Journal: bioRxiv
Article Title: The Unkempt RNA binding protein reveals a local translation program in centriole overduplication
doi: 10.1101/2024.07.29.605660
Figure Lengend Snippet: (A) Plk4 overexpression promotes centriole overduplication. Structured Illumination Microscopy (SIM) images of RPE-1 cells with endogenous and overexpressed Plk4. Plk4, magenta and centrioles, green. Scale bar 0.5 μm. (B) Unk is elevated upon Plk4 overexpression. Left panel – MA plot of mRNA levels based on RNA seq of RPE-1 cells with overexpressed Plk4, 16 hours after overexpression. PLK4 and UNK transcripts are elevated by 1.5- and 1.4-fold, respectively. Right panel – frequencies of PLK4 and UNK mRNAs in RPE-1 cells with endogenous and overexpressed Plk4. Graph values expressed as the means of biological replicates and SD. P values were determined using one-way ANOVA with Šídák post hoc test. (C) Unk protein is elevated at the centrosome by Plk4 overexpression. Left panels – SIM images of endogenous Unk in RPE-1 cells with endogenous and overexpressed Plk4. Unk, magenta and centrioles, green. Scale bar, 5 μm. Insets scale bar, 1 μm. Middle panels – 8 μm radial fluorescence intensity and corresponding ratio quantification of Unk using Centrin as the centrosome centroid. Right panel – centrosomal Unk fluorescence intensity based on binned central 2 μm. Graph values expressed as the means of biological replicates and SD. P value was determined using an unpaired two-tailed t test. (D) Unk is required for Plk4-induced centriole overduplication. Left panels – SIM images of RPE-1 cells showing centrioles in Unk depleted cells with endogenous and overexpressed Plk4. Unk, magenta and centrioles, green. Scale bar, 0.5 μm. Middle panel – frequency of cells with centriole overduplication. Right panel – number of cells with greater than, equal to, and less than four centrioles. Graph values expressed as the means of biological replicates and SD. P values were determined using one-way ANOVA with Šídák post hoc test.
Article Snippet: Libraries were constructed using the Nugen Universal Plus mRNA-SEQ library construction kit (Nugen 0508) and sequenced on an Illumina NovaSEQ 6000 sequencer by the Genomics and Microarray shared resource at the University of Colorado Cancer Center.
Techniques: Over Expression, Microscopy, RNA Sequencing Assay, Fluorescence, Two Tailed Test
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