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.

过渡金属包括Cu、Mn、Zn和Fe，是所有生命形式的必需营养素。微生物病原体 在获取这些重要元素方面面临挑战，因为宿主故意试图使微生物缺乏它们的营养。 金属通过统称为营养免疫的过程。成功的病原体已经进化出 精心设计的方法，以逃避营养免疫和捕获金属从他们的主机;一个机制， 分泌“金属载体”。金属载体是小分子（≤1.5 kDa），可与细胞外金属结合， 高亲和力并将离子传递给微生物以供吸收。细菌分泌各种金属载体的铁，铜或 锌，虽然没有被确定为锰。致病真菌被认为只分泌铁金属载体 而且只存在于某些物种中。白色念珠菌是一种机会性真菌病原体， 酵母是任何种类的金属载体，尽管酵母的致病机理依赖于宿主金属。最近，我们的实验室 挑战了假丝酵母菌无金属载体的教条，并获得了假丝酵母菌有金属载体的证据。白色念珠菌在 事实上，产生对结合Mn 2+或Cu 2+表现出强选择性的金属团样化合物。我们称 这些Mn-MBC和Cu-MBC为金属结合络合物。Mn-MBC和Cu-MBC的金属配位 不同，因为它们的色谱特性，表明Mn-MBC和Cu-MBC是不同的 分子。有趣的是，Cu-MBC的产生，而不是Mn-MBC，是由Fe诱导的，表明Cu-MBC的作用。 铁稳态中的MBC。目前我们还不知道MBC分子的化学性质， 在不同真菌中的存在，或它们在真菌致病的金属代谢中的作用。在这两个- 一年的研究计划，我们将测试MBC生产从真菌以外的假丝酵母属;我们将化学 识别C.白念珠菌Mn-MBC和Cu-MBC，并将开始探索其作用机制。目标1： 调查不同真菌的MBC，并从化学上鉴定C.白色念珠菌使用尺寸 排阻色谱/ICP-质谱/ENDOR光谱法，我们已经开发， 鉴定Mn-和Cu-MBC，我们将测试两种非念珠菌属真菌病原体中MBC生产。Mn-MBC和 Cu-MBC来源于C.将通过质谱法对白色念珠菌进行化学鉴定并确定其组成。 实验将吸引ENDOR光谱学和质谱学方面的专家合作者。目标2： 深入了解MBC的基础生物学。我们的初步研究表明，Mn-和Cu- MBC可以作为C.白色念珠菌培养。我们将测试 使用MBC的金属摄取途径，并检查MBC金属递送的金属酶靶标。 如果时间允许，我们将使用适当的C.白色 突变体和念珠菌病的小鼠模型，其中真菌病原体面临金属饥饿应激。总的来说， 这个为期2年的项目的成功完成将揭示真菌王国的第一个非铁金属载体 以及真菌捕获致病所必需的金属营养素的新机制。