Spindle microtubule organization requires the CT-35 of TPX2. (A) Endogenous TPX2 and Eg5 colocalize on spindle microtubules in LLC-Pk1 cells. Immunofluorescence images with anti-TPX2 and -Eg5 antibodies are shown. DNA was stained with DAPI. (B) Western blots of extracts of the cell lines used in these experiments. The blot was stained with anti-TPX2, which recognizes both pig and mouse TPX2. None, no transgene. Control (Con) or siRNA-targeting pig TPX2 is shown. (C) mTPX2^LAP, but not mTPX2-710^LAP, rescues mitotic index after siRNA to deplete pig TPX2. (D) Spindle microtubule organization in parental cells (top row) and in cells expressing the indicated transgene and treated with siRNA-targeting pig TPX2. Cells were stained for γ- and α-tubulin. (E) Quantification of the spindle phenotype in fixed parental cells and cells expressing the indicated transgene and treated with pTPX2 siRNA. (C and E) For each condition, n = 400; error bars represent SD. Bars, 5 µm.