Bioinformatic analysis predicts that ethanol exposure during early development causes alternative splicing alterations of genes involved in RNA post-transcriptional regulation

Abstract

Prenatal ethanol exposure is associated with neurodevelopmental defects and long-lasting cognitive deficits, which are grouped as fetal alcohol spectrum disorders (FASD). The molecular mechanisms underlying FASD are incompletely characterized. Alternative splicing, including the insertion of microexons (exons of less than 30 nucleotides in length), is highly prevalent in the nervous system. However, whether ethanol exposure can have acute or chronic deleterious effects in this process is poorly understood. In this work, we used the bioinformatic tools VAST-TOOLS, rMATS, MAJIQ, and MicroExonator to predict alternative splicing events affected by ethanol from available RNA sequencing data. Experimental protocols of ethanol exposure included human cortical tissue development, human embryoid body differentiation, and mouse development. We found common genes with predicted differential alternative splicing using distinct bioinformatic tools in different experimental designs. Notably, Gene Ontology and KEGG analysis revealed that the alternative splicing of genes related to RNA processing and protein synthesis was commonly affected in the different ethanol exposure schemes. In addition, the inclusion of microexons was also affected by ethanol. This bioinformatic analysis provides a reliable list of candidate genes whose splicing is affected by ethanol during nervous system development. Furthermore, our results suggest that ethanol particularly modifies the alternative splicing of genes related to post-transcriptional regulation, which probably affects neuronal proteome complexity and brain function.

Effects of In Utero EtOH Exposure on 18S Ribosomal RNA Processing: Contribution to Fetal Alcohol Spectrum Disorder

“…The effects of prenatal alcohol exposure on early fetal development are difficult to determine in utero using current imaging technology. Thus, our study selected a biomarker that could be detected early in fetuses during the vulnerable developmental period between three and six weeks after fertilization [ 49 , 50 ], when EtOH can damage the cranial neural crest cells that are critical for development of facial features, and when rRNAs are involved in a marked reduction of cranial neural crest cell proliferation and survival, impaired migration, and increased apoptosis [ 19 , 22 ]. Here, we performed developmental studies on sr18S-RNA in human brain using ddPCR, and demonstrated: (i) developmental regulation of srRNAs in human fetal brain ( Figure 1 A) and in FB-Es ( Figure 1 B); (ii) the effects of in utero EtOH exposure on srRNA in developing brain ( Figure 1 and Figure 3 ); (iii) the association of MBP gene-binding protein, Purα, with 18S ribosomal RNA, which results in its cleavage, with the formation of small RNAs (smRNA) ( Figure 2 , Figure 4 and Figure 5 ); (iv) the role of Purα and 18S ribosomal RNA in viral–host interaction ( Figure 6 ); and (v) DNase and RNase nuclease activity and subsequent unwinding activity of the MBP gene-binding Purα ( Figure 5 and Figure 7 ).…”

“…The mechanism by which EtOH affects RNAs is unknown, but recently, it was suggested that EtOH modifies the alternative splicing of genes related to post-transcriptional regulation, which likely affects neuronal proteome complexity and brain function [ 22 ].…”

“…Some rRNA processing and translation in mammalian cells was assayed using a synthetic 18S rRNA expression system [ 21 ]. Bioinformatic analysis predicted that EtOH exposure during early development would cause alternative splicing alterations of genes involved in RNA post-transcriptional regulation [ 22 ].…”