Microtubule nucleation from the centrosome is reduced in the differentiation process.

<p>(A) Deconvolved optical sections of differentiated muscle cells after re-growth of microtubules for 20 seconds, stained for immunofluorescence of (left) microtubules, (middle) the centrosome protein pericentrin, and (right) stained with the DNA marker DAPI. In a myotube early after fusion, pericentrin is visible both around the nuclear surface and on the centrosomes (arrows). Microtubule re-growth is seen from these centrosomes, in addition to perinuclear and cytoplasmic sites. (B) Myotube, after cold-induced depolymerization of microtubules and re-growth for 10 seconds. Immunofluorescence of the plus-end-binding protein EB3 and of pericentrin are shown. Cytoplasmic, nuclear, and centrosomal sites of emerging EB3 comets are indicated on the left. The table indicates the percentage of EB3 comets growing from cytoplasmic, nuclear, and centrosomal locations in undifferentiated cells (n = 6), as well as in differentiating cells before fusion (mononucleate, n = 15), and after fusion (n = 12). The total number of EB3 comets per cell ranged from 39 (undifferentiated) to 1151 (differentiated, fused). (C) Left: nucleus of a myoblast at an early stage of differentiation, stained for pericentrin (top) and microtubules (bottom), recovering for 10 seconds from cold-induced depolymerization. The number of microtubules grown from the nuclear surface was quantified by counting EB3 comets as in (B). The results for each nucleus were plotted against the intensity of perinuclear pericentrin (arbitrary values, after subtraction of background) in undifferentiated cells (n = 6), and in mononucleate (n = 15) and fused differentiated cells (n = 22). Horizontal and vertical bars indicate standard deviations of pericentrin intensity and of the number of EB3 comets, respectively, in differentiated mononucleate and fused cells. Bar in (A), 10 µm; identical magnification in (B), (C).