Roles for small noncoding RNAs in silencing of retrotransposons in the mammalian brain

Abstract

Piwi-interacting RNAs (piRNAs), long thought to be restricted to germline, have recently been discovered in neurons of Aplysia, with a role in the epigenetic regulation of gene expression underlying long-term memory. We here ask whether piwi/piRNAs are also expressed and have functional roles in the mammalian brain. Large-scale RNA sequencing and subsequent analysis of protein expression revealed the presence in brain of several piRNA biogenesis factors including a mouse piwi (Mili), as well as small RNAs, albeit at low levels, resembling conserved piRNAs in mouse testes [primarily LINE1 (long interspersed nuclear element1) retrotransposon-derived]. Despite the seeming low expression of these putative piRNAs, single-base pair CpG methylation analyses across the genome of Mili/piRNA-deficient (Mili −/− ) mice demonstrate that brain genomic DNA is preferentially hypomethylated within intergenic areas and LINE1 promoter areas of the genome. Furthermore, Mili mutant mice exhibit behavioral deficits such as hyperactivity and reduced anxiety. These results suggest that putative piRNAs exist in mammalian brain, and similar to the role of piRNAs in testes, they may be involved in the silencing of retrotransposons, which in brain have critical roles in contributing to genomic heterogeneity underlying adaptation, stress response, and brain pathology. We also describe the presence of another class of small RNAs in the brain, with features of endogenous siRNAs, which may have taken over the role of invertebrate piRNAs in their capacity to target both transposons, as well as protein-coding genes. Thus, RNA interference through gene and retrotransposon silencing previously encountered in Aplysia may also have potential roles in the mammalian brain.piwi-interacting RNA | transposon | DNA methylation | behavior | endogenous siRNA P iwi-interacting RNAs (piRNAs) are 26-to 32-nt small noncoding RNAs that associate with the piwi clade of the Argonaute family of proteins and whose primary functions in mammals are associated with the silencing of transposons in the germline through de novo DNA methylation (1-4). Transposons constitute 44% of the human genome and could when active contribute to genetic heterogeneity, to alteration of behavior during adaptation, and to responses to stress. Somatic retrotransposition and its associated insertional mutagenesis have particularly important implications for brain disorders and are often associated with schizophrenia, Rett syndrome, and neurodegenerative disorders (5-7). The LINE1 (long interspersed nuclear element1) family of retrotransposons in particular is active in human and mouse hippocampus (8, 9).Although it has long been thought that piRNA expression and function are restricted to the germline, our laboratory and others have previously found that the piRNA pathway is functional in invertebrate neurons as well (10, 11). Aplysia neurons, in particular, have a strikingly abundant expression of piRNAs (contributing to at least 5% of the total noncoding RNA population); they in turn hav...

A Neuronal piRNA Pathway Inhibits Axon Regeneration in C. elegans

“…However, PIWI/piRNAs are reportedly expressed in the nervous system of flies, Aplysia, and mice (Lee et al, 2011; Nandi et al, 2016; Perrat et al, 2013; Rajasethupathy et al, 2012). In Aplysia sensory neurons neuronal piRNAs function in long-term facilitation (Rajasethupathy et al, 2012).…”

Hippocampal knockdown of Piwil1 and Piwil2 enhances contextual fear memory in mice

“…A more recent exploration of neuronal piRNAs identified approximately 30,000 piRNAs from mouse brain tissue (Ghosheh, Seridi, Ryu, Takahashi, Orlando, Carninci, and Ravasi, 2016) and suggested that these brain-derived piRNAs are most similar to those associated with PIWIL2 in mouse testes. Another study identified low-level expression of truncated PIWIL2 in the adult mouse brain, with hypomethylation of transposons and behavioural defects observed in Piwil2 knockout mice (Nandi, Chandramohan, Fioriti, Melnick, Hébert, Mason, Rajasethupathy, and Kandel, 2016). We have also previously demonstrated that the Piwi-like genes Piwil1 and Piwil2 are expressed in cultured mouse neurons, and have shown that they are dynamically regulated by neuronal activation, and are involved in transcriptional regulation of activityinduced gene expression (Leighton, Zhao, Li, Dai, Marshall, Liu, Wang, Zajaczkowski, Khandelwal, and Kumar, 2018).…”

“…These reports are congruent with the finding that de novo coding mutations in human Piwil genes are strongly associated with autism-spectrum disorders (Iossifov et al, 2014) which are increasingly understood to involve aberrant organisation of cortical neurons (Nunes, Peatfield, Vakorin, and Doesburg, 2018;Packer, 2016). In addition, Nandi et al found that Piwil2 knockout mice are hyperactive and that males show reduced anxiety (Nandi et al, 2016); because the genetic absence of Piwil2 is present from conception, it is likely that this phenotype is also developmental. In the present study, we have demonstrated for the first time that targeted manipulation of the Piwi pathway in the adult mammalian brain affects behaviour.…”

Alterations in retrotransposition, synaptic connectivity, and myelination implicated by transcriptomic changes following maternal immune activation in non-human primates

“…is a master regulator of piRNA mediated DNA methylation and an inhibitor of retrotransposition (47,48). PIWIL2 also regulates the expression of plasticityrelated genes in the adult brain, and disruption of the piwi pathway in HC enhances contextual fear memory in mice (49).…”