Suppressors of the ndc10-2 mutation: A role for the ubiquitin system in Saccharomyces cerevisiae kinetochore function

Abstract

We have isolated a new conditional-lethal mutation, ndc10-2, in the NDC10/CBF2/CTF14 gene that encodes the 110-kD subunit of the Saccharomyces cerevisiase CBF3 kinetochore complex. At the restrictive temperature of 37°, ndc10-2 cells are able to assemble anaphase spindles, but fail to segregate their DNA, consistent with a defect in kinetochore function. To identify other factors that play a role in kinetochore assembly or function, we isolated both dosage and second site suppressors of the ndc10-2 mutation. These screens identified UBC6 as a dosage suppressor, and mutation in UBC6 and UBC7 as second-site suppressors of ndc10-2 heat sensitivity. Both UBC6 and UBC7 encode ubiquitin-conjugating enzymes that function in ubiquitin- mediated protein degradation. Furthermore, overexpression of a mutant ubiquitin suppresses the ndc10-2 mutation. These results implicate the ubiquitin system in the regulation of ndc10-2 function and suggest a role for the ubiquin system in kinetochore function.