为监管工具或技术的承诺，悖论和陷阱的新认识。因此，我们的目标是产生一个新的有利位置，从中可以查看公民参与的 “设计” 和 “设计” 的价值观和社会技术想象力。在这样做的过程中，我们不仅展示了公众 (不) 如何监管技术，还展示了公民自己是如何通过参与技术来监管的。

BACKGROUND & AIMS: :An RNA-binding protein called PABPC1 has an important role in determining protein synthesis rates and hypertrophy in the heart.

背景与目标： : 一种称为PABPC1的RNA结合蛋白在确定蛋白质合成速率和心脏肥大方面具有重要作用。

BACKGROUND & AIMS: :The second messenger cyclic dimeric GMP (c-di-GMP) is almost ubiquitous among bacteria as are the c-di-GMP turnover proteins, which mediate the transition between motility and sessility. EAL domain proteins have been characterized as c-di-GMP-specific phosphodiesterases. While most EAL domain proteins contain additional, usually N-terminal, domains, there is a distinct family of proteins with stand-alone EAL domains, exemplified by Salmonella enterica serovar Typhimurium proteins STM3611 (YhjH/PdeH), a c-di-GMP-specific phosphodiesterase, and the enzymatically inactive STM1344 (YdiV/CdgR) and STM1697, which regulate bacterial motility through interaction with the flagellar master regulator, FlhDC. We have analyzed the phylogenetic distribution of EAL-only proteins and their potential functions. Genes encoding EAL-only proteins were found in various bacterial phyla, although most of them were seen in proteobacteria, particularly enterobacteria. Based on the conservation of the active site residues, nearly all stand-alone EAL domains encoded by genomes from phyla other than proteobacteria appear to represent functional phosphodiesterases. Within enterobacteria, EAL-only proteins were found to cluster either with YhjH or with one of the subfamilies of YdiV-related proteins. EAL-only proteins from Shigella flexneri, Klebsiella pneumoniae, and Yersinia enterocolitica were tested for their ability to regulate swimming and swarming motility and formation of the red, dry, and rough (rdar) biofilm morphotype. In these tests, YhjH-related proteins S4210, KPN_01159, KPN_03274, and YE4063 displayed properties typical of enzymatically active phosphodiesterases, whereas S1641 and YE1324 behaved like members of the YdiV/STM1697 subfamily, with Yersinia enterocolitica protein YE1324 shown to downregulate motility in its native host. Of two closely related EAL-only proteins, YE2225 is an active phosphodiesterase, while YE1324 appears to interact with FlhD. These results suggest that in FlhDC-harboring beta- and gammaproteobacteria, some EAL-only proteins evolved to become catalytically inactive and regulate motility and biofilm formation by interacting with FlhDC.IMPORTANCE The EAL domain superfamily consists mainly of proteins with cyclic dimeric GMP-specific phosphodiesterase activity, but individual domains have been classified in three classes according to their functions and conserved amino acid signatures. Proteins that consist solely of stand-alone EAL domains cannot rely on other domains to form catalytically active dimers, and most of them fall into one of two distinct classes: catalytically active phosphodiesterases with well-conserved residues of the active site and the dimerization loop, and catalytically inactive YdiV/CdgR-like proteins that regulate bacterial motility by binding to the flagellar master regulator, FlhDC, and are found primarily in enterobacteria. The presence of apparently inactive EAL-only proteins in the bacteria that do not express FlhD suggests the existence of additional EAL interaction partners.

背景与目标： : 第二信使环二聚体GMP (c-di-GMP) 和c-di-GMP转换蛋白几乎在细菌中无处不在，它们介导了运动性和稳健性之间的过渡。EAL结构域蛋白已被表征为c-二GMP特异性磷酸二酯酶。虽然大多数EAL结构域蛋白都包含额外的 (通常是N末端) 结构域，但有一个独特的具有独立EAL结构域的蛋白质家族，例如沙门氏菌肠道血清型鼠伤寒杆菌蛋白STM3611 (YhjH/PdeH)，一种c-di-GMP特异性磷酸二酯酶，以及酶促失活的STM1344 (YdiV/CdgR) 和STM1697，它们通过与鞭毛主调节剂FlhDC相互作用来调节细菌的运动。我们已经分析了仅EAL蛋白的系统发育分布及其潜在功能。在各种细菌门中都发现了仅编码EAL蛋白的基因，尽管其中大多数在变形杆菌中，尤其是肠杆菌中。基于活性位点残基的保守性，几乎所有由变形杆菌以外的门基因组编码的独立EAL结构域似乎都代表功能性磷酸二酯酶。在肠杆菌中，发现仅EAL蛋白与YhjH或与YdiV相关蛋白的亚科之一聚集。测试了福氏志贺氏菌，肺炎克雷伯菌和小肠结肠炎耶尔森氏菌的仅EAL蛋白调节游泳和蜂群运动以及形成红色，干燥和粗糙 (rdar) 生物膜形态的能力。在这些测试中，YhjH相关蛋白S4210，KPN_01159，KPN_03274和YE4063显示出酶活性磷酸二酯酶的典型特性，而S1641和YE1324的表现类似于YdiV/STM1697亚家族的成员，小肠结肠炎耶尔森氏菌蛋白YE1324显示出下调其天然宿主的运动。在两种密切相关的仅EAL蛋白中，YE2225是一种活性磷酸二酯酶，而YE1324似乎与FlhD相互作用。这些结果表明，在携带 β-和gammaproteobacteria的FlhDC中，一些仅EAL蛋白通过与FlhDC相互作用而进化为催化失活并调节运动和生物膜形成。重要性EAL结构域超家族主要由具有环二聚体GMP特异性磷酸二酯酶活性的蛋白质组成，但是，根据其功能和保守的氨基酸特征，将各个结构域分为三类。仅由独立的EAL结构域组成的蛋白质不能依靠其他结构域来形成催化活性二聚体，并且它们大多数属于两个不同的类别之一: 具有活性位点和二聚化环的保守残基的催化活性磷酸二酯酶，和催化失活的YdiV/CdgR样蛋白，通过与鞭毛主调节剂FlhDC结合来调节细菌的运动，主要在肠杆菌中发现。在不表达FlhD的细菌中存在明显无活性的仅EAL蛋白，这表明存在其他EAL相互作用伙伴。

BACKGROUND & AIMS: :Coenzyme A (CoA) biosynthesis in bacteria and eukaryotes is regulated primarily by feedback inhibition towards pantothenate kinase (PanK). As most archaea utilize a modified route for CoA biosynthesis and do not harbour PanK, the mechanisms governing regulation of CoA biosynthesis are unknown. Here we performed genetic and biochemical studies on the ketopantoate reductase (KPR) from the hyperthermophilic archaeon Thermococcus kodakarensis. KPR catalyses the second step in CoA biosynthesis, the reduction of 2-oxopantoate to pantoate. Gene disruption of TK1968, whose product was 20-29% identical to previously characterized KPRs from bacteria/eukaryotes, resulted in a strain with growth defects that were complemented by addition of pantoate. The TK1968 protein (Tk-KPR) displayed reductase activity specific for 2-oxopantoate and preferred NADH as the electron donor, distinct to the bacterial/eukaryotic NADPH-dependent enzymes. Tk-KPR activity decreased dramatically in the presence of CoA and KPR activity in cell-free extracts was also inhibited by CoA. Kinetic studies indicated that CoA inhibits KPR by competing with NADH. Inhibition of ketopantoate hydroxymethyltransferase, the first enzyme of the pathway, by CoA was not observed. Our results suggest that CoA biosynthesis in T. kodakarensis is regulated by feedback inhibition of KPR, providing a feasible regulation mechanism of CoA biosynthesis in archaea.

背景与目标： : 细菌和真核生物中的辅酶a (CoA) 生物合成主要通过对泛酸激酶 (PanK) 的反馈抑制来调节。由于大多数古细菌利用改良的CoA生物合成途径并且不携带PanK，因此控制CoA生物合成的机制尚不清楚。在这里，我们对来自嗜热古细菌热球菌kodakarensis的酮托酸酯还原酶 (KPR) 进行了遗传和生化研究。KPR催化CoA生物合成的第二步，即2-氧尿囊酸酯还原为泛酸酯。TK1968的基因破坏 (其产物与先前来自细菌/真核生物的KPRs 20-29% 相同) 导致了具有生长缺陷的菌株，该菌株通过添加泛酸补充。TK1968蛋白 (Tk-KPR) 显示出对2-氧尿囊酸酯具有特异性的还原酶活性，并且首选NADH作为电子供体，与细菌/真核NADPH依赖性酶不同。在CoA存在下，tk-kpr活性显着降低，无细胞提取物中的KPR活性也受到CoA的抑制。动力学研究表明，CoA通过与NADH竞争来抑制KPR。未观察到CoA对该途径的第一个酶-酮体羟甲基转移酶的抑制作用。我们的研究结果表明，科达卡氏菌CoA的生物合成受KPR的反馈抑制调节，为古菌CoA的生物合成提供了可行的调控机制。