The Surveillance Mechanism of the Spindle Position Checkpoint in YeastJ Neil R. Adames, Jessica R. Oberle, and John A. Cooper Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110 Abstract. The spindle position checkpoint in Saccharo- myces cerevisiae delays mitotic exit until the spindle has moved into the mother–bud neck, ensuring that each daughter cell inherits a nucleus. The small G protein Tem1p is critical in promoting mitotic exit and is con- centrated at the spindle pole destined for the bud. The presumed nucleotide exchange factor for Tem1p, Lte1p, is concentrated in the bud. These ﬁndings suggested the hypothesis that movement of the spindle pole through the neck allows Tem1p to interact with Lte1p, promot- ing GTP loading of Tem1p and mitotic exit. However, we report that deletion of LTE1 had little effect on the timing of mitotic exit. We also examined several mu- Introduction During cell division in eukaryotes, the mitotic spindle sep- arates each duplicated pair of chromosomes to opposite poles of the cell. Cytokinesis then occurs between the poles, ensuring that each cell receives a full complement of chromosomes. In most eukaryotes, the position of the cleavage plane is determined by the position of the spindle (Oegema and Mitchison, 1997). In budding yeasts, such as Saccharomyces cerevisiae, cell division occurs at the mother–bud neck. Therefore, budding yeast must move the spindle into the neck before dividing. S. cerevisiae has two partially redundant mechanisms to move the spindle into the neck (Heil-Chapdelaine et al., 1999). A major mechanism for spindle movement into the neck is dynein-dependent sliding of microtubule sides along the bud cortex (Yeh et al., 1995; Adames and Cooper, 2000). A backup mechanism for spindle movement into the neck involves capture of microtubule ends at the bud cortex by Kar9p (Miller and Rose, 1998) and subsequent microtu- bule shrinkage to pull the nucleus toward the bud and help orient the spindle (Shaw et al., 1997; Adames and Cooper, 2000). Both mechanisms require cytoplasmic microtubules to pass through the neck and interact with the bud cortex. In cells lacking functional dynein or dynactin, spindle movement into the neck can be delayed (Eshel et al., 1993; J The online version of this article contains supplemental material. Address correspondence to Neil Adames, Department of Cell Biology and Physiology, Washington University School of Medicine, 660 South Euclid Ave., St. Louis, MO 63110. Tel.: (314) 362-4606. Fax: (314) 362- 0098. E-mail: nadames@cellbio.wustl.edu tants in which some cells inappropriately exit mitosis even though the spindle is within the mother. In some of these cells, the spindle pole body did not interact with the bud or the neck before mitotic exit. Thus, some alternative mechanism must exist to coordinate mitotic exit with spindle position. In both wild-type and mutant cells, mitotic exit was preceded by loss of cytoplasmic microtubules from the neck. Thus, the spindle position checkpoint may monitor such interactions. Key words: checkpoint • mitosis • dynein • EBI • Saccharomyces cerevisiae Li et al., 1993; Yeh et al., 1995). When this happens, cyto- plasmic microtubules passing through the neck continue to probe the bud cortex and grow longer, often pushing the spindle further from the neck (Adames and Cooper, 2000). When spindle movement into the neck is delayed, cytokinesis and mitotic exit is also delayed (Yeh et al., 1995). Inappropriate mitotic exit, defined here as mitotic exit when the spindle is not positioned in the neck, is ex- tremely rare. Therefore, a checkpoint mechanism appears to monitor spindle position and prevents mitotic exit. Pre- viously, we found that some mutations caused a failure of the tight coupling between spindle position and mitotic exit (Muhua et al., 1998). Recent work has shed light on how the spindle position checkpoint might work. Exit from mitosis is controlled by the small G protein Tem1p, which is an essential pro- tein that activates proteins of the mitotic exit network (Shirayama et al., 1994; Shou et al., 1999). The major func- tion of the mitotic exit network is to promote degradation of the mitotic cyclins and to inhibit the mitotic cyclin–cdk complex (Visintin et al., 1998; Shou et al., 1999). Tem1p is located at the spindle pole body (SPB)1 destined for the bud (Bardin et al., 2000; Bloecher et al., 2000). Lte1p has homology to guanine–nucleotide exchange factors, is pre- sumed to activate Tem1p, and is located in the bud (Bar- din et al., 2000; Pereira et al., 2000). Thus Tem1p and 1Abbreviations used in this paper: SEP, standard error of the proportion; SPB, spindle pole body. ª The Rockefeller University Press, 0021-9525/2001/04/159/10 $5.00 The Journal of Cell Biology, Volume 153, Number 1, April 2, 2001 159–168 http://www.jcb.org/cgi/content/full/153/1/159 159