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Circular RNAs (circRNAs) are a novel category of covalently-closed non-coding RNAs mainly derived from the back-splicing of exons or introns of protein-coding genes. In addition to their inherent high overall stability, circRNAs, have been shown to have strong functional effects on gene expression via a multitude of transcriptional and post-transcriptional mechanisms. Furthermore, circRNAs, appear to be particularly enriched in the brain and able to influence both prenatal development and postnatal brain function. However, little is known about the potential involvement of circRNAs in the long term influence of prenatal alcohol exposure (PAE) in the brain and their relevance for Fetal Alcohol Spectrum Disorders (FASD). Using circRNA-specific quantification, we have found that circHomer1 , an activity-dependent circRNA derived from Homer protein homolog 1 ( Homer1 ) and enriched in postnatal brain, is significantly down-regulated in the male frontal cortex and hippocampus of mice subjected to modest PAE. Our data further suggest that the expression of H19 , an imprinted embryonic brain-enriched long non-coding RNA (lncRNA), is significantly up-regulated in the frontal cortex of male PAE mice. Furthermore, we show opposing changes in the developmental- and brain region specific- expression of circHomer1 and H19 . Lastly, we show that knockdown of H19 results in robust increases in circHomer1 but not linear HOMER1 mRNA expression in human glioblastoma cell lines. Taken together, our work uncovers notable sex- and brain region-specific alterations in circRNA and lncRNA expression following PAE and introduces novel mechanistic insights with potential relevance to FASD., Competing Interests: NM has a financial interest in Circular Genomics Inc., a company focused on the use of circRNAs as biomarkers for psychiatric disorders. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Papageorgiou, Amoah, Pierotti, Otero, Eckel, Coffey, Allan, Caldwell and Mellios.)

Fetal alcohol spectrum disorders (FASD) are heterogeneous disorders associated with alcohol exposure to the developing fetus that are characterized by a range of adverse neurodevelopmental deficits. Despite the numerous genomics and genetic studies on FASD models, the comprehensive molecular understanding of the mechanisms that underlie FASD-related neurodevelopmental deficits remains elusive. Circular RNAs (circRNAs) are a subtype of long non-coding RNAs that are derived from back-splicing and covalent joining of exons and/or introns of protein-coding genes. Recent studies have shown that circRNAs are highly enriched in the brain, where they are developmentally regulated. However, the role of the majority of brain-enriched circRNAs in normal and pathological brain development and function has not been explored yet. Here we carried out the first systematic profiling of circRNA expression in response to prenatal alcohol exposure (PAE) in male and female embryonic day 18 (E18) whole brains. We observed that the changes in circRNA expression in response to PAE were notably sex-specific and that PAE tended to erase most of the sex-specificity in circRNA expression present in control (saccharin-treated) mice. On the other hand, RNA sequencing (RNA-seq) in the same samples showed that changes in protein-coding gene expression were not predominantly sex-specific. Using circRNA quantitative real-time PCR (qRT-PCR), we validated that circSatb2 , which is generated from the special AT-rich sequence-binding protein 2 ( Satb2 ) gene, is significantly upregulated in the brain of E18 male PAE mice. We also show that circPtchd2 , a circRNA synthesized from dispatched RND transporter family member 3 ( Disp3 , also known as Ptchd2 ), exhibits significantly higher expression in E18 control but not PAE female mouse brain relative to males. Taken together, our results demonstrate that PAE differentially alters circRNA expression in the developing brain in a sex-specific manner., (Copyright © 2020 Paudel, Pierotti, Lozano, Amoah, Gardiner, Caldwell, Allan and Mellios.)