Regulatory small RNA pathways in the nematode C. elegans and the ant T. rugatulus
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Abstract
Epigenetic regulation via RNA interference (RNAi) is an important method for altering gene expression and for silencing transposons; transposable genetic elements which can self-insert into the genome at various locations, thus wreaking havoc on the genome when not kept in check. RNAi functions via proteins known as Argonautes which associate with small RNAs, using these to direct them to their targets via sequence complementarity. This thesis concerns different pathways related to RNAi in the nematode Caenorhabditis elegans, in which RNAi has long been studied, as well as in the ant, Temnothorax rugatulus, in which RNAi has not priorly been studied. In Chapter I, we show how the worm-specific Argonaute WAGO-3 is paternally inherited via the sperm in C. elegans, where it localizes to the newly found PEI-granule, whose phase-separation is controlled by the proteins PEI-1 and PEI-2. We show evidence to support the segregation into budding spermatids of PEI-1/2 and WAGO-3 via anchoring to fibrous body–membranous organelles (FB-MOs). We also show that WAGO-3 is important for paternal inheritance of transgenerational epigenetic memory (RNAe).
In Chapter II, we show that T. rugatulus expresses at least two different classes of small RNAs; miRNAs and piRNAs. We show that miRNAs are related to caste-differentiation and that the piRNA pathway, relevant for transposon silencing, is active even in rudimentary ovaries incapable of reproducing.
Finally, in Chapter III, we provide new insights into the functionalities of the three germline-expressed, worm-specific Argonautes WAGO-1, WAGO-3, and WAGO-4 in C. elegans. We show evidence that WAGO-1 and WAGO-4 influence the paternal ALG-3/4 pathway, possibly via interaction with the nuclear Argonaute WAGO-9/HRDE-1. WAGO-3, on the other hand, we find to influence the maternal ERGO-1 pathway, most likely also via interaction with one or more nuclear Argonautes. We find that Argonaute regulation depends on the lifestage of C. elegans, and we lastly show that mRNA misregulation caused by loss of WAGO-4 is
remembered transgenerationally.