The Plasmodial Surface Anion Channel And Malaria Parasite Nutrient Acquisition

疟原虫表面阴离子通道与疟原虫营养获取

• 批准号: 7592254
• 负责人: SANJAY A DESAI
• 金额: $ 75.53万
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7592254
• 关键词: Affect; Affinity; Alanine; Ammonium; Anions; Antimalarials; Basic Science; Biochemical; Biological; Biological Assay; Biophysics; Cells; Cellular biology; Cloning; Collection; Dependence; Drug resistance; Electrophysiology (science); Erythrocytes; Exhibits; Genes; Genetic Materials; Goals; Growth; Human; In Vitro; Infection; Informatics; Ion Channel; Ion Transport; Isoleucine; Lead; Libraries; Lipid Biochemistry; Malaria; Mediating; Membrane; Membrane Proteins; Modeling; Molecular; Molecular Biology; Numbers; Nutrient; Parasites; Pathway interactions; Permeability; Pharmacology; Phenotype; Physiology; Predisposition; Property; Proteins; Range; Resistance; Role; Route; Sorbitol; Specificity; Structure-Activity Relationship; Surface; Temperature; Therapeutic; Thinking; Toxin; Work; blasticidin S; cost; drug development; fitness; gene cloning; high throughput screening; inhibitor/antagonist; killings; mutant; novel strategies; resistance mechanism; small molecule; solute; uptake

The Apicomplexan Molecular Physiology Section continued its studies of the plasmodial surface anion channel (PSAC) and made two significant contributions. First, we identified and characterized the first PSAC mutant (PNAS 104:1063-1068, 2007). This mutant was generated by in vitro selection with blasticidin S, a toxin that reaches its intracellular target via permeation through PSAC. Because blasticidin S resistance correlated with reduced permeability to multiple solutes, we predicted there may be marked changes in PSAC. Single-channel and whole-cell electrophysiology confirmed this prediction by revealing altered channel gating, selectivity, pharmacology, and functional copy number/infected cell. Mutant parasites cultured without blasticidin S reverted to the wild-type channel phenotype and exhibited restored susceptibility to killing by blasticidin S. These findings 1) confirm that PSAC is the primary mechanism of organic solute uptake after infection because changes in PSAC affected the permeability of each solute, 2) implicate parasite genes in the expression of PSAC because human erythrocytes lack heritable genetic material, 3) provide a new approach to cloning PSACs gene, 4) reveal a new drug resistance mechanism in malaria parasites, and 5) suggest that PSAC serves an essential role for the intracellular parasite because a fitness cost was associated with blasticidin S resistance. Second, we identified solute-inhibitor interactions within PSACs pore (Mol. Pharmacol. 71:1241-50, 2007). In this study, we found that phenyltrimethyl ammonium and isoleucine transport through PSAC were less effectively inhibited by known PSAC antagonists than the uptake of sorbitol and alanine. This observation was unexpected because all four solutes were thought to share a single transport mechanism. We excluded uptake via unrelated channels because specific PSAC inhibitors also exhibited solute-dependent affinities. Mixtures of permeating solutes, whole-cell electrophysiology, and temperature-dependence studies suggested that a single ion channel with two separate routes for permeating solutes is the most conservative explanation for our findings. Such a model may permit fine-tuning of PSACs unusual selectivity and allow the parasite to acquire a diverse collection of nutrients. These findings also suggest that PSAC antagonists must effectively block both routes to be suitable lead compounds for antimalarial drug development.

Apicomplexan分子生理学部分继续研究了质体表面阴离子通道（PSAC），并做出了两个重要的贡献。 首先，我们确定并表征了第一个PSAC突变体（PNAS 104：1063-1068，2007）。 该突变体是通过用blasticidin s进行体外选择而产生的，blasticidin s是通过PSAC渗透到其细胞内靶标的毒素。 由于Blasticidin的耐药性与多种溶质的渗透性降低相关，因此我们预测PSAC可能发生明显变化。 单通道和全细胞电生理学通过揭示了变化的通道门控，选择性，药理学和功能拷贝数/感染细胞来证实这一预测。 没有蓝乳肽的培养的突变寄生虫恢复为野生型通道表型，并表现出恢复的易感性，可恢复杀死blasticidin的S。这些发现1）确认，PSAC是感染后有机溶质吸收的主要机制提供了一种克隆PSAC基因的新方法，4）揭示了疟疾寄生虫的新耐药性机制，5）表明PSAC对细胞内寄生虫起着至关重要的作用，因为健身成本与blasteridin的耐药性有关。 其次，我们确定了PSACS孔中的溶质抑制剂相互作用（Mol。Pharmacol。71：1241-50，2007）。 在这项研究中，我们发现已知的PSAC拮抗剂比山梨糖醇和丙氨酸的摄取对通过PSAC的苯基三甲基铵和异亮氨酸转运的抑制较少。 这一观察结果是出乎意料的，因为所有四种溶质都被认为具有单一的运输机制。 我们通过无关的通道排除了吸收，因为特定的PSAC抑制剂也表现出溶质依赖性亲和力。 渗透溶质，全细胞电生理学和温度依赖性研究的混合物表明，具有两种单独渗透溶质途径的单个离子通道是我们发现的最保守的解释。 这样的模型可以允许对PSAC进行微调异常的选择性，并允许寄生虫获得多种营养的收集。 这些发现还表明，PSAC拮抗剂必须有效地阻断两种途径，以作为抗疟药开发的合适铅化合物。