We report the mechanistic basis guiding the migration pattern of multiple nuclei 
in hyphae of Ashbya gossypii. Using electron tomography, we reconstructed the 
cytoplasmic microtubule (cMT) cytoskeleton in three tip regions with a total of 
13 nuclei and also the spindle microtubules of four mitotic nuclei. Each spindle 
pole body (SPB) nucleates three cMTs and most cMTs above a certain length grow 
according to their plus-end structure. Long cMTs closely align for several 
microns along the cortex, presumably marking regions where dynein generates 
pulling forces on nuclei. Close proximity between cMTs emanating from adjacent 
nuclei was not observed. The majority of nuclei carry duplicated side-by-side 
SPBs, which together emanate an average of six cMTs, in most cases in opposite 
orientation with respect to the hyphal growth axis. Such cMT arrays explain why 
many nuclei undergo short-range back and forth movements. Only occasionally do 
all six cMTs orient in one direction, a precondition for long-range nuclear 
bypassing. Following mitosis, daughter nuclei carry a single SPB with three 
cMTs. The increased probability that all three cMTs orient in one direction 
explains the high rate of nuclear bypassing observed in these nuclei. The A. 
gossypii mitotic spindle was found to be structurally similar to that of 
Saccharomyces cerevisiae in terms of nuclear microtubule (nMT) number, length 
distribution and three-dimensional organization even though the two organisms 
differ significantly in chromosome number. Our results suggest that two nMTs 
attach to each kinetochore in A. gossypii and not only one nMT like in S. 
cerevisiae.