Metal nutrients and metallophore-like molecules for a fungal pathogen

真菌病原体的金属营养物和类金属载体分子

• 批准号: 10231544
• 负责人: Valeria C Culotta
• 金额: $ 24.56万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-01 至 2023-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10231544
• 关键词: Affinity; Anabolism; Animals; Bacteria; Binding; Biochemical; Biological Assay; Biology; Candida; Candida albicans; Candidiasis; Cells; Chemicals; Chemistry; Collaborations; Complex; Electron Nuclear Double Resonance; Elements; Environment; Exhibits; Face; Generations; Growth; Homeostasis; Human; Infection; Innate Immune Response; Invaded; Ions; Life; Mass Spectrum Analysis; Metals; Methods; Microbe; Micronutrients; Molecular Sieve Chromatography; Molecular Weight; Nature; Nutrient; Nutritional Immunity; Organism; Pathogenesis; Pathway interactions; Process; Production; Property; Public Health; Research; Role; Siderophores; Site; Spectrum Analysis; Starvation; Stress; Surveys; Testing; Time; Transition Elements; Virulence; Washington; Yeasts; experimental study; extracellular; fungus; insight; metal metabolism; metalloenzyme; mouse model; mutant; pathogen; pathogenic fungus; pathogenic microbe; programs; success; uptake; virtual

Transition metals including Cu, Mn, Zn and Fe are essential nutrients for all life forms. Microbial pathogens face challenges in acquiring these vital elements, as the host deliberately attempts to starve microbes of their metals through processes collectively known as nutritional immunity. Successful pathogens have evolved elaborate methods to evade nutritional immunity and capture metals from their host; one mechanism involves secretion of ‘metallophores”. Metallophores are small (≤1.5 kDa) molecules that bind extracellular metals with high affinity and deliver the ion to the microbe for uptake. Bacteria secrete various metallophores for Fe, Cu or Zn, although none have been identified for Mn. Pathogenic fungi are thought to only secrete Fe-metallophores and only in certain species. Candida albicans is an opportunistic fungal pathogen believed to not produce metallophores of any kind, although the yeast is dependent on host metals for pathogenesis. Recently, our lab has challenged the dogma of no metallophores for Candida sp and obtained evidence that C. albicans does in fact produce metallophore-like compounds that exhibit strong selectivity for binding Mn2+ or Cu2+. We call these Mn-MBC and Cu-MBC for metal binding complex. The metal coordination sites of Mn-MBC and Cu-MBC are distinct, as are their chromatographic properties, indicating that Mn-MBC and Cu-MBC are different molecules. Interestingly, production of Cu-MBC, but not Mn-MBC, is induced by Fe, indicating a role for Cu- MBC in Fe homeostasis. Currently we do not know the chemical nature of the MBC molecules, their occurrence among diverse fungi, or their roles in metabolism of metals for fungal pathogenesis. Over this two- year research plan, we shall test for MBC production from fungi outside of Candida sp; we will chemically identify C. albicans Mn-MBC and Cu-MBC, and will begin to probe their mechanism of action. Aim 1: To survey MBC across diverse fungi and chemically identify the molecules from C. albicans. Using a size exclusion chromatography/ICP-mass spectrometry/ENDOR spectroscopy approach that we have developed to identify Mn- and Cu-MBC, we will test for MBC production in two non-Candida fungal pathogens. Mn-MBC and Cu-MBC from C. albicans will be chemically identified and composition defined by mass spectrometry. Experiments will engage expert collaborators in ENDOR spectroscopy and mass spectrometry. Aim 2: To gain insight into the fundamental biology of MBC. Our preliminary studies show that both Mn- and Cu- MBC can serve as donors of their respective metals for uptake by C. albicans in culture. We shall test the pathway of metal uptake that uses the MBCs and examine metalloenzyme targets for metal delivery by MBC. Time permitting, we shall test the role of Mn- and Cu-MBC in pathogenesis using appropriate C. albicans mutants and a murine model of candidiasis where the fungal pathogen faces metal starvation stress. Overall, successful completion of this 2-year program will reveal the first non-Fe metallophores for the fungal kingdom and new mechanisms by which fungi can capture metal nutrients that are essential for pathogenesis.

包括Cu，Mn，Zn和Fe在内的过渡金属是所有生命形式的必需营养素。微生物病原体 在获得这些重要元素时面临挑战，因为主人故意试图饿死他们的微生物 金属通过共同称为营养免疫的过程。成功的病原体发展了 详细的方法来逃避营养免疫并从其宿主那里捕获金属；一种机制涉及 分泌“金属流团”。金属噬菌体是小（≤1.5kDa）的分子，与细胞外金属结合 高亲和力并将离子输送到微生物以吸收。细菌分泌各种金属球体的Fe，Cu或 Zn，尽管没有针对Mn确定。病原真菌被认为仅分泌Fe-Inallophothores 并且仅在某些物种中。白色念珠菌是一种机会性真菌病原体，据信不产生 尽管酵母取决于宿主金属的发病机理，但任何种类的金属泳道。最近，我们的实验室 已经挑战了无金属泳道的念珠菌的教条，并获得了白色念珠菌在 事实产生了类似金属球的化合物，具有结合MN2+或Cu2+的强选择性。我们打电话 这些用于金属结合复合物的MN-MBC和CU-MBC。 MN-MBC和CU-MBC的金属协调位点 与它们的色谱特性一样不同，表明MN-MBC和CU-MBC不同 分子。有趣的是，Fe诱导Cu-MBC而非MN-MBC的产生，这表明Cu-发挥了作用 MBC在FE稳态中。目前，我们不知道MBC分子的化学性质， 潜水员真菌之间的发生，或它们在金属代谢中的作用，用于真菌发病机理。在这个两个 年度研究计划，我们将测试Candida SP以外的真菌的MBC生产；我们将化学 识别白色念珠菌Mn-MBC和Cu-MBC，并将开始探究其作用机理。目标1：到 调查MBC跨潜水员真菌，并从化学鉴定白色念珠菌的分子。使用尺寸 我们已经开发出的排除色谱/ICP质量光谱法/内托光谱法 识别MN-和CU-MBC，我们将在两种非candida真菌病原体中测试MBC的产生。 Mn-MBC和 来自白色念珠菌的CU-MBC将被化学鉴定，并由质谱法定定义。 实验将吸引专家合作者参与端光谱和质谱法。目标2：到 深入了解MBC的基本生物学。我们的初步研究表明，Mn-和Cu-均可 MBC可以用作其各自金属的捐助者，以吸引白色念珠菌在培养物中的吸收。我们将测试 使用MBC的金属摄取途径并检查MBC的金属酶靶标。 允许的时间，我们应使用适当的白色念珠菌测试Mn-和Cu-MBC在发病机理中的作用 真菌病原体面临金属饥饿应激的突变体和鼠类念珠菌病模型。全面的， 这项为期两年计划的成功完成将揭示真菌王国的第一个非FE金属体 真菌可以捕获发病机理必不可少的金属营养素的新机制。