Identification of Entamoeba small RNAs by pyrosequencing

描述（由申请方提供）：溶组织内阿米巴是一种原生动物寄生虫，是一种重要的人类病原体。E.溶组织菌包括痢疾和肝脓肿，且该生物体是全球范围内主要寄生虫性死因。基因表达的调节是使寄生虫在组织入侵期间转变为包囊期并传播疾病和适应宿主环境的关键因素。在大肠histolytica中，我们已经鉴定了一群27 nt大小的小RNA，其（i）具有52-聚磷酸末端，（ii）与基因反义，和（iii）与E.溶组织菌Argonaute蛋白。全基因组微阵列表达分析显示，基本上所有的基因，反义小RNA地图不表达滋养体条件下，寄生虫阶段，从小RNA克隆。然而，其中许多基因在其他大肠杆菌中表达。结果表明，在溶组织菌菌株中，小RNA和基因表达水平之间存在负相关性，表明这些小RNA介导同源基因的沉默。总的来说，我们的结果表明，E。histolytica具有丰富的27 nt群体，具有与来自C. elegans，并且似乎调节基因表达。这些数据表明，由52-聚磷酸siRNA介导的沉默途径延伸到单细胞真核生物。我们的假设是，RNAi样沉默途径在大肠杆菌中起作用。可能调节寄生虫分化和毒力的关键基因。我们的目标是获得E. histolytica，重点是sRNA差异表达的毒力和无毒力的寄生虫和阶段转换。这项工作将提供一个内阿米巴小RNA的参考数据库，很容易与更广泛的科学界分享，并成为该领域的重大进展。 公共卫生相关性：溶组织内阿米巴是一种重要的病原体，在全球范围内影响人类健康。虽然大多数疾病发生在发展中国家，但这种寄生虫可以在任何地方引起感染，从而对水净化系统产生不利影响。我们有兴趣了解寄生虫用于引起疾病的分子机制。

DESCRIPTION (provided by applicant): Entamoeba histolytica, a protozoan parasite, is an important human pathogen. Diseases caused by E. histolytica include dysentery and liver abscesses and this organism is a leading parasitic cause of death on a global scale. Regulation of gene expression is a key factor that enables the parasite to convert to the cyst stage and propagate disease and adapt to the host environment during tissue invasion. In E. histolytica, we have identified a population of small RNAs of 27nt size that (i) have a 52-polyphosphate termini, (ii) map antisense to genes, and (iii) associate with an E. histolytica Argonaute protein. Whole genome microarray expression analysis revealed that essentially all genes to which antisense small RNAs map were not expressed under trophozoite conditions, the parasite stage from which the small RNAs were cloned. However, a number of these genes were expressed in other E. histolytica strains with an inverse correlation between small RNA and gene expression level suggesting that these small RNAs mediate silencing of the cognate gene. Overall, our results demonstrate that E. histolytica has an abundant 27nt population, with features similar to secondary siRNAs from C. elegans, and which appear to regulate gene expression. These data indicate that a silencing pathway mediated by 52-polyphosphate siRNAs extends to single celled eukaryotic organisms. Our hypothesis is that an RNAi-like silencing pathway is functional in E. histolytica and may regulate genes key to parasite differentiation and virulence. Our goal is to obtain a comprehensive profile of small RNA species in E. histolytica with an emphasis on sRNAs differentially expressed in virulent and non-virulent parasites and during stage conversion. This work will provide a reference database of Entamoeba small RNAs, be readily shared with the broader scientific community and be a significant advance for the field. PUBLIC HEALTH RELEVANCE: Entamoeba histolytica is an important pathogen with an impact on human health on a global scale. Although the majority of disease is in developing countries, this parasite can cause infections anywhere that water purification systems get adversely affected. We are interested in understanding the molecular mechanisms that the parasite uses to cause disease.