The polarity and dynamics of microtubule assembly in the budding yeast Saccharomyces cerevisiae.
Microtubule assembly in Saccharomyces cerevisiae is initiated from sites within spindle pole bodies (SPBs) in the nuclear envelope. Microtubule plus ends are thought to be organized distal to the SPBs, while minus ends are proximal. Several hypotheses for the function of microtubule motor proteins in force generation and regulation of microtubule assembly propose that assembly and disassembly occur at minus ends as well as at plus ends. Here we analyse microtubule assembly relative to the SPBs in haploid yeast cells expressing green fluorescent protein fused to alpha-tubulin, a microtubule subunit. Throughout the cell cycle, analysis of fluorescent speckle marks on cytoplasmic astral microtubules reveals that there is no detectable assembly or disassembly at minus ends. After laser-photobleaching, metaphase spindles recover about 63% of the bleached fluorescence, with a half-life of about 1 minute. After anaphase onset, photobleached marks in the interpolar spindle are persistent and do not move relative to the SPBs. In late anaphase, the elongated spindles disassemble at the microtubule plus ends. These results show for astral and anaphase interpolar spindle microtubules, and possibly for metaphase spindle microtubules, that microtubule assembly and disassembly occur at plus, and not minus, ends.
Maddox
P S
PS
Department of Biology, CB3280, University of North Carolina, Chapel Hill, North Carolina 27599-3280, USA. pmaddox@email.unc.edu
Bloom
K S
KS
Salmon
E D
ED
D000705:Anaphase / Q000502:physiology; D003593:Cytoplasm / Q000378:metabolism; D016193:G1 Phase / Q000502:physiology; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D007202:Indicators and Reagents / Q000378:metabolism; D007834:Lasers; D008164:Luminescent Proteins / Q000235:genetics; D008677:Metaphase / Q000502:physiology; D008856:Microscopy, Fluorescence / Q000379:methods; D008870:Microtubules / Q000737:chemistry / Q000378:metabolism*; D008938:Mitosis / Q000502:physiology; D016196:S Phase / Q000502:physiology; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000235:genetics / Q000378:metabolism*; D008941:Spindle Apparatus / Q000502:physiology; D013692:Telophase / Q000502:physiology
D007202:Indicators and Reagents; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins