Extended Data Fig. 7: Modulating tubulin glutamylation affects ER distribution.
From: ER proteins decipher the tubulin code to regulate organelle distribution
a, Representative images of U2OS and COS7 cells sequentially immunolabeled for polyE (polyglutamylation), polyE plus GT335 (glutamylation) and Alexa 647 conjugated α-tubulin. GT335 signal indicates monoglutamylation in this case because polyglutamylation is pre-saturated with polyE antibody. b, Quantifications of signal distributions for cells as in (a). Data points from the same cell are linked by solid lines. n = 20 cells for U2OS, 35 cells for COS7. c, Western blot analysis of U2OS, COS7, HeLa and RPE1 cells. d, MDR ratios of monoE (indicated by GT335 labeling) to polyE, or microtubules (indicated by α-tubulin labeling) to polyE. n = 20 cells for U2OS, 35 cells for COS7. e, Representative images of U2OS cells overexpressing the indicated TTLLs or CCPs. f, g, Quantifications of ER and microtubule MDRs of cells as in e; n = 37, 40, 34, 40, 40, 41, 39 cells (left to right) for (f); n = 29, 38, 37, 51, 38, 46, 39 cells for (g). h, Quantifications of ER MDR of cells in p180/KTN1 double knockout cells overexpressing the indicated TTLLs or CCPs; n = 63, 61, 64, 63, 61 cells (left to right). i, Quantifications of ER MDR of wild-type or KTN1 knockout cells overexpressing TTLL7; n = 40, 64 cells (left to right). j, U2OS cells transfected with the indicated siRNAs for 72 h were analyzed by western blotting. k, U2OS cells transfected with the indicated siRNAs for 48 h were analyzed by real time PCR. n = 4 repeats. l, m, Representative image and quantification of ER in control or CCP5 siRNA transfected cells. Scale bars, 10 μm. n = 64, 70, 69, 39, 39, 39 cells (left to right). Data are mean ± s.d. P values shown on top, two-tailed t-test. See Supplementary Information for uncropped western blots.
