Journal: bioRxiv
Article Title: A DIFFERENTIAL REQUIREMENT FOR THE CILIOPATHY GENE RPGRIP1L IN HUMAN AND MOUSE SPINAL PROGENITOR FATE SPECIFICATION
doi: 10.1101/2024.02.28.582477
Figure Lengend Snippet: (A) Analysis of SHH target genes GLI1 and PTCH1 . Log2(tpm+1) data from bulk RNASeq analysis are displayed as mean ± SEM (WT: n=4; KO: n=3). (B, C) Immunofluorescence analysis of ciliary proteins on WT and RPGRIP1L-deficient spinal organoids at day 6 of differentiation. (B) Cilia are labeled by ARL13B in red and gamma-TUBULIN in grey. Cilia are present at apical sites pointing into inner cavities of the organoids. The scale bar represents a length of 300 µm. (C) Cilia are labeled by IFT81 in green. The scale bar represents 1 µm. The graph shows unaltered ciliary length in RGPRIP1L -/- organoids compared to WT. Data are shown as mean ± SEM (WT: n=4; KO: n=4). (D) Stimulated-Emission-Depletion (STED) images of WT and RPGRIP1L-deficient cilia at day 6 of differentiation. Cilia are labeled by ARL13B in green. RPGRIP1L is marked in red. The scale bar represents 1 µm. Quantification of RPGRIP1L was measured on confocal images. Data are shown as mean ± SEM. Asterisks denote statistical significance according to unpaired t tests with Welch′s correction (***P < 0.001) (WT: n=4; KO: n=4). (E) Immunofluorescence of WT and RPGRIP1L- deficient cilia on spinal organoids at day 6 of differentiation. Cilia are labeled by ARL13B in red and INPP5E in green. The basal body is labeled by gamma-TUBULIN in grey. The scale bar represents 1 µM. Quantifications show an unaltered amount of INPP5E but a decreased amount of ARL13B in RPGRIP1L-deficient cilia. Data are shown as mean ± SEM. Asterisks denote statistical significance according to unpaired t tests with Welch′s correction (***P < 0.001) (WT: n=4; KO: n=4). (F) Immunofluorescence of WT and RPGRIP1L-deficient cilia on spinal organoids at day 6 of differentiation. Cilia are labeled by ARL13B in green and AC3 in red. The basal body is labeled by gamma-TUBULIN in grey. The scale bar represents 1 µm. Quantifications show a decreased amount of AC3 in RPGRIP1L-deficient cilia. Data are shown as mean ± SEM. Asterisks denote statistical significance according to unpaired t tests with Welch′s correction (***P < 0.001) (WT: n=4; KO: n=4).
Article Snippet: STED images were acquired on the STEDYCON confocal and STED module (Abberior Instruments GmbH, Germany) connected to the lateral port of an IX83 inverted microscope (Evident Scientific).
Techniques: Immunofluorescence, Labeling
