Phosphorylation of Tyr19 in Cdc28 by Swe1 protein kinase
negatively regulates B-cyclin-bound forms of Cdc28. Swe1 itself is
negatively regulated by a protein kinase, Hsl1, and a putative
protein-arginine methyltransferase, Hsl7. Hsl1, Hsl7 and Swe1 localize
to the bud neck in a septin-dependent manner at bud
emergence. Moreover, Hsl1 and Hsl7 are required for Swe1 modification
and degradation, an event necessary for timely entry into
mitosis. Hsl7 physically interacts with both Hsl1 and Swe1 [see
Shulewitz et al. (1999) MCB 19: 7123-7137]. These data suggest that
Hsl1 and Hsl7 couple modulation of Cdc28 activity to morphogenetic
events occurring during early stages of budding. To further dissect
functional interactions in this regulatory complex, we mapped a region
of Hsl7 essential for its binding to Hsl1 using a variation of the
reverse two-hybrid method. We also delineated a region of Hsl1
required for its attachment to the septins. Using GFP fusions in live
cells, and indirect immunofluorescence and immuno-electron microscopy
in fixed cells, we have found that, late in telophase and in early G1,
Hsl7 disappears from the bud neck and relocalizes to the cytoplasmic
face of the spindle pole body (SPB). At the time of bud emergence,
Hsl7 shifts from the SPB to the bud neck in an Hsl1-dependent
fashion. The physiological relevance of the dual location of Hsl7 and
the molecular mechanism by which Hsl7 action contributes to Swe1
inactivation are under active study.
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