Saunders W, Hornack D, Lengyel V, Deng C
Department of Biological Sciences, University of Pittsburgh, Pennsylvania 15260, USA.
J Cell Biol. 1997 Apr 21;137(2):417-31. doi: 10.1083/jcb.137.2.417.
The Saccharomyces cerevisiae kinesin-related motor Kar3p, though known to be required for karyogamy, plays a poorly defined, nonessential role during vegetative growth. We have found evidence suggesting that Kar3p functions to limit the number and length of cytoplasmic microtubules in a cell cycle-specific manner. Deletion of KAR3 leads to a dramatic increase in cytoplasmic microtubules, a phenotype which is most pronounced from START through the onset of anaphase but less so during late anaphase in synchronized cultures. We have immunolocalized HA-tagged Kar3p to the spindle pole body region, and fittingly, Kar3p was not detected by late anaphase. A microtubule depolymerizing activity may be the major vegetative role for Kar3p. Addition of the microtubule polymerization inhibitors nocodazol or benomyl to the medium or deletion of the nonessential alpha-tubulin TUB3 gene can mostly correct the abnormal microtubule arrays and other growth defects of kar3 mutants, suggesting that these phenotypes result from excessive microtubule polymerization. Microtubule depolymerization may also be the mechanism by which Kar3p acts in opposition to the anaphase B motors Cin8p and Kip1p. A preanaphase spindle collapse phenotype of cin8 kip1 mutants, previously shown to involve Kar3p, is markedly delayed when microtubule depolymerization is inhibited by the tub2-150 mutation. These results suggest that the Kar3p motor may act to regulate the length and number of microtubules in the preanaphase spindle.
酿酒酵母中与驱动蛋白相关的马达蛋白Kar3p，虽然已知其在核融合过程中是必需的，但在营养生长期间发挥的作用定义不明确且并非必需。我们发现有证据表明，Kar3p以细胞周期特异性方式发挥作用，限制细胞质微管的数量和长度。KAR3基因的缺失导致细胞质微管显著增加，这种表型在同步培养物中从起始点到后期开始时最为明显，但在后期后期则不太明显。我们已将带有HA标签的Kar3p免疫定位到纺锤极体区域，并且恰当地，在后期后期未检测到Kar3p。微管解聚活性可能是Kar3p在营养生长中的主要作用。向培养基中添加微管聚合抑制剂诺考达唑或苯菌灵，或缺失非必需的α-微管蛋白TUB3基因，大多可以纠正kar3突变体异常的微管阵列和其他生长缺陷，这表明这些表型是由微管过度聚合导致的。微管解聚也可能是Kar3p与后期B马达蛋白Cin8p和Kip1p起相反作用的机制。cin8 kip1突变体的后期前纺锤体塌陷表型（先前已证明涉及Kar3p），在微管解聚被tub2 - 150突变抑制时会明显延迟。这些结果表明，Kar3p马达蛋白可能在调节后期前纺锤体中微管的长度和数量方面发挥作用。
