TB Membrane Transporters and Intrinsic Resistance

结核病膜转运蛋白和固有耐药性

• 批准号: 8868006
• 负责人: Georgiana E. Purdy
• 金额: $ 38.5万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-05 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8868006
• 关键词: ATP-Binding Cassette Transporters; Anti-Bacterial Agents; Antibiotic Resistance; Antibiotic Therapy; Antibiotics; Antimicrobial Cationic Peptides; Antimicrobial Resistance; Bacteria; Binding; Biological Assay; Cathepsins; Cell Membrane Permeability; Cell Wall; Circular Dichroism; Data; Development; Disease; Drug Efflux; Environment; Exposure to; Future; Genes; Genus Mycobacterium; Goals; Human; Immune response; In Vitro; Infection; Knowledge; Lead; Membrane; Membrane Proteins; Membrane Transport Proteins; Metabolic; Molecular Profiling; Mus; Mycobacterium tuberculosis; NMR Spectroscopy; Peptide Antibiotics; Peptide Hydrolases; Peptides; Permeability; Pharmaceutical Preparations; Pharmacotherapy; Play; Predisposition; Property; Public Health; Regulation; Resistance; Rest; Role; Signal Transduction; System; Therapeutic; Time; Transmembrane Transport; Tuberculosis; Ubiquitin; Virulence; Work; antibiotic efflux; antimicrobial; antimicrobial peptide; bactericide; base; differential expression; disorder control; improved; inhibitor/antagonist; killings; macrophage; mouse model; mutant; mycobacterial; novel therapeutic intervention; novel therapeutics; research study; resistance mechanism; response; therapeutic target; tool

DESCRIPTION (provided by applicant): Tuberculosis continues to pose a threat to public health, and resistance to commonly used antibiotics is an increasingly prevalent problem in efforts to control this disease. Thus, there is a great need to develop new drugs and therapeutic strategies. Mycobacteria display intrinsic resistance to antibiotics due to the low permeability of the mycobacterial cell wall and active efflux of antibiotics. An understanding of drug efflux systems role in intrinsic antibiotic resistance and their regulation will lead to new therapeutic approaches to TB since inhibition of these intrinsic systems may improve the efficacy of current drugs and shorten treatment time. Previously we showed that ubiquitin-derived peptides (Ub-peptides) contribute the mycobactericidal repertoire of macrophages (Alonso et al., 2007; Purdy et al., 2009). Our first objective is to better understand the bactericidal actions of these peptides. Initial studies indicate that Ub-peptides behave like antimicrobial peptides to impair mycobacterial membrane function. Mycobacterium tuberculosis (Mtb) has co-evolved with the human immune response and likely possesses resistance mechanisms to sub-lethal concentrations of host antimicrobial compounds. In modern disease, these systems may provide intrinsic resistance to antibiotics. To identify Mtb intrinsic resistance mechanisms, two complementary approaches were taken using mycobactericidal Ub-peptides as a tool. 1.) We directly identified ABC and MFS transporter mutants more susceptible to treatment with Ub-peptides in a transposon mutant screen. 2.) The expression profile of Mtb exposed to Ub-peptides was analyzed. Ten percent of Ub-peptide regulated genes encoded predicted membrane proteins including ATP Binding Cassette (ABC) or Major Facilitator Superfamily (MFS) transport systems. These differentially expressed genes alter the bacterial response to the presence of antimicrobial peptides and antibiotics. To show this we acquired Mtb mutants that lack ABC and MFS transporters and found that they are more susceptible to antimicrobial peptides and antibiotics. Our working hypothesis is that Ub- peptides are host antimicrobial peptides and that Mtb ABC and MFS membrane transport systems contribute to Mtb intrinsic antimicrobial resistance and virulence. The long-term goal of this work will direct therapeutic targeting of these membrane transport systems to increase the inherent sensitivity of Mtb to host antimicrobial compounds and antibiotic treatment. Our aims are the following: 1) We will characterize the mechanism of Ub-peptide action. 2) To better understand intrinsic resistance of Mtb we will define the role of the ABC transporters Pst and Rv0986-Rv0987, and the MFS transporters Rv1634, and Rv3239c in the intrinsic resistance of Mtb. 3) We will determine if these ABC and MFS transporters contribute to M. tuberculosis virulence through macrophage survival assays and mouse infections. These studies will expand our knowledge of M. tuberculosis intrinsic resistance to antimicrobial Ub-peptides and antibiotics, and will identify targets for future drug therapy approaches.

描述（由申请人提供）：结核病继续对公共卫生构成威胁，对常用抗生素的抵抗是控制这种疾病的努力，这是一个日益普遍的问题。因此，非常需要开发新药和治疗策略。分枝杆菌表现出对抗生素的内在抗性，这是由于分枝杆菌壁的渗透性低和抗生素的主动排出。了解药物外排系统在内在抗生素耐药性中的作用及其调节将导致新的TB治疗方法，因为抑制这些内在系统可能会提高当前药物的功效并缩短治疗时间。 以前我们表明，泛素衍生的肽（UB肽）贡献了巨噬细胞的分枝杆菌曲目（Alonso等，2007； Purdy等，2009）。我们的第一个目标是更好地了解这些肽的杀菌作用。初步研究表明，UB肽的行为像抗菌肽一样损害分枝杆菌膜功能。结核分枝杆菌（MTB）与人免疫反应共同发展，可能具有对宿主抗菌化合物亚致死浓度的抗性机制。在现代疾病中，这些系统可能会提供对抗生素的内在抗性。为了识别MTB固有电阻机制，使用分枝杆菌UB肽作为工具采用了两种互补方法。 1.）我们直接鉴定出在转座子突变体筛选中更容易受到UB肽治疗的ABC和MFS转运蛋白突变体。 2.）分析了暴露于UB肽的MTB的表达曲线。 UB肽调节的基因中有百分之十，编码了预测的膜蛋白，包括ATP结合盒（ABC）或主要的促进剂超家族（MFS）传输系统。这些差异表达的基因改变了对抗菌肽和抗生素的存在的细菌反应。为了证明这一点，我们获得了缺乏ABC和MFS转运蛋白的MTB突变体，发现它们更容易受到抗菌肽和抗生素的影响。我们的工作假设是，UB-肽是宿主抗菌肽，MTB ABC和MFS膜传输系统有助于MTB固有的抗微生物耐药性和毒力。这项工作的长期目标将指导这些膜运输系统的治疗靶向，以提高MTB对携带抗菌化合物和抗生素治疗的固有敏感性。 我们的目标是：1）我们将表征UB肽作用的机制。 2）为了更好地理解MTB的固有电阻，我们将定义ABC转运蛋白PST和RV0986-RV0987，以及MFS转运蛋白RV1634和RV3239C在MTB内在电阻中的作用。 3）我们将确定这些ABC和MFS转运蛋白是否通过巨噬细胞存活分析和小鼠感染有助于结核分枝杆菌的毒力。这些研究将扩大我们对抗菌UB肽和抗生素的结核分枝杆菌固有耐药性的了解，并将确定未来药物治疗方法的靶标。

项目成果

Crystal structure of the transcriptional regulator Rv1219c of Mycobacterium tuberculosis.

结核分枝杆菌转录调节因子 Rv1219c 的晶体结构。

• DOI: 10.1002/pro.2424
• 发表时间: 2014
• 期刊: Protein science : a publication of the Protein Society
• 作者: Kumar,Nitin;Radhakrishnan,Abhijith;Wright,CatherineC;Chou,Tsung-Han;Lei,Hsiang-Ting;Bolla,JaniReddy;Tringides,MariosL;Rajashankar,KanagalaghattaR;Su,Chih-Chia;Purdy,GeorgianaE;Yu,EdwardW
• 通讯作者: Yu,EdwardW

Confinement-Induced Drug-Tolerance in Mycobacteria Mediated by an Efflux Mechanism.

• DOI: 10.1371/journal.pone.0136231
• 发表时间: 2015
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Luthuli BB;Purdy GE;Balagaddé FK
• 通讯作者: Balagaddé FK

Reply to Brennan, "Biofilms and Mycobacterium tuberculosis".

回复布伦南，“生物膜和结核分枝杆菌”。

• DOI: 10.1128/iai.00436-17
• 发表时间: 2017
• 期刊: Infection and immunity
• 影响因子: 3.1
• 作者: Purdy,GeorgianaE