Transfer RNA Dynamics During Neural Differentiation - Transfer RNA Dynamics During Neural Differentiation Far from serving as invariant mRNA decoders, transfer RNAs (tRNAs) play a dynamic role in shaping gene expression. For example, tRNA abundance is expected to affect mRNA stability: transcripts enriched in optimal codons (decoded by abundant tRNAs) are stable while transcripts enriched in non-optimal codons (decoded by rare tRNAs) are unstable. We will test this model using differentiation of Drosophila neural stem cells (neuroblasts) as a highly tractable system with relevance to human neurodevelopment and neurological disease. We hypothesize that tRNA levels differ between neuroblasts and their neuronal progeny, thereby influencing mRNA stability and fine-tuning gene expression to meet the needs of each cell type. We also hypothesize that tRNA gene (tDNA) transcription during neural differentiation is regulated by two non-exclusive mechanisms: widespread repression of RNA polymerase III (RPOL3) by a neural-specific inhibitor and more targeted repression of RPOL3 due to overlapping RNA polymerase II (RPOL2)-dependent transcription. We will test these hypotheses using quantification of tRNA abundance, tRNA transcription and mRNA decay in neuroblasts and neurons, combined with pharmacologic and genetic manipulation of predicted regulators of tDNA transcription. We will determine the functional significance of differential tRNA expression using targeted tRNA knockdown and overexpression in assays of mRNA decay, neurodevelopment and behavior. This work will be made possible through the development of novel tools for altering tRNA expression, including a CRISPR / Cas9-based approach. Defective tRNA expression is implicated in several neurodevelopmental and neurological diseases, including neurodegenerative diseases, multiple forms of microcephaly, and brain cancers. We will use our expertise in Drosophila neurobiology and RNA biology to significantly advance the understanding of tRNA expression dynamics that regulate normal development and, when defective, contribute to disease.
