The role of the proteins Kar9 and Myo2 in orienting the mitotic spindle of budding yeast.
BACKGROUND
Two genetic 'pathways' contribute to the fidelity of nuclear segregation during the process of budding in the yeast Saccharomyces cerevisiae. An early pathway, involving Kar9p and other proteins, orients the mitotic spindle along the mother-bud axis. Upon the onset of anaphase, cytoplasmic dynein provides the motive force for nuclear movement into the bud. Loss of either pathway results in nuclear-migration defects; loss of both is lethal. Here, to visualize the functional steps leading to correct spindle orientation along the mother-bud axis, we imaged live yeast cells expressing Kar9p and dynein as green fluorescent protein fusions.


RESULTS
Transport of Kar9p into the bud was found to require the myosin Myo2p. Kar9p interacted with microtubules through the microtubule-binding protein Bim1p and facilitated microtubule penetration into the bud. Once microtubules entered the bud, Kar9p provided a platform for microtubule capture at the bud cortex. Kar9p was also observed at sites of microtubule shortening in the bud, suggesting that Kar9p couples microtubule shortening to nuclear migration.


CONCLUSIONS
Thus, Kar9p provides a key link between the actin cytoskeleton and microtubules early in the cell cycle. A cooperative mechanism between Kar9p and Myo2p facilitates the pre-anaphase orientation of the spindle. Later, Kar9p couples microtubule disassembly with nuclear migration.
Beach
D L
DL
Department of Biology, University of North Carolina, Chapel Hill, North Carolina 27599-3280, USA.
Thibodeaux
J
J
Maddox
P
P
Yeh
E
E
Bloom
K
K
D000199:Actins / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D016764:Cell Polarity; D003593:Cytoplasm / Q000378:metabolism; D004268:DNA-Binding Proteins; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008870:Microtubules / Q000378:metabolism; D018995:Myosin Heavy Chains; D024462:Myosin Type II; D024701:Myosin Type V; D009218:Myosins / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D012331:RNA, Fungal / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000502:physiology*; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; D008941:Spindle Apparatus / Q000502:physiology*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
C098733:ASH1 protein, S cerevisiae; D000199:Actins; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; C110443:KAR9 protein, S cerevisiae; D008164:Luminescent Proteins; C077512:MYO2 protein, S cerevisiae; C434211:MYO2 protein, S pombe; D009687:Nuclear Proteins; D012331:RNA, Fungal; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; D014157:Transcription Factors; D049452:Green Fluorescent Proteins; D024462:Myosin Type II; D024701:Myosin Type V; D018995:Myosin Heavy Chains; D009218:Myosins