Functional Characterisation of Spc29p, a Component of the Saccharomyces cerevisiae Spindle Pole Body

Elliott, Sarah L (2001) Functional Characterisation of Spc29p, a Component of the Saccharomyces cerevisiae Spindle Pole Body. PhD thesis, University of Glasgow.

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In the budding yeast Saccharomyces cerevisiae, microtubule organising functions are carried out by the spindle pole body (SPB). The SPB is a disk-like, multilayered structure that is embedded in the nuclear membrane throughout the cell cycle. The major substructures of the SPB are the outer, central and inner plaques and the half-bridge. The central plaque is embedded in the nuclear membrane. The outer and inner plaques are positioned on the cytoplasmic and nuclear side of the central plaque where they organise the cytoplasmic and nuclear microtubules respectively. The half-bridge extends from the central plaque along the nuclear envelope and is involved in SPB duplication and organises cytoplasmic microtubules during G1 of the cell cycle and during mating. Described within is the identification and characterisation of a novel SPB component, Spc29p. Spc29p was found as a component of a salt stable SPB subcomplex composed of central and inner plaque SPB components. This complex contained Spc29p, the central plaque component Spc42p, the ?-tubulin-complex-binding protein Spe110p and calmodulin. Spc29p acts as a linker between Spc42p and the globular C-terminus of Spc110p at the central plaque. Evidence is provided that the calmodulin-binding site of Spc110p influences the binding of Spc29p to Spc110p. In addition, co-overexpression of Spc29p, Spc42p and Spc110p resulted in a three fold increase in the diameter of the SPB, indicating that expression levels of SPB components may be involved in the size control of the SPB. Construction and analysis of spc29 temperature sensitive mutants uncovered another important function of Spc29p. Using immunofluorescence, FACS analysis and electron microscopy the spc29(ts) mutants were found to have an SPB duplication defect. A role for Spc29p in SPB duplication was supported by the genetic and biochemical interactions between Spc29p and other SPB components involved in SPB duplication, namely the half-bridge components Cdc3 1p and Karlp. Evidence is also provided for an interaction between Spc29p at the periphery of the central plaque, with the Bbplp/Mps2p complex, which is involved in embedding the SPB in the nuclear envelope. Spc29p was also found to be phosphorylated in a cell cycle dependent manner.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Additional Information: Adviser: Elmar Schiebel
Keywords: Molecular biology, Microbiology
Date of Award: 2001
Depositing User: Enlighten Team
Unique ID: glathesis:2001-76025
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 19 Dec 2019 09:15
Last Modified: 19 Dec 2019 09:15

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