Activation of endogenous retroviruses during brain development causes neuroinflammation

Endogenous retroviruses (ERVs) make up a large fraction of mammalian genome and are thought to contribute to human disease, including brain disorders. Aberrant activation of ERVs constitute a potential trigger for neuroinflammation, but mechanistic insight into this phenomenon remains unclear. Using CRISPR/Cas9-based gene disruption of the epigenetic co-repressor protein Trim28, we found a dynamic H3K9me3-dependent regulation of ERVs in proliferating neural progenitor cells (NPCs), but not in adult neurons. In vivo disruption of Trim28 in cortical NPCs during brain development resulted in viable offspring expressing high levels of ERVs in excitatory neurons in the adult brain of mice. Neuronal ERV expression was linked to inflammation, including activated microglia, and aggregates of ERV-derived proteins. This study demonstrates that brain development is a critical period for the silencing of ERVs and provides causal in vivo evidence demonstrating that transcriptional activation of ERV in neurons results in neuroinflammation. Overall design: In vitro CRISPR Trim28-KO in mouse neural progentiro cells (NPCs) with 3 guide RNAs (guide 3, 4 and 13 targeting exon 3, 4 and 13 of Trim28) where each guide has two duplicates. Two controls (where guide RNA towards LacZ were used). In vivo CRISPR Trim28-KO (cas knockin mice): five Cas9-GFP knock in mice (referred as Cas in this document) where injected with guide RNAs 3 or 4 or 13 (targeting exon 3, 4 and 13), or all three guide RNA combined, plus one control (guideRNA towards LacZ). All guideRNAs were co-injected with AAV vectors expressing Cre. In vivo CRISPR Trim28-KO (Stop-Cas knockin mice): five stop-Cas9-GFP knock in mice (referred as StopCas in this document) where injected with AAV vectors expressing guide RNAs 3 or 4 or 13 (targeting exon 3, 4 and 13), or all three guide RNA combined, plus one control (guideRNA towards LacZ). All guideRNAs were co-injected with AAV vectors expressing Cre. In vivo Trim28-KO (Trim28-fl mice): AAV.Cre injections into Trim28-fl animals (+/- and +/+): 4 KO samples and 4 controls. In vivo Trim28-KO during development: Emx1-Cre/Trim28-fl animals. Three KOs (homozygote for Trim28-fl) and three control (heterozygote for Trim28-fl). H3K9me3 cut and run: One sample in NPCs and its respective IgGs. Trim28-KO Single cell sequencing: Two KO and two controls from the Emx mice were sequenced in single cell.