Polymorphisms link Protein Kinase C-α to Cryptosporidia susceptibility in 1st year of life

项目摘要 隐孢子虫是一种对全球健康有重要影响的病原体。总的来说，疟疾是 估计造成10%的5岁以下儿童死亡率，最近的全球 隐孢子虫病是仅次于轮状病毒的第二大致死原因， 5岁以下儿童死亡（5）。硝唑尼特，目前唯一批准的隐孢子虫药物 感染，对于治疗免疫活性患者仅是中等有效的。此外，它只是 在严重免疫功能低下患者中相当于安慰剂。隐孢子虫的临床表现 感染的严重程度是可变的;一些患者经历了严重的牙周病，而一些感染仍然存在。 亚临床。尽管对这种广谱疾病的原因知之甚少，但有几项研究表明， 证明了无症状和有症状感染的长期临床影响。我们进行了 全基因组关联研究（GWAS）在孟加拉国的婴儿在生命的第一年。我们评估了 人类基因组中650万个单核苷酸多态性（SNP）与 三个独立患者队列中的症状性隐孢子虫感染。这一分析揭示了一个高度 PRKCA基因中的多个SNP与隐孢子虫易感性的显著统计学关联 腹泻每拷贝一个风险等位基因，在第一次试验中感染隐孢子虫病的风险增加2.4倍。 年的生活。PRKCA中最显著的SNP是GTex数据库中的eQTL，然而， PKCα与隐孢子虫之间的关系尚未确定。PKCα激酶是一种主要的 肌动蛋白和细胞内入侵期间，隐孢子虫诱导宿主细胞肌动蛋白重塑。进一步调查 PRKCA基因变异如何与PKCα功能和对隐孢子虫感染的易感性相关， 提供了一个重要的人类病原体致病机理的见解。我们假设 隐孢子虫通过肌动蛋白重构侵入肠上皮细胞需要宿主PKCα 细胞骨架基于这些初步数据，我们的目标是表征（i）PKCα在隐孢子虫中的作用 （ii）确定SNP对PRKCA表达的影响 和下游功能，和（iii）鉴定PRKCA表达是否介导儿童疾病的严重程度。到 为了实现这一目标，研究计划包括在野外和野外对隐孢子虫进行积极的调查。 长凳。在这里，我们建议通过使用分子工具来弥合科学发现的两种方法， 检查PRKCA并监测基因分型人类儿童的临床结果。的首要目标 这些独立但相互作用的目标将是宿主PKCα的翻译发展， 旨在推进隐孢子虫病急需的治疗。

PROJECT SUMMARY Cryptosporidia is a diarrheal pathogen with an important impact on global health. Overall, diarrheal disease is estimated to be responsible for 10% of under 5-year old child mortality, and the recent Global Burden of Diseases analysis of diarrheal mortality found that Cryptosporidia was second only to rotavirus as a cause of death in children under five years old (5). Nitazoxanide, currently the only approved drug for Cryptosporidia infection, is only moderately effective for treatment of immunocompetent patients. Furthermore, it is only equivalent to placebo in severely immunocompromised patients. Clinical presentation of Cryptosporidia infection is variable in severity; some patients experience severe diarrheal illness while some infections remain subclinical. Although the cause for the wide spectrum of disease is poorly understood, several studies have demonstrated long-term clinical implications of both asymptomatic and symptomatic infection. We performed a genome-wide association study (GWAS) on infants in Bangladesh within the first year of life. We evaluated the association between 6.5 million single nucleotide polymorphisms (SNPs) across the human genome and symptomatic Cryptosporidia infection in three independent patient cohorts. This analysis revealed a highly significant statistical association of multiple SNPs in the PRKCA gene and susceptibility to Cryptosporidia diarrhea. Each copy of the risk allele increased the risk of contracting cryptosporidiosis by 2.4 times in the first year of life. The most significant SNP in PRKCA is an eQTL in the GTex database, however, a definitive link between PKCα and Cryptosporidia has not yet been established. The PKCα kinase is a primary regulator of actin and during intracellular invasion, Cryptosporidia induces host cell actin remodeling. Further investigation into how the PRKCA gene variation relates to PKCα function and susceptibility to Cryptosporidia infection will provide insight into the pathogenesis of an important human pathogen to cause disease. We hypothesize that host PKCα is required for Cryptosporidia invasion of intestinal epithelium through remodeling of actin cytoskeleton. Based on this preliminary data, we aim to characterize (i) the role of PKCα in Cryptosporidia invasion of the intestinal epithelium in vitro and in vivo (ii) define the impact of the SNPs on PRKCA expression and downstream function, and (iii) identify if PRKCA expression mediates severity of disease in children. To accomplish this, the research proposal includes active investigation of Cryptosporidia both in the field and at the bench. Here, we propose bridging two approaches for scientific discovery by using molecular tools to examine PRKCA and monitoring clinical outcomes in genotyped human children. The overarching goal of these independent yet interacting aims will be the translational development of host PKCα as a potential target to advance critically needed treatments for cryptosporidiosis.