Journal: bioRxiv
Article Title: Phosphorylation of the rod-tail hinge region of cingulin regulates its interaction with nonmuscle myosin-2B
doi: 10.64898/2026.04.02.716052
Figure Lengend Snippet: Inhibition of either CK1 or CK2 promotes the rescue of junctional NM2B in cingulin-KO cells expressing the depho-6 mutant of cingulin. (A) Top: scheme of GFP-tagged canine CGN (cCGN-FL), with the GFP tag (green), globular head (gray), coiled-coil rod (white) and globular tail (blue) domains (amino-acid residue boundaries are indicated above the scheme). Bottom: C-terminal sequences of WT canine CGN (cCGN-FL, sequence 1140–1190 with specific residues indicated in the region) and corresponding sequences of the cCGN-dephosphomimetic-6 mutant (depho-6), which does not rescue junctional NM2B (Fig. S1K’). (B-G) IF microscopy analysis and localization (left) and quantification of junctional labeling (relative fluorescence intensity) (right) of NM2B in CGN-KO MDCK cells rescued with GFP-cCGN-FL (B, D and F), or with GFP-cCGN-depho-6 mutant (C, E and G) treated either with DMSO (B and C) or with CK1 inhibitor (D and E: 25 μM, 8h) or with CK2 inhibitor (F and G: 25 μM, 14h). Arrows and arrowheads show increased, normal and decreased/undetected junctional labeling, respectively. Quantifications of relative fluorescent intensity (RFI) shows the ratio between the junctional staining of NM2B versus the junctional marker PLEKHA6 (n=70 junctions) from three independent experiments. Data in quantifications are represented as mean±SD. Statistical significance was determined by unpaired Mann-Whitney’s test. ***p≤0.001. Scale bar (G)= 10 μm.
Article Snippet: Drugs treatments were as follows (final concentration, duration, catalog number and source): CK1 inhibitor (25 μM, 8 h, D4476 MedChemExpress) and CK2 inhibitor (5 μM, 14 h, CX4945 Selleckchem).
Techniques: Inhibition, Expressing, Mutagenesis, Residue, Sequencing, Microscopy, Labeling, Fluorescence, Staining, Marker
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