RNA Dynamics in Aging Bacterial Spores. To view the full text, please login as a subscribed user or purchase a subscription. Click here to view the full text on ScienceDirect. Figure 1 rRNA Dynamics in Aging Spores (A–C) RNA was extracted from decoated B. subtilis (PY79) spores at the indicated age and temperature and analyzed in a microfluidic gel (see Experimental Procedures). Arrows designate the positions of 23S rRNA (upper) and 16S rRNA (lower). (D) qRT-PCR analysis was performed on cDNA derived from aging B. subtilis (PY79) spores incubated at the indicated temperatures (see Experimental Procedures).
(E) Bioanalyzer pseudogel of RNA extracted from decoated spores and vegetative cells of the WT strain (PY79) (see Experimental Procedures). See also Figure S1. Figure 2 Depletion of RNaseY Reduces Degradation of Spore RNA (B and C) qRT-PCR analysis of select transcripts in aging WT (PY79) spores (green) versus RNaseY-depleted spores (ES83: Pxylose-rny-cm) (purple) (see Experimental Procedures). See also Figure S2 and Table S1. ? Genome-wide Identification of Polycomb-Associated RNAs by RIP-seq. To view the full text, please login as a subscribed user or purchase a subscription. Click here to view the full text on ScienceDirect. Figure 1 The RIP-seq Technique and Analysis of Pilot Libraries (A) RIP-seq schematic.
(B) Western blot analysis (right panel) of Ezh2 protein in wild-type (WT) and Ezh2−/− ESCs. Coomassie staining (left panel) shows equal loading. (C) Preparatory agarose gel for RIP product size selection. (D) Pilot library statistics for WT and control libraries for an equivalent number of cells (column 2), reads after filtering using criteria shown in Figure S1 (column 3), and distinct reads after removing duplicates and repetitive elements (column 4). (E) CCs of indicated libraries in pairwise comparisons against the original WT library. (F) The cumulative frequency of WT reads mapping to elements with indicated genome copy numbers. (G) Relative frequencies of various repeats in the WT library. (H) Alignments of distinct WT pilot reads to the mouse X chromosome. Figure 2. Live Cell Imaging of Telomerase RNA Dynamics Reveals Cell Cycle-Dependent Clustering of Telomerase at Elongating Telomeres. To view the full text, please login as a subscribed user or purchase a subscription.
Click here to view the full text on ScienceDirect. Figure 1 A Green Fluorescent Reporter System to Visualize Telomerase RNA in Living Yeast Cells (A) Description of the MS2-GFP reporter system. (B) Fluorescent in situ hybridization on TLC1 RNA shows colocalization between TLC1-10xMS2 RNA and MS2-GFP foci. Scale bar: 1 μm. (C) Time-lapse images (taken from Movies S1, S2, and S3 at the indicated times) of cells in G1 (a–d); S phase (i–l); and G2 (q–t) expressing TLC1 RNA-GFP foci. Figure 2 Dynamics of TLC1 RNA-GFP Foci and Clusters (A) Comparison of TLC1 RNA-GFP particle dynamics in different phases of the cell cycle; a diffusion coefficient (D). (B) Comparison of the MSD of TLC1 RNA-GFP foci in G1 or G2, TLC1 RNA-GFP clusters in S phase, and TEL IVR-LacO foci. (C) Occurrence of TLC1 RNA-GFP clusters during cell-cycle phases. (D) Quantification of the number of TLC1 RNA-GFP cluster per late S phase cell. Figure 3. Single-Molecule Imaging of Transcriptionally Coupled and Uncoupled Splicing.
Open Archive Summary Introns are removed from pre-mRNAs during transcription while the pre-mRNA is still tethered to the gene locus via RNA polymerase. However, during alternative splicing, it is important that splicing be deferred until all of the exons and introns involved in the choice have been synthesized. We have developed an in situ RNA imaging method with single-molecule sensitivity to define the intracellular sites of splicing.
Using this approach, we found that the normally tight coupling between transcription and splicing is broken in situations where the intron's polypyrimidine tract is sequestered within strong secondary structures. We also found that in two cases of alternative splicing, in which certain exons are skipped due to the activity of the RNA-binding proteins Sxl and PTB, splicing is uncoupled from transcription. PaperClip <span class="MMCvInstallJS">To listen to this audio, enable JavaScript on your browser. Graphical Abstract Highlights Introduction Results Figure 1. No-Nonsense Functions for Long Noncoding RNAs.