| Mitochondrial and nuclear segregation in S. cerevisiae require the Mdm1p intermediate filament-like cytoskeleton. MDM1 was mutagenized to identify mdm1 alleles defective in organelle inheritance but competent for Mdm1p assembly. Complex intragenic interactions among mutations defining two regions of Mdm1p (each responsible for mitochondrial inheritance and nuclear segregation) suggest Mdm1p forms complex assemblies in vivo. These structures must assemble such that the function of one molecule can be influenced by neighboring molecules, and must present surfaces with which many cellular machineries interact. mdm1-252, an allele defective for mitochondrial inheritance, and mdm1-217, an allele defective for nuclear segregation, were used in for suppressor analysis to identify proteins that interact with Mdm1p to mediate mitochondrial inheritance and nuclear segregation.;The smm1 and smm2 mutations in Rsp5p and Bul1p, respectively, suppress mdm1-252. smm1 causes mitochondrial inheritance defects, and overexpression of mitochondrial inheritance. In support, Rsp5p binds to Mdm1p in-vitro/. Bul1p interacts with Rsp5p (Yashiroda et al., 1996), a ubiquitin-protein ligase (Huibregetse et al., 1995), suggesting ubiquitination is required for mitochondrial inheritance. Rsp5p ubiquitin ligase activity is required for mitochondrial inheritance. Furthermore, ubiquitin overexpression suppresses smm1, and overexpression of the ubiquitin K63R mutation perturbs wild type mitochondrial inheritance, suggesting Rsp5p-dependent lysine 63 linked ubiqitination is required for mitochondrial inheritance.;Mdm1p is also required for mitotic spindle orientation. If S-phase synchronized cells are shifted to non-permissive temperatures, spindles become rapidly misaligned in Class IIIa but not Class IIIb mdm1 cells. If cells are released from G1 block at 37... |
