Control of gene expression is regulated at many stages. One aspect of this regulation, the localization of specific mRNA transcripts to distinct cellular locations, has only recently been fully appreciated. Correct targeting of these messages can be crucial for many cellular and higher-order processes. For example, disruption of dendritic targeting of specific messages in mice leads to a decrease in long-term potentiation, implying that this process may be important for learning and memory. Recent studies have shown that up to two thousand mRNAs are localized to axons in neuronal cells. However, little is known about how messages are tagged for transport or the factors that mediate it. Although the mechanism of localization of a handful of messages has been dissected, how such mechanisms may operate on a transcriptome-wide scale is unknown. The studies outlined in this proposal aim to define the cis-acting sequences and trans-acting factors responsible for the localization of many messages. Specifically, I will address this question using a combination of RNA biochemistry, molecular genetics, and computational genomics. Using RNA-seq from axons and cell bodies, I will define the RNA sequences necessary for localization of specific messages. I will then refine these sequences using an MS2-GFP reporter system and search the localized transcriptome both computationally and experimentally for other instances of these motifs, whether they be contained in primary or secondary RNA structure. The factors that bind these motifs will then be identified biochemically and their effects on the localization of other messages will be identified using RNAi and localized RNA-seq. The results of these studies will allow the identification of families of messages that use similar motifs and factors for their localization, as well as the prediction of motif sequences in other messages. This has important clinical and therapeutic implications as mutations in many RNA-localizing proteins are associated with several neuronal diseases, including spinomuscular atrophy and ALS.
Misregulation of RNA localization is implicated in many neuronal diseases. A more complete understanding of the sequences and factors mediating localization is required for understanding their molecular mechanisms and their potential for treatment.
