期刊
CELL CYCLE
卷 8, 期 19, 页码 3182-3190出版社
TAYLOR & FRANCIS INC
DOI: 10.4161/cc.8.19.9731
关键词
cytoplasmic dynein; cortical dynein; spindle orientation; PH domain; CAAX motif; PIP2
类别
资金
- University of Massachusetts Amherst Biology Department HHMI
- NIH/NIGMS [1R01GM076094]
During mitosis in budding yeast, cortically anchored dynein exerts pulling forces on cytoplasmic microtubules, moving the mitotic spindle into the mother-bud neck. Anchoring of dynein requires the cortical patch protein Num1, which is hypothesized to interact with PI(4,5)P-2 via its C-terminal pleckstrin homology (PH) domain. Here we show that the PH domain and PI(4,5)P-2 are required for the cortical localization of Num1, but are not sufficient to mediate the cortical assembly of Num1 patches. A GFP fusion to the PH domain localizes to the cortex in foci containing -2 molecules, whereas patches of full-length Num1-GFP contain similar to 14 molecules. A membrane targeting sequence containing the CAAX motif from the yeast Ras2 protein can compensate for the PH domain to target Num1 to the plasma membrane as discrete patches. The CAAX-targeted Num1 exhibits overlapping but largely distinct localization from wild-type Num1. However, it is fully functional in the dynein pathway. More importantly, cortical PI(4,5)P-2 is dispensable for the localization and function of the CAAX-targeted Num1. Together, these results demonstrate that cortical assembly of Num1 into functional dynein-anchoring patches is independent of its interaction with PI(4,5)P-2.
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