Postdoctoral Researcher Mater Research Institute Brisbane, Queensland, Australia
Abstract Authors: Stephanie Workman1, Sandra R Richardson1.
1. Mater Research Institute, Brisbane, Australia
Abstract Text: The autonomous retrotransposon Long INterspersed Element 1 (LINE-1) comprises approximately 17 and 18% of the human and mouse genomes respectively. Utilizing a ‘copy and paste’ mechanism of retrotransposition, a subset of evolutionarily young LINE-1 elements haveretained the ability to insert new copies of themselves throughout the genome.As functionally selfish elements,the drive to successfully pass their genetic information onto the next generation and beyondrequires LINE-1s to generate new insertions in pluripotent embryonic cells prior to germ line specification, or in cells of the germ lineage.Not only does LINE-1 activity cause inflammation and aberrant immune activation, but the resulting mutation at insertion sites can be deleterious to genomic function and stability, sometimes contributing to isolated cases of disease in humans.To restrict LINE-1 activity and safeguard their genome, male germ cells employ well characterized defense mechanisms including promoter methylation and the piRNA pathway. Failures in these regulatory systems result in sterility in mice and contribute to cases of human male infertility.However, in oocytes the extent of LINE-1 activityand the regulatory mechanisms in place to curb that activity remain a mystery.We hypothesized that the prolonged state of hypomethylation and transcription endured by arrested oocytes during normal oocyte development leaves them vulnerable to LINE-1 expression and retrotransposition,requiring an oocyte specific defense mechanism not yet characterized.To determine the extent of LINE-1 activity in the oocyte we have carried out a novel analysis of LINE-1 mRNA and protein expression in the murine ovary using a combination of RNA Scope in situ hybridization and immunofluorescence. Strikingly,we foundmajorLINE-1 protein ORF1p expressed in distinct puncta throughout the oocyte cytoplasm across all stages of oocyte maturation from primary to pre-ovulatory follicle.To determine which structures ORF1p was residing in we carried out co-immunostaining with markers of recently characterized specialized oocyte protein compartment Endo Lysosomal Vesicle Assemblies (ELVAs). ORF1pshowed aconsistent co-localization with lysosome component LAMP1 and autophagyprotein LC3A/B in all oocytes.These results suggest that during its maturation the oocyte is exposed to significant LINE-1 expression and utilizes ELVAs to restrict LINE-1 retrotransposition activity.Given that ELVAs have been shown to break down in the zygote, the underlying mechanisms of this female specific regulatory system and its effectiveness against retrotransposition in the early embryo are yet to be explored. Importantly, a disruption to this system may contribute to poor oocyte health, age-associated fertility decline, and early miscarriage.
