Fig. 4

Fig. 4 CITK regulates acetylated tubulin levels and co-immunoprecipitates with HDAC6.

a Western blot analysis of total lysate from ONS-76 48 h after transfection of the indicated siRNA. The levels of CITK and acetylated tubulin (ac-TUB) were analyzed, and the internal loading control was α-tubulin (α-TUB). b Quantification of the relative density of ac-TUB in samples described in (a). c Schematic diagram of CITK protein domains (Kinase domain; Coiled coil, Rho-binding domain (Rho BD); Zinc finger (ZF), pleckstrin-homology domain (PH), Citron–Nik1 homology domain (CNH), PDZ domain) and of the different constructs used in the following panels. d Western blot analysis of total cell lysates (left) or RFP-Trap-based immunoprecipitations (right), obtained from 293-T cells transfected with constructs expressing Cherry-tagged CITK or with empty control vecto. The levels of RFP, HDAC6, and CYLD were analyzed. e Western blot analysis of total cell lysates (left) or anti-Myc immunoprecipitations (right), obtained from 293-T cells transfected with vectors expressing the indicated Myc-tagged CITK proteins. The levels of Myc, HDAC6 and CYLD were analyzed. f Western blot analysis of total lysate from ONS-76 48 h after transfection with the indicated siRNAs. The levels of CITK and HDAC6 were analyzed, and the internal loading control was Vinculin (VINC). Quantification of HDAC6 levels in siCITK cells relative to siCtrl. Nuclear (g) and cytoplasmic (h) fractionation of ONS-76 48 h after siRNA transfection. The levels of CITK and HDAC6 were analyzed, and the internal loading control was lamin A/C (LAM A/C) for the nucleus and α-tubulin (α-TUB) for the cytoplasm. Each dot indicates an independent biological replicate. Error bars, SEM. *P < 0.05, **P < 0.01. unpaired two-tailed Student’s t-test. A.U. arbitrary unit.