Role of erythrocyte band 3 in malaria adhesion

红细胞带 3 在疟疾粘附中的作用

• 批准号: 10669600
• 负责人: Athar H. Chishti
• 金额: $ 53.05万
• 依托单位: TUFTS UNIVERSITY BOSTON
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-07-10 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10669600
• 关键词: Address; Adhesions; Adhesives; Affinity; Amino Acids; Anions; Binding; Binding Proteins; Biochemical; Biological Assay; Biological Process; Blood Vessels; Brain; C-Type Lectins; Cell Line; Cells; Cerebral Malaria; Cerebrum; Cessation of life; Child; Coma; Complementary DNA; Complex; Cytoplasm; Cytoskeleton; Deposition; Economic Development; Endothelial Cells; Endothelium; Erythrocyte Anion Exchange Protein 1; Erythrocyte Membrane; Erythrocytes; Extracellular Domain; Functional disorder; Glutamic Acid; Health; Human; Hypoxia; Imaging Device; Immunoprecipitation; Infection; Inflammation; Integral Membrane Protein; Kinetics; Knock-out; Laboratories; Lesion; Ligands; Link; Location; Malaria; Mass Spectrum Analysis; Mediating; Membrane; Membrane Proteins; Monoclonal Antibodies; Nature; Oxygen; Parasites; Pathogenesis; Pathway interactions; Peptides; Persons; Phage Display; Phenotype; Phosphorylation; Plasmodium falciparum; Property; Proteins; Recombinants; Role; Series; Surface; TNF gene; Testing; Therapeutic; Thrombospondins; Vacuole; Vascular Endothelial Cell; Vascular Endothelium; cDNA Library; ectoplasm; extracellular; histidine-rich proteins; malaria infection; mortality; neoantigens; novel; postcapillary venule; protein complex; protein function; real time monitoring; receptor; recruit; synthetic peptide; targeted treatment; therapeutic target

Plasmodium falciparum infections cause malaria resulting in the deaths of ~435,000 people, mostly children, in malaria-endemic regions each year. A distinct feature of P. falciparum is sequestration; the adhesion of infected RBCs (iRBCs) containing mature parasites to endothelial cells. iRBCs display surface bumps known as knobs that mediate the attachment of iRBCs to endothelium. Both parasite and host proteins located at the knobs contribute to the cytoadhesion phenotype. Strategies that can disrupt interactions of knobs offer potential therapeutic means of attenuating their adhesive properties; however, their precise mechanisms remain poorly understood. Using phage display cDNA screens, we identified a new knob parasite protein termed PfGARP. P. falciparum glutamic acid-rich protein (PfGARP) is exported outside of the RBCs and binds to an extracellular segment of band 3. This novel localization of PfGARP suggests a functional role in the adhesion phenotype of iRBCs. Based on these findings, we hypothesize that PfGARP contributes to the cytoadhesion of malaria infected human RBCs to vascular endothelium in a PfEMP1-independent manner by engaging a novel ligand-receptor(s) complex on the endothelial surface. We will test this hypothesis as follows: Specific Aim 1: Mechanism of PfGARP interactions with human erythrocytes/RBCs. Experimental support comes from our immuno-colocalization studies with known knob markers, PfEMP1 and KAHRP, placing PfGARP at the knobs. Since a truncated form of band 3, which binds to PfGARP, is also localized to knobs, this raises the possibility that the PfGARP-Band 3 complex may function as an PfEMP1-independent interaction linking iRBCs to the endothelial cells. We propose to identify the precise ectoplasmic segment of band 3 and critical amino acid residues that mediate Band 3-PfGARP interaction. We plan to use the high-affinity peptides of band 3 for assessing the functional effects of PfGARP binding to human RBCs including adhesion, deformability, and band 3 phosphorylation. We will employ immuno-EM, immunoprecipitation and mass spectrometry to investigate the localization and composition of PfGARP complex on knobs. A PfGARP knockout parasite strain will be generated to evaluate the adhesive properties of iRBCs by multiple cytoadhesion assays. Aim 2: Identification of host endothelial receptors for PfGARP. We generated a phage display cDNA library from human endothelial cells (TNFα-activated HUVECs) and screened with recombinant PfGARP as bait. Among several potential receptors identified in the screens, we are currently characterizing a single-pass transmembrane receptor termed CLEC14A (C-type lectin domain containing 14A) as a potential binding partner of PfGARP. To test the possibility that PfGARP engages CLEC14A to recruit iRBCs to the vascular endothelium, we will use state-of-the-art imaging tools developed for endothelial cell lines in the Herman laboratory at Tufts to monitor real-time kinetics of cytoadherence ex vivo. Together, the proposed studies will contribute to our understanding of the biological function of knobs and role of PfGARP in the adhesion of iRBCs to microvascular endothelial cells.

恶性疟原虫感染导致疟疾，导致约 435,000 人死亡，其中大部分是儿童 每年疟疾流行地区的恶性疟原虫的一个显着特征是感染者的粘附。 含有成熟内皮细胞寄生虫的红细胞（iRBC）显示出称为旋钮的表面凸起。 介导 iRBC 与内皮细胞的附着，寄生虫和宿主蛋白均位于旋钮处。 有助于细胞粘附表型的策略可以破坏旋钮的相互作用提供潜在的机会。 减弱其粘附特性的治疗方法；然而，其精确机制仍然很差。 通过噬菌体展示 cDNA 筛选，我们鉴定出一种新的旋钮寄生虫蛋白，称为 PfGARP P。 恶性疟原虫富含谷氨酸的蛋白（PfGARP）被输出到红细胞外并与细胞外结合 带 3 的片段。PfGARP 的这种新定位表明其在粘附表型中的功能作用 基于这些发现，我们发现 PfGARP 有助于疟疾的细胞粘附。 通过使用一种新颖的方法，以不依赖于 PfEMP1 的方式将人类红细胞感染到血管内皮 内皮表面上的配体-受体复合物 我们将按如下方式检验该假设： 具体目标。 图 1：PfGARP 与人红细胞/红细胞相互作用的机制来自实验支持。 我们使用已知的旋钮标记 PfEMP1 和 KAHRP 进行免疫共定位研究，将 PfGARP 置于 由于与 PfGARP 结合的带 3 的截短形式也定位于旋钮，因此这提高了 PfGARP-Band 3 复合物可能作为连接 iRBC 的 PfEMP1 独立相互作用发挥作用 我们建议鉴定带 3 和关键氨基的精确胞质片段。 介导带 3-PfGARP 相互作用的酸残基 我们计划使用带 3 的高亲和力肽。 评估 PfGARP 与人类红细胞结合的功能影响，包括粘附、变形性和条带 我们将采用免疫电镜、免疫沉淀和质谱法来研究 3 磷酸化。 PfGARP 复合物在旋钮上的定位和组成将产生 PfGARP 敲除寄生虫菌株。 通过多种细胞粘附测定评估 iRBC 的粘附特性 目标 2：宿主的鉴定。 我们从人内皮细胞中生成了噬菌体展示 cDNA 文库。 （TNFα 激活的 HUVEC）并以重组 PfGARP 作为诱饵进行筛选。 在屏幕中识别出，我们目前正在表征一种单程跨膜受体，称为 CLEC14A（含有 14A 的 C 型凝集素结构域）作为 PfGARP 的潜在结合伙伴来测试可能性。 PfGARP 利用 CLEC14A 将 iRBC 招募到血管内皮，我们将使用最先进的技术 塔夫茨大学赫尔曼实验室为内皮细胞系开发的成像工具，用于监测实时动力学 总之，拟议的研究将有助于我们对生物学的理解。 旋钮的功能以及 PfGARP 在 iRBC 与微血管内皮细胞粘附中的作用。

项目成果

BDA-410 inhibits SARS-CoV-2 main protease activity and viral replication in mammalian cells.

• DOI: 10.1111/jcmm.17442
• 发表时间: 2022-10
• 期刊: Journal of cellular and molecular medicine
• 影响因子: 5.3

Plasmodium falciparum signal peptide peptidase is a promising drug target against blood stage malaria.

• DOI: 10.1016/j.bbrc.2009.01.083
• 发表时间: 2009-03-13
• 期刊: BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
• 影响因子: 3.1
• 作者: Li, Xuerong;Chen, Huiqing;Bahamontes-Rosa, Noemi;Kun, Jurgen F. J.;Traore, Boubacar;Crompton, Peter D.;Chishti, Athar H.
• 通讯作者: Chishti, Athar H.

The cytoadherence ligand Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) binds to the P. falciparum knob-associated histidine-rich protein (KAHRP) by electrostatic interactions.

细胞粘附配体恶性疟原虫红细胞膜蛋白 1 (PfEMP1) 通过静电相互作用与恶性疟原虫旋钮相关富含组氨酸蛋白 (KAHRP) 结合。

• DOI: 10.1016/s0166-6851(00)00281-4
• 发表时间: 2000
• 期刊: Molecular and biochemical parasitology
• 影响因子: 1.5
• 作者: Voigt,S;Hanspal,M;LeRoy,PJ;Zhao,PS;Oh,SS;Chishti,AH;Liu,SC
• 通讯作者: Liu,SC

Host receptors in malaria merozoite invasion.

疟疾裂殖子入侵中的宿主受体。

• DOI: 10.1007/3-540-29088-5_8
• 发表时间: 2005
• 期刊: Current topics in microbiology and immunology
• 作者: Oh,SS;Chishti,AH
• 通讯作者: Chishti,AH

Safe and effective delivery of supplemental iron to healthy adults: a two-phase, randomized, double-blind trial - the safe iron study.

• DOI: 10.3389/fnut.2023.1230061
• 发表时间: 2023
• 期刊: FRONTIERS IN NUTRITION
• 影响因子: 5
• 作者: Lewis, Erin D.;Ortega, Edwin F.;Dao, Maria Carlota;Barger, Kathryn;Mason, Joel B.;Leong, John M.;Osburne, Marcia S.;Magoun, Loranne;Nepveux, V. Felix J.;Chishti, Athar H.;Schwake, Christopher;Quynh, Anh;Gilhooly, Cheryl H.;Petty, Gayle;Guo, Weimin;Matuszek, Gregory;Pereira, Dora;Reddy, Manju;Wang, Jifan;Wu, Dayong;Meydani, Simin N.;Combs Jr, Gerald F.
• 通讯作者: Combs Jr, Gerald F.