A Genome-Wide Screen for Dendritically Localized RNAs Identifies Genes Required for Dendrite Morphogenesis

Abstract

Localizing messenger RNAs at specific subcellular sites is a conserved mechanism for targeting the synthesis of cytoplasmic proteins to distinct subcellular domains, thereby generating the asymmetric protein distributions necessary for cellular and developmental polarity. However, the full range of transcripts that are asymmetrically distributed in specialized cell types, and the significance of their localization, especially in the nervous system, are not known. We used the EP-MS2 method, which combines EP transposon insertion with the MS2/MCP in vivo fluorescent labeling system, to screen for novel localized transcripts in polarized cells, focusing on the highly branched Drosophila class IV dendritic arborization neurons. Of a total of 541 lines screened, we identified 55 EP-MS2 insertions producing transcripts that were enriched in neuronal processes, particularly in dendrites. The 47 genes identified by these insertions encode molecularly diverse proteins, and are enriched for genes that function in neuronal development and physiology. RNAi-mediated knockdown confirmed roles for many of the candidate genes in dendrite morphogenesis. We propose that the transport of mRNAs encoded by these genes into the dendrites allows their expression to be regulated on a local scale during the dynamic developmental processes of dendrite outgrowth, branching, and/or remodeling.

Recent advances in branching mechanisms underlying neuronal morphogenesis

“…mTORC2, Src, and mRNA binding-dependent S181 phosphorylation of ZBP1 is essential for ZBP1 mobility and distribution along dendrites and regulation of dendritic branching in hippocampal neurons 133 . An unbiased genome-wide screen identified 55 new candidate mRNA transcripts that localize specifically to dendrites of Drosophila class IV dendritic arborization sensory neurons 134 . Further validation identified 18 genes whose mRNA is transported specifically to dendrites for local translation and regulation of branching.…”

Enclosure of Dendrites by Epidermal Cells Restricts Branching and Permits Coordinated Development of Spatially Overlapping Sensory Neurons

“…Additionally, a missense mutation in the human homologue EPB41LI is associated with severe non-syndromic intellectual disability, suggesting a specific role for EPB41LI in neuronal function (Hamdan et al, 2011). Intriguingly, cora mRNA localizes to dendrites of C4da neurons, and knockdown of cora expression results in increased dendrite branching (Misra et al, 2016). …”

Drosophila as a Model for Assessing the Function of RNA-Binding Proteins during Neurogenesis and Neurological Disease

“…The fact that almost all of these candidates are conserved across metazoa suggests that RNA regulatory mechanisms are critical to the development and function of the nervous system. This idea is also supported by the fact that a large number of RNAs are localized to neurites in many different species [ 96 , 97 , 98 , 99 , 100 , 101 , 102 , 103 , 104 ]. Importantly, the ability to visualize specific RNA transcripts in vivo, using the MS2/MCP in vivo fluorescent labeling system, makes Drosophila an ideal model for investigating the regulation and dynamics of mRNA localization within neurons [ 90 , 91 , 96 ].…”

“…This strategy identified 55 dendritically localized transcripts. Moreover, knockdown of many of the genes encoding these dendritically localized transcripts resulted in dendritic defects, underscoring the utility of this unbiased screening approach to identify new players in dendrite morphogenesis [ 96 ].…”