Validation and Characterisation of Novel miRNAs Identified in the E15.5 Mouse Brain
MicroRNAs (miRNAs) are small noncoding elements that downregulate gene expression and play an important role in maintaining robust neurodevelopment. Since abnormal expression of miRNAs has been associated with disorders including schizophrenia and autism spectrum disorder, characterising miRNAs is of growing biomedical relevance. However, individual miRNA expression levels can vary considerably between species, tissue types and even within the same tissue throughout developmental timepoints. Consequently, many potentially important developmental miRNAs remain undiscovered due to their transient expression. Thus, identification and characterisation of more miRNAs may contribute important insights to the current knowledge base surrounding the function of miRNAs in neurodevelopment. Previously, our lab has identified several novel miRNAs by small RNA sequencing of the E15.5 mouse brain. The aim of this project was to combine bioinformatic and experimental techniques in order to validate and functionally characterise a selection of these novel miRNAs. Firstly, interspecies conservation of these novel miRNAs was analysed using both miRbase and the BLAT genome browser, which revealed 4 of the 10 miRNAs analysed are conserved throughout vertebrates, while the others appear to be mouse specific. Subsequently, miRNA expression levels in various mouse tissues was determined using real-time quantitative reverse transcriptase PCR (RT-qPCR), demonstrating that each miRNA analysed has tissue- and time-specific expression levels. Interestingly, three of the novel miRNAs have significantly different expression levels between male and female E15.5 brains, highlighting the importance of considering sex differences when classifying miRNAs. Further investigation of the expression patterns of a portion of these miRNAs by in situ hybridisation uncovered some of the miRNAs may have atypical biogenesis. Lastly, to gain further understanding of the function of these novel miRNAs, their mRNA targets were bioinformatically predicted. Gene ontology analysis of predicted targets suggested that many of the miRNAs target mRNAs involved in neurodevelopment and neural functioning. This project validates and characterises a selection of the novel miRNAs identified and provides evidence to suggest that some have roles in regulating neurodevelopment within mice.
Advisor: Wilson, Megan
Degree Name: Bachelor of Biomedical Sciences with Honours
Degree Discipline: Anatomy
Publisher: University of Otago
Keywords: miRNA; microRNA; neurodevelopment; sex bias; mouse
Research Type: Thesis