Fig. 1.

Reporter mRNAs display directed dendritic transport in hippocampal neurons. a Scheme of both MS2 only and MS2+Rgs4 reporter constructs and tdMCP-GFP expression cassettes (upper) and the MS2 system (lower). pRSV Rous sarcoma virus promoter, pUBC Ubiquitin C promoter, ORF open reading frame, NLS nuclear localization signal, tdMCP tandem MS2 coat protein, UTR untranslated region. b Phase contrast, GFP fluorescence (reporter), MS2 single molecule FISH and overlay in a rat hippocampal neuron expressing both tdMCP-GFP and MS2+Rgs4 MS2 reporter mRNA (scheme). Arrowheads indicate overlapping tdMCP-GFP bound MS2 reporter mRNA and MS2 smFISH. Fluorescent images were deconvolved to assess overlap (for unprocessed images see Supplementary Fig. 1e). Scale bar 20 µm. Boxed region is magnified in right panels. Representative kymographs (left) and extracted tracks (right) illustrating differences in unidirectional MS2+Rgs4 3′-UTR mRNA granule transport speed, displacement and directionality (c), as well as interrupted (d) and multidirectional transport (e). Anterograde and retrograde transport are indicated in green or red arrowheads and lines, respectively. f Quantification of relative transport dynamics of MS2 only and MS2+Rgs4 3′-UTR reporter mRNAs in 1 and 10-min time-series acquisitions, respectively