| dc.description.abstract | Piwi-interacting RNAs (piRNAs) are an animal-specific class of germline-enriched small non-coding RNAs that play a crucial role in germline development and fertility in diverse organisms. In Caenorhabditis elegans (C. elegans), mature piRNAs are uniformly 21 nucleotides (nt) long, and begin with monophosphorylated uridine (therefore, also known as 21U-RNA). They are processed from 25-30 nt long capped RNA precursors, which are transcribed by RNA polymerase II and initiate precisely 2 nt upstream of the 5’-end of mature 21U-RNAs. Therefore, to generate 21 nt long mature piRNAs, the 5’ cap and first two nucleotides from the 5’-end must be removed, and extra nucleotides must be trimmed from the 3’-end. It has previously been shown that PARN-1, a conserved 3’-5’ exoribonuclease, contributes to the 3’-end processing of the precursors. However, the enzyme(s) responsible for processing the 5’-end of precursor piRNAs is still unknown.
Objective 1: Identification and characterization of factor(s) involved in the processing of precursor piRNAs in C. elegans.
Here, we report that the endoribonuclease activity of XRN-2 is involved in the 5’-end processing of a small set of piRNAs in C. elegans. Our findings also demonstrate that XRN-2 is capable of degrading precursor piRNAs that are not bound and protected by PRG-1, revealing a potential surveillance mechanism for ensuring quality control during precursor piRNA processing. Our research suggests that piRNAs originating from longer precursor molecules (≥ 60 nt) undergo an initial ribonucleolytic processing of their 3’-ends, which is mediated by ENDU-1, either directly or indirectly. Finally, we show that XRN-2 is not only important for the generation of mature piRNAs and piRNA-dependent 22G-RNAs, but through yet unknown pathways, it also affects piRNA-independent 22G-RNAs that shape transcriptome, as well as contribute to genomic integrity via regulation of transposable elements.
Objective 2: Elucidating a tailing-mediated surveillance pathway for the degradation of unprocessed intermediate precursor 21UR-1 in C. elegans.
Here, we report on a surveillance pathway mediated by 3’-tailing that targets unprocessed intermediate precursor 21UR-1 in the adult germline of C. elegans. The absence of proper 5’- and 3’-end processing events leads to the tailing of intermediate precursor 21UR-1, which acts as a signal for their degradation. CID-1 is identified as the mediator of 3’-tailing of precursor 21UR-1. Additionally, it has been shown that tailing affects the stability of PRG-1. Abrogation of the tailing event leads to a significant accumulation of unprocessed intermediate precursor 21UR-1 in C. elegans embryo, which would otherwise be degraded during the maternal to zygotic transition.
Overall, our study provides insight into precursor piRNA processing and quality control mechanisms in C. elegans. | en_US |
