Fig 3

Fig 3

(A) smFISH images of a single optical section of a myelin sheath in a 3-dpf larva spinal cord. mbpa transcripts line the myelin sheath. Arrows highlight clusters of mbpa mRNA transcripts. (B) smFISH images of a single optical section of myelin tracts in the hindbrain of a 5-dpf larva. Boxed area magnified to highlight sheath termini (arrows). (C) smFISH images of a single optical section in transverse plane of myelin sheaths in a 5-dpf larva midbrain. Scale bars (A, B, and D), 5 μm; (C, boxed enlargements), 1 μm. (D) Representative images from MS2 system showing colocalization of mRNA containing mbpa 3′ UTR and F-actin in a myelinating oligodendrocyte. Asterisk marks the cell body, and boxes are magnified to highlight sheath termini. Arrows highlight sheaths with mRNA, and arrowheads highlight sheaths lacking mRNA. (E) Proportion of sheaths with mRNA enriched in sheath termini at 4 dpf using the MS2 system. Proportion measured as (sheaths with enrichment / number of sheaths) = 10/35 sv40, 18/38 mbpa. (F) Average fluorescence intensity of MS2 mRNA reporter containing the sv40 or mbpa 3′ UTRs across a 7-μm distance, at 0.2-μm intervals, from myelin sheath termini at 4 dpf. Each line scan was normalized to the average fluorescent intensity per sheath. All normalized values for each distance were then averaged. Shaded area represents 95% confidence interval. Statistical significance was evaluated every 0.2 μm using Wilcoxon test, and the distance between 0.8–1.0 μm was statistically significant (blue line). sv40 3′ UTR n = 5 larvae, 35 sheaths. mbpa 3′ UTR n = 6 larvae, 38 sheaths. The underlying numerical data can be found in S1 and S2 Data. 3' UTR, 3' untranslated region; dpf, days post fertilization; F-actin, filamentous actin; smFISH, single molecule fluorescent in situ hybridization.