Supplemental Material

This article contains the following supporting material:

• Materials and Methods
• Table 1
• Figure 1 - The 18Xmyc-tagged Dia2 allele is functional. A) Dia2-18Xmyc rescues the cold-sensitive and HU-sensitive defects of the dia2 allele. Serial dilutions of the indicated diploid strains were spotted on rich media containing the indicated amounts of HU and were incubated at 25°C. B) Expression of the 18Xmyc-tagged Dia2 protein. Whole cell extracts of Y80 and the Dia2-18Xmyc strain were subjected to immunoblotting using anti-Myc antibodies. The 18Xmyc tagged Dia2 protein exists as two major forms. Anti-Pgk1 antibodies were used as a loading control. C) The Dia2 F-box deletion mutant does not bind Skp1. Skp1 was immunoprecipitated as described in the Materials and Methods section from cells expressing either wildtype Dia2 or the ƒ´F-box mutant tagged with 18 myc epitopes.

• Figure 2 - Overexpression of CLB5 and CLB6 is toxic todia2 cells. WT and dia2 cells transformed with galactose-inducible vectors expressing CLB4, CLB5 and CLB6 were grown on selective media containing galactose at 30°C. All strains grew on selective media containing glucose (data not shown).

• Figure 3 - Skp1 associates with ARS603 in nocodazole-arrested cells. Chromatin immunoprecipitation was performed as described in Materials and Methods.