High impact information on SPC110

• Microtubule organization requires the yeast gamma-tubulin complex (Tub4p complex) which binds to the nuclear side of the SPB at the N-terminal domain of Spc110p [1].
• Finally, we isolated a complex containing Spc110p, Spc42p, calmodulin and a 35 kDa protein, suggesting that these four proteins interact in the SPB [2].
• We created temperature-sensitive SPC110 mutations by combining PCR mutagenesis with a plasmid shuffle strategy [3].
• To confirm this, we examined the phenotype of ts alleles of SPC110, one of which contains a mutation in the calmodulin binding site and was suppressed by overexpression of calmodulin [4].
• This suggests that the essential function of Spc110p is to connect the nuclear microtubules to the SPB [4].

Biological context of SPC110

• Deletion of HCM1 does not affect the basal level of SPC110 transcription, but reduces the induction that occurs late in G1 of the cell cycle [5].
• The promoter of SPC110 contains a match to the consensus binding site [5].
• Nuf1p/Spc110p, one of the essential targets, is a spindle pole body component that is required for proper mitosis [6].
• Additionally, these mutants displayed characteristic mitotic defects: an increased ratio of artificial chromosome loss, which could be suppressed by the CaM-independent dominant allele of NUF1, and aberrant microtubule structures [6].
• However, the interaction was restored in a heterozygous diploid of the two cmd1 alleles, suggesting that intragenic complementation between these cmd1 alleles occurs by a novel mechanism, whereby co-presence of both mutant proteins rescues the interaction with Nuf1p [6].

Anatomical context of SPC110

• The amino terminus of Spc110p faces the inner plaque, the substructure from which spindle microtubules radiate [7].
• Spc110p is an essential structural component of the SPB and spans between the central and inner plaques of this multilamellar organelle [7].
• The yeast spindle pole body (SPB) component Spc110p (Nuf1p) undergoes specific serine/threonine phosphorylation as the mitotic spindle apparatus forms, and this phosphorylation persists until cells enter anaphase [8].
• In budding yeast, this attachment is provided by the coiled-coil protein Spc110p, which links the yeast gamma-tubulin complex to the core of the yeast centrosome [9].

Associations of SPC110 with chemical compounds

• Ser(60), Thr(64), and Thr(68) are the major sites in Spc110p phosphorylated by Mps1p in vitro, and alanine substitution at these sites abolishes the mitosis-specific isoform in vivo [8].

Physical interactions of SPC110

• Immunoprecipitation analysis showed that two cmd1 mutations belonging to two distinct intragenic complementation groups had the most severely impaired complex formation with Nuf1p at the restrictive temperature [6].
• Cdc31p was also able to bind to the carboxy terminus of Nuflp/Spc110p, another component of the SPB (Kilmartin, J. V., S. L. Dyos, D. Kershaw, and J. T. Finch. 1993. J. Cell Biol. 123:1175-1184) [10].
• We tested whether binding of the recombinant Tub4p complex to the spindle pole body docking protein Spc110p affects its nucleating activity [11].

Regulatory relationships of SPC110

• The temperature-sensitive growth of these cmd1 mutants was suppressed by a CaM-independent dominant allele of NUF1 [6].

Other interactions of SPC110

• Spc29p is a component of the Spc110p subcomplex and is essential for spindle pole body duplication [12].
• Together, these studies provide evidence that Spc110p directly links the Tub4p complex to the SPB [7].
• In contrast, the spc97 alleles are synthetic lethal with spc110 alleles that encode mutations in either the N terminus or the C terminus [7].
• We also found that mutations in SPC110, which encodes a major SPB component, showed genetic interactions with both CDC31 and the PKC1 pathway [13].
• In support of the model that the PKC1 pathway regulates SPB duplication, one of the phosphorylated forms of Spc110p was absent in pkc1 and mpk1Delta mutants [13].

Analytical, diagnostic and therapeutic context of SPC110

• Using the two-hybrid assay, we show that the interactions of Spc110p with Spc97p and Spc98p are not equivalent [7].
• In addition, immunofluorescence results indicate that mammalian cells contain a NUF1-related nuclear protein [14].
• Sequence analysis of the NUF1 gene predicts a protein 945 amino acids in length that contains three domains: a large 627 residue central domain predicted to form a coiled-coil structure flanked by nonhelical amino-terminal and carboxy-terminal regions [14].
• Antibodies to NUF1 fusion proteins were generated, and affinity-purified antibodies were used for immunoblot analysis and indirect immunofluorescence localization [14].
• Asynchronous wild-type cultures contain two electrophoretically distinct isoforms of Spc110p as detected by Western blot analysis, suggesting that Spc110p is modified in vivo [15].

References