Identification of bona fide B2 SINE retrotransposon transcription through single-nucleus RNA-seq of the mouse hippocampus

Abstract

Currently, researchers rely on generalized methods to quantify transposable element (TE) RNA expression, such as RT-qPCR and RNA-seq, that do not distinguish between TEs expressed from their own promoter (bona fide) and TEs that are transcribed from a neighboring gene promoter such as within an intron or exon. This distinction is important owing to the differing functional roles of TEs depending on whether they are independently transcribed. Here we report a simple strategy to examine bona fide TE expression, termed BonaFide-TEseq. This approach can be used with any template-switch based library such as Smart-seq2 or the single-cell 5 ′ gene expression kit from 10x, extending its utility to single-cell RNAsequencing. This approach does not require TE-specific enrichment, enabling the simultaneous examination of TEs and protein-coding genes. We show that TEs identified through BonaFide-TEseq are expressed from their own promoter, rather than captured as internal products of genes. We reveal the utility of BonaFide-TEseq in the analysis of single-cell data and show that short-interspersed nuclear elements (SINEs) show cell type-specific expression profiles in the mouse hippocampus. We further show that, in response to a brief exposure of home-cage mice to a novel stimulus, SINEs are activated in dentate granule neurons in a time course that is similar to that of protein-coding immediate early genes. This work provides a simple alternative approach to assess bona fide TE transcription at single-cell resolution and provides a proof-of-concept using this method to identify SINE activation in a context that is relevant for normal learning and memory.

Corticosterone dynamically regulates retrotransposable element expression in the rat hippocampus and C6 cells

“…First, they showed that B2 SINE RNA was enriched in fos + DG cells following novel environment exposure ( Lacar et al, 2016 ). Subsequently, this group confirmed that these B2 SINE RNAs were in fact bona fide transcripts featuring the necessary machinery for independent transcriptional activation - not simply an artifact of “read-through” transcriptional events ( Linker et al, 2020 ). These findings suggest the capacity for hippocampal B2 SINE transposons to be transcriptionally activated on the same time scale as immediate early genes by acute stress, supporting a potential role for rapid activation B2 SINEs by corticosteroids.…”

“…While these experiments have only included males, preliminary findings have suggested that hippocampal B2 SINE transcript abundance may be sexually dimorphic ( Lambert et al, 2019 ). Future studies leveraging next-generation sequencing approaches are needed to determine both the presence of internal promoters within CORT activated transposon transcripts, so called bona fide transcripts, as well as the precise genomic loci of transposon-derived RNA ( Linker et al, 2020 ; Guffanti et al, 2018 ).…”

Retrotransposons as Drivers of Mammalian Brain Evolution

“…To what extent, and according to which species-specific paths, brain functionality in mammals relies on retrotransposon exaptation will become increasingly clear as the particularly challenging aspects of retrotransposon molecular biology become easier to address. Major challenges include: (i) the unambiguous identification of expressed retrotransposon loci, together with the related issue of insertional and internal sequence polymorphisms [ 192 ], which becomes highly relevant when studies of retrotransposon impact are to be conducted at the population level; (ii) the implementation of a single-cell perspective while addressing the expression and genomic impact of retrotransposons, which appears particularly relevant in the case of brain biology given the extremely diverse specialization of neurons [ 193 , 244 ]; (iii) the experimental assessment of cis -regulatory roles of retrotransposons by means of their systematic perturbation, e.g., through CRISPR-based approaches [ 120 ].…”

“…In particular, the Alu-derived human NDM29 transcript induces a neuron-like phenotype when transfected into undifferentiated neuroblastoma cells [ 188 ], and members of the snaR family of ncRNAs, with a possible role in translation, were found to be differentially expressed in different brain regions [ 189 ]. Given the recent improvement of methods of detecting and quantifying the expression of individual SINE loci in cells and tissues [ 144 , 190 , 191 , 192 ], including single neurons [ 193 ], it is likely that more cases will be revealed of exapted SINEs producing ncRNAs involved in brain function.…”

Transposable elements and their KZFP controllers are drivers of transcriptional innovation in the developing human brain

“…We propose that increased postnatal TE expression may possibly be reflective of the development of cell types not present in early prenatal stages, such as astrocytes, microglia and oligodendrocytes, the developmental and transcriptional trajectories of which were identified by scRNA-seq analyses . To determine the transposcriptome in scRNA-seq data remains technically challenging because many TE-derived transcripts are lowly abundant, a limitation that will hopefully be alleviated by progress in sequencing techniques and computational approaches (Linker et al 2020). Of note, TEs heavily contribute to long non-coding RNAs (lncRNAs), which are abundant in the human brain (Derrien et al 2012;Kelley and Rinn 2012;Zimmer-Bensch 2019).…”