Genome-wide screen reveals APC-associated RNAs enriched in cell protrusions

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Abstract
RNA localization is important in many biological processes that involve the establishment or maintenance of polarity. Previously there has been no comprehensive identification of RNAs that localize during the polarization of mammalian cells, but that has now been achieved in a study of fibroblasts responding to migratory stimuli. A genome-wide screen identified more than 50 RNAs that localize to cell protrusions extending from mouse fibroblasts. The RNAs are anchored in granules concentrated at the 'plus' ends of microtubules, a novel RNA anchoring mechanism and an unanticipated function for microtubule plus ends. RNAs in these granules are associated with the tumour suppressor protein adenomatous polyposis coli (APC), a multi-function protein extensively studied as a component in the Wnt signalling pathway, also thought to be involved in cell migration, cell adhesion and mitosis. The cover image shows RNA granules (blue) at the tip of a cell protrusion, which has also been stained for actin filaments. RNA localization is important in diverse biological process including establishment of polarity. This study has focused on migrating fibroblasts that polarize to form a leading edge and tail, in a process that involves assymetric distribution of RNAs. On a genome-wide scale RNAs that are localized to cell protrusions are identified. Through their 3' UTRs, these transcripts are anchored in granules concentrated at the plus end of microtubules and associate with the APC tumour suppressor. RNA localization is important for the establishment and maintenance of polarity in multiple cell types. Localized RNAs are usually transported along microtubules or actin filaments1 and become anchored at their destination to some underlying subcellular structure. Retention commonly involves actin or actin-associated proteins2,3,4,5,6,7, although cytokeratin filaments and dynein anchor certain RNAs8,9. RNA localization is important for diverse processes ranging from cell fate determination to synaptic plasticity; however, so far there have been few comprehensive studies of localized RNAs in mammalian cells. Here we have addressed this issue, focusing on migrating fibroblasts that polarize to form a leading edge and a tail in a process that involves asymmetric distribution of RNAs10,11,12. We used a fractionation scheme13 combined with microarrays to identify, on a genome-wide scale, RNAs that localize in protruding pseudopodia of mouse fibroblasts in response to migratory stimuli. We find that a diverse group of RNAs accumulates in such pseudopodial protrusions. Through their 3′ untranslated regions these transcripts are anchored in granules concentrated at the plus ends of detyrosinated microtubules. RNAs in the granules associate with the adenomatous polyposis coli (APC) tumour suppressor and the fragile X mental retardation protein (FMRP). APC is required for the accumulation of transcripts in protrusions. Our results suggest a new type of RNA anchoring mechanism as well as a new, unanticipated function for APC in localizing RNAs.