Identification and characterisation of novel piRNA processing factors in C. elegans

秀丽隐杆线虫新型piRNA加工因子的鉴定与表征。小RNA分子可以在基因表达途径中起调节作用。靶标的精确识别基于碱基配对，sRNA作为特异性因子，其加工过程需要严格控制。其中一个保守的小RNA沉默途径是所谓的Piwi途径。这种途径是生殖细胞中控制转座因子的主要监视系统之一，由称为pirna的小rna操纵。鉴于转座子序列的广泛多样性和多样性，需要很好地控制piRNA序列库，以避免宿主序列的识别。这是通过选择特定的转录本作为piRNA前体，然后进行不同的处理步骤来实现的。一般来说，选择过程和处理步骤都没有得到很好的理解。在秀丽隐杆线虫（C. elegans）中，pirna来源于特异性的短RNA聚合酶II转录物。随后，它们被一种名为PETISCO的专用蛋白质复合物结合，这种复合物允许一种尚未识别的核酸酶处理它们的5 '端。这一步骤是不同活动的组合，如去盖，然后是核糖核酸外溶酶修剪，还是由一种不同的核糖核酸内溶酶驱动，目前尚不清楚。我们提出的初步数据表明，核糖核酸内酶是由一种异二聚体蛋白复合物构成的，该复合物特异性作用于秀丽隐杆线虫中petisco结合的piRNA前体。提出的工作计划旨在巩固这一假设，并了解这种新的酶活性。我们想了解它的三维结构以及它是如何与PETISCO联系在一起的。此外，我们确定了另外两个在piRNA生物发生中起作用的因素，并在结构域水平上类似于构成二聚体内切酶的蛋白质。我们的数据表明，基于piRNA加工核酸酶的两个亚基之一，可能存在其他类型核酸酶的潜在模块化异二聚体组装。我们的目标是了解这些复合物在生化和有机体水平上的功能。

Identification and characterization of novel piRNA processing factors in C. elegans.Small RNA molecules can act as regulators in gene expression pathways. The precise recognition of targets is based on base pairing, and the sRNA thus acts as specificity factors, and consequently, their processing needs to be tightly controlled. One of the well-conserved small RNA silencing pathways is the so-called Piwi pathway. This pathway is one of the main surveillance systems in germ cells to keep transposable elements under control and is steered by small RNAs known as piRNAs. Given the wide diversity and varying nature of the transposon sequences, the piRNA sequence repertoire needs to be well controlled to avoid the recognition of host sequences. This happens by selecting specific transcripts as piRNA precursors, followed by distinct processing steps. Neither the selection process nor the processing steps are in general well understood.In the nematode Caenorhabditis elegans (C. elegans), the piRNAs are derived from specific, short RNA polymerase II transcripts. Following this, they are bound by a dedicated protein complex named PETISCO, which allows the processing of their 5′ end by an as-yet-unidentified nuclease. Whether this step is a compilation of different activities, such as de-capping, followed by exoribonucleolytic trimming, or is driven by a distinct endoribonucleolytic enzyme is unknown.We present preliminary data that implicate that the endoribonuclease is constituted by a heterodimeric protein complex that specifically acts on PETISCO-bound piRNA precursors in C. elegans. The proposed work plans are aimed to solidify this hypothesis and to understand this novel enzymatic activity. We want to learn about its 3-dimensional structure and how it is connected to PETISCO. In addition, we identified two additional factors that play a role in piRNA biogenesis and resemble proteins constituting dimeric endonuclease on the domain level. Our data suggest a potential modular heterodimeric assembly of potentially other types of nucleases, based on one of the two subunits of the piRNA processing nuclease. We aim to understand the function(s) of those complexes both at the biochemical and organismic levels.