BACKGROUND & AIMS: :The pH regulation of gene expression in Aspergillus nidulans is mediated by pacC, whose 678 residue-derived protein contains three putative Cys2His2 zinc fingers. Ten pacCc mutations mimicking growth at alkaline pH remove between 100 and 214 C-terminal residues, including a highly acidic region containing an acidic glutamine repeat. Nine pacC+/- mutations mimicking acidic growth conditions remove between 299 and 505 C-terminal residues. Deletion of the entire pacC coding region mimics acidity but leads additionally to poor growth and conidiation. A PacC fusion protein binds DNA with the core consensus GCCARG. At alkaline ambient pH, PacC activates transcription of alkaline-expressed genes (including pacC itself) and represses transcription of acid-expressed genes. pacCc mutations obviate the need for pH signal transduction.

背景与目标： : 构巢曲霉基因表达的pH调节由pacC介导，其678残基衍生蛋白包含三个假定的Cys2His2锌指。在碱性pH下模拟生长的十个pacCc突变去除100和214个C末端残基之间的残基，包括含有酸性谷氨酰胺重复序列的高酸性区域。模仿酸性生长条件的九个pacC +/-突变在299和505 C末端残基之间去除。整个pacC编码区的缺失模仿酸度，但还会导致生长和分生孢子变差。PacC融合蛋白与核心共识GCCARG结合DNA。在碱性环境pH下，PacC激活碱性表达基因 (包括pacC本身) 的转录并抑制酸表达基因的转录。pacCc突变消除了对pH信号转导的需要。

BACKGROUND & AIMS: :Gene expression in fungi by ambient pH is regulated via a conserved signalling cascade whose terminal component is the zinc finger transcription factor PacC/Rim1p. We have identified a pacC orthologue in the vascular wilt pathogen Fusarium oxysporum that binds the consensus 5'-GCCAAG-3' sequence and is proteolytically processed in a similar way to PacC from Aspergillus nidulans. pacC transcript levels were elevated in F. oxysporum grown in alkaline conditions and almost undetectable at extreme acidic growth conditions. PacC+/- loss-of-function mutants displayed an acidity-mimicking phenotype resulting in poor growth at alkaline pH, increased acid protease activity and higher transcript levels of acid-expressed polygalacturonase genes. Reintroduction of a functional pacC copy into a pacC+/- mutant restored the wild-type phenotype. Conversely, F. oxysporum merodiploids carrying a dominant activating pacCc allele had increased pacC transcript and protein levels and displayed an alkalinity-mimicking phenotype with reduced acid phosphatase and increased alkaline protease activities. PacC+/- mutants were more virulent than the wild-type strain in root infection assays with tomato plants, whereas pacCc strains were significantly reduced in virulence. We propose that F. oxysporum PacC acts as a negative regulator of virulence to plants, possibly by preventing transcription of acid-expressed genes important for infection.

背景与目标： : 环境pH在真菌中的基因表达通过保守的信号级联调控，其末端成分是锌指转录因子PacC/Rim1p。我们已经在血管枯萎病原体镰刀菌中鉴定了一个pacC直系同源物，该同源物结合共有的5 '-GCCAAG-3' 序列，并以与构巢曲霉的PacC相似的方式进行蛋白水解处理。在碱性条件下生长的F. oxysporum中，pacC转录水平升高，在极端酸性生长条件下几乎无法检测到。PacC/功能丧失突变体显示出模拟酸度的表型，导致在碱性pH下生长不良，酸性蛋白酶活性增加和酸表达的多聚半乳糖醛酸酶基因的转录水平较高。将功能性pacC拷贝重新引入pacC/-突变体中可以恢复野生型表型。相反，携带显性激活的pacCc等位基因的F. oxysporum merodiploids具有增加的pacC转录本和蛋白质水平，并显示出碱度模拟表型，酸性磷酸酶降低，碱性蛋白酶活性增加。在番茄植物的根系感染试验中，PacC/-突变体比野生型菌株更具毒性，而pacCc菌株的毒力显着降低。我们建议，F. oxysporum PacC可能通过阻止对感染重要的酸表达基因的转录来充当植物毒力的负调节剂。

BACKGROUND & AIMS: BACKGROUND:Non-high-density lipoprotein (HDL) cholesterol is recommended as a secondary lipid goal treated initially with lifestyle modification. However, the relationship between non-HDL and subclinical atherosclerosis is unknown. We examined the independent relationships between coronary artery calcium (CAC), lipids including non-HDL, exercise, and diet among healthy male participants of the Prospective Army Coronary Calcium (PACC) Project. METHODS:Male participants from the PACC Project (n = 1637, mean age 42.8 years; no history of coronary heart disease) were studied. We used validated surveys to measure dietary quality and habitual physical exercise. Fasting lipid concentrations and other cardiovascular risk variables were measured. Subclinical atherosclerosis was detected with the use of electron beam computed tomography for CAC. Factors independently associated with the presence of any detectable CAC (CAC score > 0), including standard CV risk variables, non-HDL, exercise, and diet, were evaluated with the use of logistic regression. RESULTS:The mean Framingham risk score was 4.6 ± 2.6%; CAC was present in 22.4%. Fasting lipid concentrations showed mean LDL-C 128 ± 32 mg/dL, HDL-C 50 ± 13 mg/dL, TG-C 130 ± 86 mg/dL, and non-HDL-C 154 ± 37 mg/dL. Men with CAC had significantly greater levels of LDL-C (135 vs 127 mg/dL), TG (148 vs 124 mg/dL), and non-HDL-C (164 vs 151 mg/dL) and less habitual physical activity (P = 0.006). There were nonsignificant trends between prevalent CAC, greater amounts of dietary fat intake, and lower HDL-C. In successive multivariable logistic regression models for the dependent variable CAC, only non-HDL-C (odds ratio [OR] 1.012 per mg/dL; 95% CI 1.002-1.023; P = .019) and age (OR 1.119 per year; 95% CI 1.063-1.178; P < .001) were independently associated with the presence of CAC, and exercise (OR 0.808; 95% CI 0.703-0.928; P = 0.003) was associated with the absence of CAC. CONCLUSIONS:Non-HDL-C and exercise are independently predictive of the presence of subclinical CAC among healthy lower-risk middle-aged men.

背景与目标：

BACKGROUND & AIMS: :In fungi, ambient pH sensing involves the activation of the Pal/PacC signalling pathway. In the dermatophyte Trichophyton rubrum, pH-dependent secretion of keratinases, which are major virulence determinants, is affected by disruption of the pacC gene. Here, the transcription profiling of the genes coding for N- and O-linked mannosyltransferases, enzymes involved in protein glycosylation, was evaluated in T. rubrum in response to disruption of the pacC gene and growth in keratin, glucose, and glucose plus glycine. We show that transcription of these mannosyltransferase genes is affected by nutrients at acidic pH and by PacC.

背景与目标： : 在真菌中，环境pH传感涉及Pal/PacC信号通路的激活。在皮肤癣菌毛癣菌中，角蛋白酶的pH依赖性分泌是主要的毒力决定因素，受pacC基因破坏的影响。在这里，响应于pacC基因的破坏以及角蛋白，葡萄糖和葡萄糖加甘氨酸的生长，在红T中评估了编码N和O连接的甘露糖基转移酶 (参与蛋白质糖基化的酶) 的基因的转录谱。我们表明，这些甘露糖基转移酶基因的转录受酸性pH下的营养物质和PacC的影响。

BACKGROUND & AIMS: :In this communication, we show that the pacC(c)14 mutation drastically reduced the mannose and N-acetylglycosamine content of the pacA-encoded acid phosphatase secreted by the fungus Aspergillus nidulans when grown at 22 degrees C, pH 5.0, compared to a control strain. The staining after PAGE was not observed for the pacA-encoded acid phosphatase, while the palD-encoded Pi-repressible alkaline phosphatase had an altered electrophoretic mobility. In addition, the secreted acid phosphatase also had a reduced number of isoforms visualized by staining after IEF and glycosylation had a protective effect against its heat inactivation. We also show that a full-length version of gene pacC-1 cloned from Neurospora crassa complemented the pacC(c)14 mutation of A. nidulans, including the remediation of both the acid and alkaline Pi-repressible phosphatases secreted at pH 5.0, which indicates that glycosylation of secreted phosphatases is mediated in A. nidulans by the conserved PacC pathway that governs pH-responsive gene expression.

背景与目标： : 在本通讯中，我们表明，与对照菌株相比，当在22 ℃，pH 5.0下生长时，pacC(c)14突变大大降低了由真菌构巢曲霉分泌的pacA编码酸性磷酸酶的甘露糖和N-乙酰基糖胺含量。PAGE后未观察到pacA编码的酸性磷酸酶的染色，而palD编码的Pi阻遏碱性磷酸酶的电泳迁移率改变。此外，分泌的酸性磷酸酶在IEF和糖基化对其热失活具有保护作用后，通过染色可见的同工型数量也减少了。我们还显示，从crassa Neurospora克隆的基因pacC-1的全长版本补充了a.nidulans的pacC(c)14突变，包括在pH 5.0下分泌的酸和碱性Pi抑制磷酸酶的修复，这表明分泌的磷酸酶的糖基化是由控制pH响应基因表达的保守的PacC途径介导的。

BACKGROUND & AIMS: :Colletotrichum gloeosporioides alkalinizes its surroundings during colonization of host tissue. The transcription factor pacC is a regulator of pH-controlled genes and is essential for successful colonization. We present here the sequence assembly of the Colletotrichum fruit pathogen and use it to explore the global regulation of pathogenicity by ambient pH. The assembled genome size was 54 Mb, encoding 18,456 genes. Transcriptomes of the wild type and ΔpacC mutant were established by RNA-seq and explored for their global pH-dependent gene regulation. The analysis showed that pacC upregulates 478 genes and downregulates 483 genes, comprising 5% of the fungal genome, including transporters, antioxidants, and cell-wall-degrading enzymes. Interestingly, gene families with similar functionality are both up- and downregulated by pacC. Global analysis of secreted genes showed significant pacC activation of degradative enzymes at alkaline pH and during fruit infection. Select genes from alkalizing-type pathogen C. gloeosporioides and from acidifying-type pathogen Sclerotinia sclerotiorum were verified by quantitative reverse-transcription polymerase chain reaction analysis at different pH values. Knock out of several pacC-activated genes confirmed their involvement in pathogenic colonization of alkalinized surroundings. The results suggest a global regulation by pacC of key pathogenicity genes during pH change in alkalinizing and acidifying pathogens.

背景与目标： : 在宿主组织定植过程中，炭疽菌将其周围环境碱化。转录因子pacC是pH控制基因的调节剂，对于成功定植至关重要。我们在这里介绍了炭疽菌果实病原体的序列组装，并使用它来探索环境pH对致病性的整体调节。组装的基因组大小为54 Mb，编码18,456个基因。通过RNA-seq建立了野生型和 Δ pacc突变体的转录组，并探索了其整体pH依赖性基因调控。分析表明，pacC上调478基因并下调483基因，包括真菌基因组的5%，包括转运蛋白，抗氧化剂和细胞壁降解酶。有趣的是，具有相似功能的基因家族都被pacC上调和下调。分泌基因的整体分析显示，在碱性pH下和水果感染期间，降解酶的pacC活化显着。通过定量逆转录聚合酶链反应分析在不同ph值下验证了碱化型病原体C. gloeosporioides和酸化型病原体菌核盘菌的选择基因。敲除几个pacC激活的基因证实了它们参与了碱化环境的致病性定植。结果表明，在碱化和酸化病原体的pH变化过程中，pacC对关键致病性基因进行了整体调节。

BACKGROUND & AIMS: :In order to survive and adapt to the environment, it is imperative for fungi to be able to sense and respond to changes in extracellular pH conditions. In ascomycetes, sensing of extracellular pH is mediated by the Pal pathway resulting in activation of the PacC transcription factor at alkaline pH. The role of PacC in regulating fungal virulence and pathogenicity has been described in several pathogenic fungi but to date not in a symbiotic fungus. Epichloë festucae is a biotrophic fungal endophyte that forms a stable mutualistic interaction with Lolium perenne. In this study, pacC deletion (ΔpacC) and dominant active (pacC(C)) mutants were generated in order to study the cellular roles of PacC in E. festucae. Deletion of pacC resulted in increased sensitivity of the mutant to salt-stress but surprisingly did not affect the ability of the mutant to grow under alkaline pH conditions. Alkaline pH was observed to induce conidiation in wild-type E. festucae but not in the ΔpacC mutant. On the other hand the pacC(C) mutant had increased conidiation at neutral pH alone. Null pacC mutants had no effect on the symbiotic interaction with ryegrass plants whereas the pacC(C) mutant increased the tiller number. Examination of the growth of the pacC(C) mutant in the plant revealed the formation of aberrant convoluted hyphal structures and an increase in hyphal breakage, which are possible reasons for the altered host interaction phenotype.

背景与目标： : 为了生存并适应环境，真菌必须能够感知并响应细胞外pH条件的变化。在子囊菌中，细胞外pH的感测是由Pal途径介导的，从而导致碱性pH下PacC转录因子的激活。在几种致病真菌中已经描述了PacC在调节真菌毒力和致病性中的作用，但迄今为止在共生真菌中尚未描述。Epichlo ë festucae是一种生物营养真菌内生菌，与黑麦草形成稳定的相互作用。在这项研究中，产生了pacC缺失 (Δ pacC) 和显性活性 (PacC (C)) 突变体，以研究pacC在festucae中的细胞作用。pacC的缺失导致突变体对盐胁迫的敏感性增加，但令人惊讶的是，它不影响突变体在碱性pH条件下生长的能力。观察到碱性pH在野生型E. festucae中诱导分生孢子，但在 Δ pacc突变体中不诱导分生孢子。另一方面，仅在中性pH下，pacC(C) 突变体的分生孢子增加。无效的pacC突变体对与黑麦草植物的共生相互作用没有影响，而pacC(C) 突变体增加了分till数。对植物中pacC(C) 突变体的生长进行的检查表明，形成了异常的回旋菌丝结构，菌丝断裂增加，这可能是宿主相互作用表型改变的原因。

BACKGROUND & AIMS: :An Aspergillus niger strain has been constructed in which the pH-dependent regulatory gene, pacC, was disrupted. The pacC gene of A. niger, like that of A. nidulans, is involved in the regulation of acid phosphatase expression. Disruptants were identified by a reduction in acid phosphatase staining of colonies. Southern analysis demonstrated integration of the disruption plasmid at the pacC locus and Northern analysis showed that the disruption strain produced a truncated pacC mRNA of 2.2 kb (as compared to 2.8 kb in the wild type). The strain carrying the pacC disruption was used to assign the pacC gene to linkage group IV; this was confirmed by CHEF electrophoresis and Southern analysis. This strain further allowed us to determine which extracellular enzyme and transport systems are under the control of pacC in A. niger. Expression of the A. niger pacC wild-type gene and the truncated pacC gene showed that, in contrast to the auto-regulated wild-type expression, which was elevated only at alkaline pH, the truncated pacC gene was deregulated, as high-level expression occurred regardless of the pH of the culture medium. Analysis of the phosphatase spectrum by isoelectric focussing and enzyme activity staining both in the wild-type and the pacC disruptant showed that at least three acid phosphatases are regulated by the pacC. For the single alkaline phosphatase no pH regulation was observed.

背景与目标： : 已经构建了黑曲霉菌株，其中pH依赖性调节基因pacC被破坏。与A.Niddulans一样，黑曲霉的pacC基因参与酸性磷酸酶表达的调节。通过减少菌落的酸性磷酸酶染色来鉴定破坏物。Southern分析证明了破坏质粒在pacC基因座上的整合，Northern分析表明，破坏菌株产生了2.2 kb的截短的pacC mRNA (与野生型中的2.8 kb相比)。携带pacC破坏的菌株用于将pacC基因分配给IV连锁组; 这已通过CHEF电泳和Southern分析得到证实。该菌株进一步使我们能够确定黑曲霉中哪些胞外酶和转运系统处于pacC的控制之下。A. niger pacC野生型基因和截短的pacC基因的表达表明，与仅在碱性pH下升高的自动调节野生型表达相反，截短的pacC基因被解除管制，无论培养基的ph值如何，都会发生高水平的表达。通过等电聚焦和酶活性染色对野生型和pacC破坏物中的磷酸酶谱进行分析表明，至少三种酸性磷酸酶受pacC调节。对于单一碱性磷酸酶，未观察到pH调节。

BACKGROUND & AIMS: INTRODUCTION:Intrathecal therapy is an important part of the pain treatment algorithm for chronic disease states. The use of this option is a viable treatment strategy, but it is inherent for pain physicians to understand risk assessment and mitigation. In this manuscript, we explore evidence and mitigating strategies to improve safety with intrathecal therapy. METHODS:A robust literature search was performed covering January 2011 to October 9, 2016, in PubMed, Embase, MEDLINE, Biomed Central, Google Scholar, Current Contents Connect, and International Pharmaceutical Abstracts. The information was cross-referenced and compiled for evidence, analysis, and consensus review, with the intent to offer weighted recommendations and consensus statements on safety for targeted intrathecal therapy delivery. RESULTS:The Polyanalgesic Consensus Conference has made several best practice recommendations to improve care and reduce morbidity and mortality associated with intrathecal therapy through all phases of management. The United States Prevention Service Task Force evidence level and consensus strength assessments are offered for each recommendation. CONCLUSION:Intrathecal therapy is a viable and relatively safe option for the treatment of cancer- and noncancer-related pain. Continued research and expert opinion are required to improve our current pharmacokinetic and pharmacodynamic model of intrathecal drug delivery, as this will undoubtedly improve safety and efficacy.

背景与目标：

BACKGROUND & AIMS: :Destruction of the pulmonary epithelium is a major feature of lung diseases caused by the mould pathogen Aspergillus fumigatus. Although it is widely postulated that tissue invasion is governed by fungal proteases, A. fumigatus mutants lacking individual or multiple enzymes remain fully invasive, suggesting a concomitant requirement for other pathogenic activities during host invasion. In this study we discovered, and exploited, a novel, tissue non-invasive, phenotype in A. fumigatus mutants lacking the pH-responsive transcription factor PacC. Our study revealed a novel mode of epithelial entry, occurring in a cell wall-dependent manner prior to protease production, and via the Dectin-1 β-glucan receptor. ΔpacC mutants are defective in both contact-mediated epithelial entry and protease expression, and significantly attenuated for pathogenicity in leukopenic mice. We combined murine infection modelling, in vivo transcriptomics, and in vitro infections of human alveolar epithelia, to delineate two major, and sequentially acting, PacC-dependent processes impacting epithelial integrity in vitro and tissue invasion in the whole animal. We demonstrate that A. fumigatus spores and germlings are internalised by epithelial cells in a contact-, actin-, cell wall- and Dectin-1 dependent manner and ΔpacC mutants, which aberrantly remodel the cell wall during germinative growth, are unable to gain entry into epithelial cells, both in vitro and in vivo. We further show that PacC acts as a global transcriptional regulator of secreted molecules during growth in the leukopenic mammalian lung, and profile the full cohort of secreted gene products expressed during invasive infection. Our study reveals a combinatorial mode of tissue entry dependent upon sequential, and mechanistically distinct, perturbations of the pulmonary epithelium and demonstrates, for the first time a protective role for Dectin-1 blockade in epithelial defences. Infecting ΔpacC mutants are hypersensitive to cell wall-active antifungal agents highlighting the value of PacC signalling as a target for antifungal therapy.

背景与目标： : 肺上皮的破坏是由霉菌病原体烟曲霉引起的肺部疾病的主要特征。尽管人们普遍认为组织入侵是由真菌蛋白酶控制的，但缺乏单个或多种酶的烟曲霉突变体仍然具有完全的侵入性，这表明在宿主入侵过程中还需要其他致病活动。在这项研究中，我们在缺乏pH响应转录因子PacC的烟曲霉突变体中发现并开发了一种新颖的组织非侵入性表型。我们的研究揭示了一种新的上皮进入模式，在蛋白酶产生之前通过Dectin-1的 β-葡聚糖受体以细胞壁依赖性方式发生。Δ pacc突变体在接触介导的上皮进入和蛋白酶表达中均有缺陷，并且在白细胞减少症小鼠中的致病性显着减弱。我们结合了鼠感染模型，体内转录组学和人肺泡上皮的体外感染，以描述两个主要且依次作用的依赖于PacC的过程，这些过程影响整个动物的上皮完整性和组织侵袭。我们证明烟曲霉孢子和发芽被上皮细胞以接触，肌动蛋白，细胞壁和Dectin-1依赖性方式内在化，并且 Δ pacc突变体在发芽生长过程中异常重塑细胞壁，无法进入上皮细胞，无论是在体外还是体内。我们进一步表明，PacC在白细胞减少的哺乳动物肺的生长过程中充当分泌分子的整体转录调节剂，并描述了侵袭性感染期间表达的分泌基因产物的完整队列。我们的研究揭示了一种组合的组织进入模式，该模式取决于肺上皮的顺序和机械上不同的扰动，并首次证明了Dectin-1阻断在上皮防御中的保护作用。感染 Δ PacC突变体对细胞壁活性抗真菌剂过敏，突出了PacC信号传导作为抗真菌治疗靶标的价值。

BACKGROUND & AIMS: :The Aspergillus PacC transcription factor undergoes proteolytic activation in response to alkaline ambient pH. In acidic environments, the 674 residue translation product adopts a 'closed' conformation, protected from activation through intramolecular interactions involving the < or = 150 residue C-terminal domain. pH signalling converts PacC to an accessible conformation enabling processing cleavage within residues 252--254. We demonstrate that activation of PacC requires two sequential proteolytic steps. First, the 'closed' translation product is converted to an accessible, committed intermediate by proteolytic elimination of the C-terminus. This ambient pH-regulated cleavage is required for the final, pH-independent processing reaction and is mediated by a distinct signalling protease (possibly PalB). The signalling protease cleaves PacC between residues 493 and 500, within a conserved 24 residue 'signalling protease box'. Precise deletion or Leu498Ser substitution prevents formation of the committed and processed forms, demonstrating that signalling cleavage is essential for final processing. In contrast, signalling cleavage is not required for processing of the Leu340Ser protein, which lacks interactions preventing processing. In its two-step mechanism, PacC processing can be compared with regulated intramembrane proteolysis.

背景与目标： : 曲霉PacC转录因子在碱性环境pH下发生蛋白水解激活。在酸性环境中，674残基翻译产物采用 “封闭” 构象，通过涉及 <或 = 150残基C-末端结构域的分子内相互作用保护免于活化。pH信号传导将PacC转化为能够在残基252-254内处理裂解的可及构象。我们证明了PacC的激活需要两个连续的蛋白水解步骤。首先，通过蛋白水解消除C末端，将 “封闭” 的翻译产物转化为可访问的，承诺的中间体。这种环境pH调节的裂解是最终的，与pH无关的加工反应所必需的，并且由不同的信号蛋白酶 (可能是PalB) 介导。信号蛋白酶在保守的24个残基 “信号蛋白酶盒” 内切割残基493和500之间的PacC。精确的删除或Leu498Ser替换可防止提交和处理的形式的形成，这表明信号裂解对于最终加工至关重要。相反，Leu340Ser蛋白的加工不需要信号裂解，因为它缺乏阻止加工的相互作用。在其两步机制中，可以将PacC处理与受调控的膜内蛋白水解进行比较。

BACKGROUND & AIMS: :A homologue of the gene encoding the transcription factor Rim101 (PacC), involved in pH signal transduction in fungi, was identified in the pathogenic basidiomycete Ustilago maydis. The gene (RIM101) encodes a protein of 827 amino acid residues, which shows highest similarity to PacC proteins from Fusarium oxysporum and Aspergillus niger. The gene had the capacity to restore protease activity to rim101 mutants from Yarrowia lipolytica, confirming its homologous function, and was expressed at both acid and neutral pH. Null Deltarim101 mutants were not affected in the in vitro pH-induced dimorphic transition, their growth rate, resistance to hypertonic sorbitol or KCl stress, and pathogenicity. However, similar to pacC (rim101) mutants in other fungi, they displayed a pleiotropic phenotype with alterations in morphogenesis, impairment in protease secretion, and increased sensitivity to Na+ and Li+ ions. Other phenotypic characteristics not previously reported in fungal pacC (rim101) mutants (morphological changes, increased sensitivity to lytic enzymes, and augmented polysaccharide secretion) were also observed in U. maydis mutants. All these modifications were alleviated by transformation with the wild-type gene, confirming that all were the result of mutation in RIM101. These data indicate that the Pal/Rim pathway is functional in U. maydis (and probably in other basidiomycetes) and plays complex roles in pH-sensing phenomena, as occurs in ascomycetes and deuteromycetes.

背景与目标： : 在致病性担子菌Ustilago maydis中鉴定了编码转录因子Rim101 (PacC) 的基因的同源物，该基因参与真菌中的pH信号转导。该基因 (RIM101) 编码827个氨基酸残基的蛋白质，该蛋白质与尖孢镰刀菌和黑曲霉的PacC蛋白具有最高的相似性。该基因具有恢复解脂耶氏酵母rim101突变体的蛋白酶活性的能力，证实了其同源功能，并在酸性和中性pH下表达。无效的Deltarim101突变体在体外pH诱导的二态转变，其生长速率，对高渗山梨醇或KCl胁迫的抗性以及致病性均不受影响。然而，与其他真菌中的pacC (rim101) 突变体相似，它们显示出多效性表型，其形态发生改变，蛋白酶分泌受损以及对Na和Li离子的敏感性增加。在U. maydis突变体中还观察到以前未在真菌pacC (rim101) 突变体中报道的其他表型特征 (形态变化，对裂解酶的敏感性增加和多糖分泌增加)。所有这些修饰都通过与野生型基因的转化而得到缓解，证实所有这些修饰都是rim101突变的结果。这些数据表明Pal/Rim途径在U. maydis (可能在其他担子菌中) 中起作用，并且在pH感应现象中起着复杂的作用，例如在子囊菌和半知菌中。

BACKGROUND & AIMS: :Penicillium expansum, the causal agent of blue mould rot, causes severe post-harvest fruit maceration simultaneously with the secretion of d-gluconic acid (GLA) and the mycotoxin patulin in colonized tissue. The factor(s) inducing patulin biosynthesis during colonization of the host acidic environment is unclear. During the colonization of apple fruit in vivo and growth in culture, P. expansum secretes pH-modulating GLA and ammonia. Although patulin and its possible opportunistic precursor GLA accumulate together during fungal development, ammonia is detected on the colonized tissue's leading edge and after extended culture, close to patulin accumulation. Here, we demonstrate ammonia-induced transcript activation of the global pH modulator PacC and patulin accumulation in the presence of GLA by: (i) direct exogenous treatment of P. expansum growing on solid medium; (ii) direct exogenous treatment on colonized apple tissue; (iii) growth under self-ammonia production conditions with limited carbon; and (iv) analysis of the transcriptional response to ammonia of the patulin biosynthesis cluster. Ammonia induced patulin accumulation concurrently with the transcript activation of pacC and patulin biosynthesis cluster genes, indicating the regulatory effect of ammonia on pacC transcript expression under acidic conditions. Electrophoretic mobility shift assays using P. expansum PacC and antibodies to the different cleaved proteins showed that PacC is not protected against proteolytic signalling at pH 4.5 relative to pH 7.0, but NH4 addition did not further enhance its proteolytic cleavage. Ammonia enhanced the activation of palF transcript in the Pal pathway under acidic conditions. Ammonia accumulation in the host environment by the pathogen under acidic pH may be a regulatory cue for pacC activation, towards the accumulation of secondary metabolites, such as patulin.

背景与目标： : 青霉青霉是蓝霉腐烂的病原体，在定植组织中分泌d-葡萄糖酸 (GLA) 和霉菌毒素棒曲霉素的同时，引起了严重的收获后浸渍。在宿主酸性环境定植过程中诱导棒曲霉素生物合成的因素尚不清楚。在苹果果实体内定植和培养过程中，P. expansum分泌调节pH的GLA和氨。尽管棒曲霉素及其可能的机会性前体GLA在真菌发育过程中积聚在一起，但在定植组织的前缘和长时间培养后检测到氨，接近棒曲霉素的积累。在这里，我们通过以下方式证明了氨诱导的转录激活全球pH调节剂PacC和棒曲霉素在GLA存在下的积累 :( i) 在固体培养基上生长的P. expansum的直接外源处理; (ii) 在定植的苹果组织上的直接外源处理; (iii) 在碳有限的自氨生产条件下生长; (iv) 分析棒曲霉素生物合成簇对氨的转录反应。氨诱导棒曲霉素的积累与pacC和棒曲霉素生物合成簇基因的转录激活同时发生，表明氨在酸性条件下对pacC转录本表达的调节作用。使用P. Expansum  pacc和针对不同裂解蛋白的抗体的电泳迁移率位移测定表明，相对于pH 7.0，在pH 4.5时，PacC不能防止蛋白水解信号传导，但NH4的添加不能进一步增强其蛋白水解切割。在酸性条件下，氨增强了Pal途径中palF转录本的活化。在酸性pH下，病原体在宿主环境中积累的氨可能是pacC活化的调节线索，从而导致次级代谢产物 (例如棒曲霉素) 的积累。

BACKGROUND & AIMS: :Cellulosic biomass represents a valuable potential substitute for fossil-based fuels. As such, there is a strong need to develop efficient biotechnological processes for the enzymatic hydrolysis of cellulosic biomass via the optimization of cellulase production by fungi. Ambient pH is an important factor affecting the industrial production of cellulase. In the present study, we demonstrate that several Aspergillus nidulans genes encoding cellulolytic enzymes are regulated by Pal-PacC-mediated pH signaling, as evidenced by the decreased cellulase productivity of the palC mutant and pacC deletants of A. nidulans. The deletion of pacC was observed to result in delayed induction and decreased expression of the cellulase genes based on time course expression analysis. The genome-wide identification of PacC-regulated genes under cellobiose-induced conditions demonstrated that genes expressed in a PacC-dependent manner included 82 % of ClrB (a transcriptional activator of the cellulase genes)-regulated genes, including orthologs of various transporter and β-glucosidase genes considered to be involved in cellobiose uptake or production of stronger inducer molecules. Together with the significant overlap between ClrB- and PacC-regulated genes, the results suggest that PacC-mediated regulation of the cellulase genes involves not only direct regulation by binding to their promoter regions but also indirect regulation via modulation of the expression of genes involved in ClrB-dependent transcriptional activation. Our findings are expected to contribute to the development of more efficient industrial cellulase production methods.

背景与目标： : 纤维素生物质是化石燃料的有价值的潜在替代品。因此，迫切需要通过优化真菌产生的纤维素酶来开发有效的生物技术方法，以酶促水解纤维素生物质。环境ph值是影响纤维素酶工业生产的重要因素。在本研究中，我们证明了编码纤维素分解酶的几个构巢曲霉基因受Pal-PacC介导的pH信号传导的调节，这可以通过下降的纤维素酶生产率证明。根据时程表达分析，观察到pcc的缺失导致纤维素酶基因的诱导延迟和表达降低。在纤维二糖诱导的条件下对PacC调节基因的全基因组鉴定表明，以PacC依赖性方式表达的基因包括ClrB (纤维素酶基因的转录激活因子) 调节基因的82%，包括被认为参与纤维二糖摄取或产生更强诱导剂分子的各种转运蛋白和 β-葡萄糖苷酶基因的直系同源物。再加上ClrB和PacC调节的基因之间的显着重叠，结果表明，PacC介导的纤维素酶基因的调控不仅涉及通过结合其启动子区域而直接调控，而且还涉及通过调节涉及的基因表达的间接调控ClrB依赖性转录激活。我们的发现有望有助于开发更有效的工业纤维素酶生产方法。

BACKGROUND & AIMS: :The transcription factor PacC/Rim101 participates in environmental pH adaptation, development and secondary metabolism in many fungi, but whether PacC/Rim101 contributes to fungal adaptation to environmental stress remains unclear. In our previous study, a homologous gene of PacC/Rim101 was identified, and PacC-silenced strains of the agaricomycete Ganoderma lucidum were constructed. In this study, we further investigated the functions of PacC in G. lucidum and found that PacC-silenced strains were hypersensitive to environmental stresses, such as osmotic stress, oxidative stress and cell wall stress, compared with wild-type (WT) and empty-vector control (CK) strains. In addition, transmission electron microscopy images of the cell wall structure showed that the cell walls of the PacC-silenced strains were thinner (by approximately 25-30%) than those of the WT and CK strains. Further analysis of cell wall composition showed that the β-1,3-glucan content in the PacC-silenced strains was only approximately 78-80% of that in the WT strain, and the changes in β-1,3-glucan content were consistent with downregulation of glucan synthase gene expression. The ability of PacC to bind to the promoters of glucan synthase-encoding genes confirms that PacC transcriptionally regulates these genes.

背景与目标： 转录因子PacC/Rim101参与了许多真菌的环境pH适应，发育和次生代谢，但PacC/Rim101是否有助于真菌对环境胁迫的适应仍不清楚。在我们先前的研究中，鉴定了PacC/Rim101的同源基因，并构建了伞菌灵芝的PacC沉默菌株。在这项研究中，我们进一步研究了灵芝中PacC的功能，发现与野生型 (WT) 和空载体控制 (CK) 菌株相比，PacC沉默菌株对环境胁迫 (如渗透应激，氧化应激和细胞壁应激) 高度敏感。此外，细胞壁结构的透射电子显微镜图像显示，经PacC沉默的菌株的细胞壁比WT和CK菌株的细胞壁薄 (约25-30%)。细胞壁组成的进一步分析表明，PacC沉默菌株中的 β-1，3-葡聚糖含量仅为WT菌株的约78-80%，并且 β-1，3-葡聚糖含量的变化与葡聚糖合酶基因表达的下调一致。PacC与葡聚糖合酶编码基因启动子结合的能力证实了PacC在转录上调节这些基因。