UNDERSTANDING GERM CELL REGULATION USING BIOINFORMATIC APPROACHES

Leanne Whitmore

Bioinformatics is a powerful tool which has been utilized to study diseases (i.e cancer) and complex biological systems including the immune system, neural interactions in the brain and testis biology. Only recently has bioinformatics been used to study the intricacies of spermatogenesis, the differentiation and development of spermatogonia into spermatozoa. Successful spermatogenesis is essential for male reproduction and understanding regulatory mechanisms necessary for ensuring production of sperm is vital for the development of male infertility treatments or reversible contraceptive options. This dissertation focuses on two regulatory mechanisms necessary for spermatogenesis in mice: 1. metabolic activity of spermatogonia and 2. piRNA control of long interspersed element-1 (L1) retrotransposon evolution. 1. The bioinformatics approach flux balance analysis (FBA) is used to model metabolic activity of spermatogonia at 8 different time points during the initial differentiation of spermatogonia. Because it is known that central and vitamin A metabolic activities are crucial for successful spermatogenesis we analyze the activities of these metabolic pathways and uncover the roles they play in regulating spermatogenesis. Additionally, reaction and enzyme in-silico knockouts, within the vitamin A metabolic pathway are performed to elucidate their importance in regulating RA availability, which is crucial for differentiation of spermatogonia. 2. Bioinformatics analysis of piRNA sequencing and transcriptome datasets from mouse testes was performed to discover if piRNA repression of L1s is directing the single lineage evolution of L1s. PiRNAs suppress L1 activity at the transcriptional and post-transcriptional stages of retrotransposition and are essential for male fertility. PiRNA and L1 expression of L1 families were analyzed to determine if there was more activity (of piRNAs or L1 transcripts) to a particular age (young vs old) of L1 family. Additionally, expression was further analyzed to understand if piRNA or L1 activity was dependent on location of L1s relative to genes. Finally, where piRNAs target L1s for silencing (5’ or 3’ end) was examined to elucidate if piRNAs are exerting selective pressure on the 5’ end thereby causing L1s to acquire a new 5’ UTR as they evolve. Overall, this thesis utilizes bioinformatics to understand spermatogenesis and advances our understanding of mammalian male reproduction.

UNDERSTANDING GERM CELL REGULATION USING BIOINFORMATIC APPROACHES

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