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Insights into the piRNA pathway through 16 strains of mice



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Meena, Narendra 


The non-coding genome generates several classes of non-coding regulatory RNAs. One such class of small RNAs - PIWI-interacting RNA (piRNAs) - are exclusively expressed in the germlines of mammals. This thesis provides an exhaustive bioinformatic exploration of piRNA biogenesis and dynamics utilising a large dual-sequencing dataset, which encompasses bulk RNA sequencing and oxidised small RNA sequencing from whole testes. The study encompasses data from 16 unique inbred strains, 12 classical inbred strains and 4 wild inbred strains, studied across three pivotal developmental timepoints in germ cell development and piRNA biogenesis: E16.5, P12.5, and P20.5, in biological triplicates. Through a differential analysis of bulk RNA sequencing, I delve into the complex pathways of piRNAs, examining the expression patterns of related developmental marker genes related to the piRNA pathway and identify strain-specific differences. Notably, gene expression differences indicate that the three wild inbred strains out of four (WSB_EiJ, CAST_EiJ, and SPRET_EiJ) and two classical inbred strain (BALB_cJ and NOD_ShiLTJ) may have a slower developmental trajectory compared to the classical inbred strains, in agreement with modest differential piRNA length distribution dynamics. In addition, CAST_EiJ, PWK_PhJ, and SPRET_EiJ were observed to have the highest counts of unique strain-specific piRNAs, including Transposable Element (TE)-mapping piRNAs across fetal pre-pachytene piRNAs, postnatal pre-pachytene piRNAs and pachytene piRNAs. Overall, TE-mapping piRNAs predominantly overlap with LINE and LTR transposon elements. Using Trinity RNA transcript assemblies with piRNA mapping, I identify previously undiscovered piRNA precursors and clusters for each strain, with a median count of piRNA precursors for E16.5 being 2206, 3240 for P12.5, and 504 for P20.5 across all 16 strains. This both reaffirms previously known piRNA precursors and broadens our understanding of potential piRNA sources and strain-specific diversity. This research also found that the insertion of the IAPEY4_L LTR/ERVK in the NOCT region is associated with the expression of piRNA precursors, as evidenced by their presence in 7 specific mouse strains (C57BL/6NJ, BALB/cJ, A/J, DBA/2J, AKR/J, NZO/HlLtJ, NOD/ShiLtJ) where these transposable elements (TE) are found. Conversely, strains lacking this TE insertion do not show similar expression patterns. In sum, this work offers new perspectives on piRNA dynamics and their influence on germ cell development across 16 diverse mice inbred strains.





Miska, Eric


mice, piRNA, TE


Doctor of Philosophy (PhD)

Awarding Institution

University of Cambridge