Fig. 5: Intron-retained nuclear TUG1 and TERT are long-lived transcripts, stably retained in the nucleus.
From: Nuclear compartmentalization of TERT mRNA and TUG1 lncRNA is driven by intron retention
a Relative stability of TUG1 and TERT exons and introns compared to GAPDH mRNA measured by RT-qPCR of random-primed cDNA from iPS, HEK293T and LN-18 cells during a 4.5 h ActD time course. GAPDH intron 2, a control for an efficiently spliced intron; 45S rRNA, a control for a precursor RNA. Each dot represents mean value from two or three replicates. b Relative abundance and stability of spliced and intron-retained transcripts in cDNA synthesized with random primers or oligo(dT) during the 4.5 h ActD treatment of iPS cells; bars, means across replicates; dots, individual replicates, n = two or three measurements. c Maximum intensity projection of LN-18 smRNA FISH targeting TUG1 exon (gray) and intron 1 (magenta) or intron 2 (magenta) at time point 0 (NT) and 4.5 h after ActD treatment. Scale bar, 5 μm. Below, smRNA FISH quantification at each time point of spliced and unspliced TUG1 transcripts in the nucleus and cytoplasm; PIR of intron 1 and intron 2 at each time point. n.s. = not significant, *P ≤ 0.05, ***P ≤ 0.001, evaluated by unpaired two-tailed t-test (equal variances) versus NT; n (nuclear TUG1, cytoplasmic TUG1, nuclear PIR intron 1) = 44 cells (NT, 40 min, 4.5 h), 43 cells (2.5 h); n (nuclear PIR intron 2) = 38 cells (NT), 39 cells (40 min, 4.5 h), 40 cells (2.5 h), two independent RNA FISH stainings. d Maximum intensity projection of LN-18 smRNA FISH targeting TERT exon (gray) and intron 11 (magenta) at time point 0 (NT) and 4.5 h after ActD treatment. Scale bar, 5 μm. Below, smRNA FISH quantification at each time point of spliced and unspliced TERT transcripts in the nucleus and cytoplasm; nuclear PIR of intron 11 at each time point. n.s. = not significant, ***P ≤ 0.001, evaluated by unpaired two-tailed t-test (equal variances) versus NT, n (nuclear, cytoplasmic TERT, nuclear PIR intron 11) = 32 cells (NT), 30 cells (40 min, 2.5 h, 4.5 h), two independent RNA FISH stainings. In c, d midline line, median; lower and upper box limits, 25th and 75th percentiles; whiskers, 1.5 times interquartile range from the 25th and 75th percentiles.
